Highlights Archive

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October 11, 2017

Review : Lignocellulose deconstruction in the biosphere
The aims of this review were to provide a better understanding of carbon recycling in the biosphere ranging from biomass synthesis to deconstruction by biotic or abiotic routes; highlight the importance of the synergism between natural microbial communities within a given ecosystem; and show the complexity of the biomass deconstruction mechanisms that have evolved in nature.

October 09, 2017

Linking Physiochemical Properties of Populus to Recalcitrance in Natural Variants
This study integrated cell wall structural components with plant genetics and presented associations between physicochemical properties, sugar release and differential gene expression. Lignin content negatively affected the sugar release of Populus, whereas lignin S/G ratio, lignin Mw, cellulose content, and cellulose accessibility were positively correlated with the conversion performance. The trend of differential gene expression in variants supports the characterization results and their effects on biomass conversion.

August 29, 2017

Schematic and TEM micrograph of the cellulose deconstruction mechanism of CelA. The GH48 and GH9 cat (...more)

Hyper Thermophilic Cellulase CelA from C. bescii Indifferent to Level of Crystallinity in Biomass
CelA was shown to break down cellulose with high crystallinity to the same extent as low crystallinity cellulose — a characteristic never reported for other cellulases. CelA exhibits a cavity forming mechanism on all types of biomass tested. This discovery demonstrates that nature's strategies for biomass conversion are diverse and that enzymes with new characteristics can still be discovered and isolated for improving biomass deconstruction applications.

August 28, 2017

Net ethanol and acetate production from the wild type C. thermocellum strain (Cth_WT) and a m (...more)

Enchi Corporation Licenses Joint Dartmouth College and ORNL Technology
"Engineered microbe could potentially move the payback period for a commercial cellulosic ethanol operation from 15 years down to two years," Bill Brady, Enchi CEO.

August 01, 2017

Society for Industrial Microbiology and Biotechnology (SIMB) Awardees in BESC–2017
Brian H. Davison, Chief Scientist for Systems Biology and Biotechnology in the Bioscience Division at ORNL and the Science Coordinator in BESC, awarded SIMB Fellow 2017. Fellowship status is a special grade of membership in SIMB, acknowledging a career-long sustained record of significant research and/or service contributions to the profession of industrial microbiology and/or biotechnology.

Amanda Williams-Rhaesa, a fifth-year Ph.D. candidate in Biochemistry and Molecular Biology at the University of Georgia (Dr. Michael Adams' lab), received a 2017 SIMB Diversity Travel Award.

July 18, 2017

BESC glycosyl transferase work featured on cover

Mechanism of Fucosylation in Xyloglucan Biosynthesis revealed in Arabidopsis thaliana
FUT1 is a plant fucosyltransferase that carries out the final step in the synthesis of the hemicellulose xyloglucan, which is a major component of the plant cell wall and is involved in cell growth and expansion, energy metabolism, and signaling. X-ray crystallography revealed the structural architecture of FUT1 in complex with bound donor and acceptor substrate analogs, forming the basis for in silico studies that unraveled the mechanistic basis for fucosylation in GT37 enzymes. Structural and mutagenic analyses determined that there was no active site amino acid residue that could act as a base for catalyzing the reaction. In silico studies revealed the presence of a crucial water molecule that could conduct catalysis by shuttling the proton to enable a SN1-like mechanism.

July 14, 2017

Comprehensive Assessment of Glycosyl Residue Composition Analysis Methods
This is the first paper that provides side-by-side comparison of the efficacy of four different established methods in the analysis of glycosyl residue composition of cell walls from both primary wall-enriched leaf tissues and secondary wall-enriched wood/stem tissues of both dicot (Arabidopsis, Populus) and grass (rice, switchgrass) species.

July 11, 2017

2017 BESC Science Retreat
The Year 10 BESC Science Retreat was in Chattanooga, Tennessee from July 11-13, 2017 with 165 attendees from 13 BESC partner institutes.

July 05, 2017

adhA, nfnAB, and adhEG544D expression in C. thermocellum

Engineering T. saccharolyticum Ethanol Pathway into C. thermocellum Improves Biofuel Production
Clostridium thermocellum is a candidate for ethanol production from cellulose, but requires metabolic engineering to improve yield and titer. Thermoanaerobacterium saccharolyticum was previously engineered to produce ethanol at high yield and titer. Four of the genes that are responsible for ethanol production are adhA, nfnA/B, and adhEG544D. Introducing the T. saccharolyticum genes into C. thermocellum significantly increased ethanol yield.

July 03, 2017

Linking Overflow Metabolism and Growth Cessation in C. thermocellum
Findings of growth cessation and overflow metabolism provide metabolic engineering and fermentation strategies for high substrate loading fermentations to achieve high product yields.

June 19, 2017

Improving Bioavailability of Carbohydrate to C. bescii in Natural and Transgenic Switchgrasses
The bioavailability of carbohydrates to C. bescii was evaluated in the switchgrass lines unmodified Cave-in-Rock (CR), transgenic COMT-KD and Myb4-OE, and their corresponding parental lines. Each switchgrass line was examined as either hydrothermally treated (180ºC for 25 min) or untreated. C. bescii growth and carbohydrate solubilization (cellulose and hemicellulose) were determined for each treatment scheme (microbial, hydrothermal, or a combination of both). While C. bescii could significantly solubilize the transgenic switchgrass lines, including Cave-in-Rock, hydrothermal treatment was needed to solubilize 50% or more of the carbohydrate content.

May 01, 2017

Microbial hydrolysis changes poplar cell wall surfaces. Cellulose (yellow) decreases and lignin (blue or gray) becomes more prominent.

New Insights On Recalcitrance From Lignin Inhibition Of Microbial Deconstruction
Microbial solubilization of biomass selectively targets carbohydrates. Clostridium thermocellum, a highly effective cellulolytic microbe, can hydrolyze ~93% of 100gL-1 pure cellulose. The microbe solubilizes only partially hydrolyzes cellulose in untreated energy crops. Techniques used were quantitative fluorescence colocalization analysis and surface chemical imaging. Quantitative evidence for the cause of feedstock recalcitrance: depletion of surface carbohydrate increases lignin exposure which leads to inhibition of enzymatic activity, while the bulk residual biomass retains significant undigested sugar content. Limited hydrolysis is not caused by inhibitory hydrolyzed sugars nor fermentation products (i.e., ethanol).

May 01, 2017

Jonathan Mielenz, retired researcher from ORNL and BESC, was awarded the 2017 Charles D. Scott award (...more)

BRCs well represented at 39th Symposium on Biotechnology for Fuels and Chemicals
As part of the overall symposium the three DOE BRCs presented a combined total of twenty-two talks, including two as collaborations between the BRCs, and thirty-nine posters representing 28% of the talks and 15% of the posters. The presentations and posters highlighted the accomplishments, as well as common challenges in organization and research, in carrying out impactful mission-directed R&D in bioenergy.

April 17, 2017

Photos of the field experiments in the first (2013), second (2014), and third (2015) growing seasons (...more)

Multi-Year Study Of Transgenic Switchgrass Plants In Regulation Shows Improved Yield And Dependency On Expression Levels
Four promising lines with moderate or low microRNA (miR156) overexpression from prior greenhouse study were selected for a field experiment to assess miR156 expression levels and biomass yields over three years. The best performing line consistently produced more biomass (25-56%) than the control in all three seasons, which translated to the production of 30% more biofuel per plant in the final season. Transgenic plants with modifications to transcriptional regulators, which impact such a broad range of genes, will likely require more extensive field evaluations than those with single and simple trait gene modifications to evaluate the stability of the desired growth phenotypes.

April 12, 2017

CBP and Cotreatment

First-Of-A-Kind Study Of Thermophilic CBP With Cotreatment
Milling during lignocellulosic fermentation (or cotreatment) is an alternative to thermochemical pretreatment to enhance biological solubilization of lignocellulose. Carbohydrate solubilization was measured in consolidated bioprocessing with Clostridium thermocellum with cotreatment or with hydrothermal pretreatment. High carbohydrate solubilization was demonstrated without thermochemical pretreatment and added saccharolytic enzymes. C. thermocellum appears able to attack all the major linkages in cellulosic biomass provided that these linkages are accessible. The ability of C. thermocellum to withstand high intensity milling supports the feasibility of cotreatment. Less modified lignin may foster production of value-added coproducts.

March 27, 2017

Use of Caldicellulosiruptor saccharolyticus as a biological probe to report on changes in recalcitrance of plant biomass
C. saccharolyticus whole genome microarrays were used to access the transcriptome when grown on purified polysaccharides and chemically pretreated or genetically modified lignocellulosic substrates. Differential regulation of carbohydrate degradation, transport, and metabolism genes in C. saccharolyticus can be used to assess the availability of various plant polysaccharides to the microbe. This allows for inferences about the consequences of a chemical pretreatment or genetic modification of lignocellulose with an eye towards biofuels production.

March 21, 2017

Operating cost and revenue comparison

CBP-CT: an R&D-Driven Innovation with Potential for Disruptive Reductions in the Cost of Cellulosic Biofuels
Compared to current technology, the CBT-CT scenario was projected to offer 4.6-fold greater annual revenue, as well as 8-fold shorter payback period, and feasibility at ~10-fold smaller scale. CBT-CT is a potentially disruptive technology and a powerful, though still speculative, example of the potential benefits of including new-paradigm as well as in-paradigm innovation in R&D portfolios.

March 21, 2017

Improving Pretreatment Inhibitor Tolerance in Clostridium thermocellum
Bioconversion performance in the presence of common pretreatment inhibitors was improved through targeted strain engineering thus reducing barriers to achieving a robust consolidated bioprocessing strategy with Clostridium thermocellum.

March 18, 2017

Structure prediction and molecular recognition feature analysis of PdDUF266A.

Overexpression of DUF266 can significantly enhance both cellulose and biomass in Populus
(BESC's 900th publication)

Little is known about the function of Domain of Unknown Function 266 (DUF266)-containing proteins and no DUF266 protein has been characterized in Populus. A bioinformatics approach was taken to identify all DUF266 proteins in Populus and their relationship with their homologs in other species. A transgenic approach was used to determine the function of one member of DUF266 proteins in Populus (PdDUF266A). The resulting phenotypes make PdDUF266A a promising target for genetic manipulation for improving biomass quality.

March 10, 2017

Increasing delignification shows changes in morphology, elasticity, plasticity and adhesion

Use of Photoacoustic AFM to Spatially Resolve Nanomechanics of Plant Cell Wall Delignification
By chemically processing biomass and employing emerging nanometrology techniques, the various stages of lignin removal may be distinguished through the observation of morphochemical and nanomechanical variations. The cell wall nanomechanical properties undergo quantifiable reductions in plasticity, adhesion energy, and elasticity. These quantitative observations can be used to characterize delignification. The observed reduction in plasticity seems counterintuitive considering that lignin adds to cell wall rigidity.

February 27, 2017

Microbial Attachment to Cellulose Substrate Produces Widespread Gene Expression Changes
Unlike the majority of biofilm forming bacteria, C. thermocellum adheres to substrata that also provide its major carbon and energy sources. Before this study, discrete omics analyses of biofilm and planktonic cell populations had not been performed. Cellulolytic bacterial cells that evolved to thrive on solid carbon sources were shown to thoroughly alter expression of their central metabolism, anabolism, and homeostatic functions in response to the availability of solid attachment interfaces and solubilizable carbohydrates. The attached bacteria were functionally strong in growth and biomass conversion, while the planktonic cells were stressed and motile due to low available substrates.

February 07, 2017

Growth rate of C. thermocellum in response to various C5 sugars

Understanding the role of pentose sugars on C. thermocellum metabolism
This work highlights the importance of removing pentose sugars during bioconversion to biofuels, which could be achieved with C5 utilizing co-cultures or engineered strains of C. thermocellum, and is the first report of a functional ArgD-type cell signaling system in a thermophilic Firmicute and further work is warranted to understand the role of cell-to-cell signaling in achieving robust fermentations of lignocellulosic biomass.

January 20, 2017

SEM image of C. bescii (blue circles) attached to cellulose walls at 48 h

Visualization of Significant Alteration of Populus biomass by C. bescii
Caldicellulosiruptor bescii targets surface cellulose and hemicellulose, but increase of surface lignin could inhibit further sugar access. Biphasic cell growth is attributed to degradation of easily accessible sugars followed by utilization of more insoluble polysaccharides.

January 10, 2017

Metabolic Adaptation Of C. thermocellum To Growth Inhibitors Released During Deconstruction Of Switchgrass
Integrated omics data of microbial growth on complex lignocellulosic biomass over time provided a detailed view of the molecular machinery (metabolites and enzymes) that reveals temporal adaptation to a complex, lignocellulose substrate — information that is critical for engineering C. thermocellum's industrial efficacy.

January 03, 2017

Engineering N-terminal End of CelA Enhances the Cellulolytic Activity of Caldicellulosiruptor bescii
To test whether alteration of the N-terminal terminus of CelA GH9 and CelA GH48 domains might improve secretion and/or catalytic efficiency of CelA, repeating aspartate tags were introduced into the N-terminal ends of these 8 domains. Introduction of repeating aspartate tags resulted in an increase in the general activity of the exoproteome and a dramatic increase in growth of C. bescii on crystalline cellulose. Most efficient cellulase systems contain highly active exocellulase enzymes capable of decrystallizing cellulose, so the observed increase in activity on Avicel bears directly upon the ability of CelA to degrade realistic biomass feedstocks destined for biofuels production.

December 22, 2016

RNA-Seq analysis for man5A (mannanase) region in ΔglyR2 mutant and parent strains

Identification of LacI Repressor-Based Regulatory Network Activity Targeting Hemicellulases in C. thermocellum
Understanding C. thermocellum gene regulation is of importance for improved fundamental knowledge of this industrially relevant bacterium. We combined use of three lacI gene deletions with transcriptomics and DNA binding assays to gain insights into LacI regulatory networks. The identification of LacI repressor activity for hemicellulase gene expression is a key result of this work and will add to the small body of existing literature on the area of gene regulation in C. thermocellum.

December 22, 2016

Correlation of sugar release with lignin content

Key Traits Leading to Reduced Recalcitrance Remain Stable Following Three Years of Field Trials
Down-regulation of the caffeic acid O-methyltransferase (COMT) gene in the lignin biosynthesis pathway of switchgrass has been previously shown by BESC to improve the thermochemical and biochemical conversion of biomass. The originally reported improvements in biomass cellulose accessibility wrought by down-regulation of COMT in switchgrass have remained stable following three years of field trials. Demonstrating stability of introduced traits for improved biofuel production in bioenergy crops is an important proof-of-principle on the path toward long-term validation of such approaches.

December 12, 2016

A representative HSQC NMR spectra of natural poplar variant lignin

Use Of Populus Natural Variants As A Research Tool To Further Determine Molecular Basis Of Recalcitrance
This project investigates the relative contributions of various factors to recalcitrance by a first-order multi-variants linear correlation analysis, which significantly helps improve the understanding of the fundamental mechanisms of biomass recalcitrance.

December 06, 2016

Metabolic network

Confirmation of Role for Malate Shunt in Clostridium thermocellum Glycolysis
The unique metabolism of C. thermocellum has presented difficulties in engineering it for improved ethanol production. Improved understanding of the malate shunt will allow the design of strains for improved ethanol production.

November 29, 2016

In situ 3D SRS imaging to track xylan distribution in deacetylated corn stover cell wall before and after xylan digestion

New Approach Allows Resolution Of Xylan Polymers In Plant Cell Wall
This approach demonstrated that xylan, an important plant polymer source, can be resolved from cellulose and lignin in situ using enzymatic digestion and label-free SRS microscopy. This can be used to enhance understanding of xylan in cell wall biosynthesis and deconstruction.

November 25, 2016

Comparison of biomass solubilization by C. thermocellum relative to fungal cellulases. The da (...more)

State of Play in Ethanol Production using Consolidated Bioprocessing with C. thermocellum and T. saccharolyticum
A comprehensive book chapter that addresses Consolidated Bioprocessing (CBP) organism development strategies; plant cell wall solubilization by C. thermocellum; bioenergetics of C. thermocellum cellulose fermentation; and metabolic engineering and the current state of strain development.

November 21, 2016

Simplified structure of plant cell walls (A), lignin isolated from poplar (B), and schematic structure of poplar lignin (C)

Mini Review Surveys Improvements In Understanding Relationship Of Lignin Structure To Biomass Recalcitrance
This review focuses on advances in understanding the specific roles of lignin properties during pre-treatment; lignin-enzyme interactions; monolignol compositional units; and hydroxycinnamates and hydroxyl and carboxylic groups in lignin.

November 03, 2016

Comparisons of five transgenic switchgrass lines to their parental controls (* indicates not a stati (...more)

Comparative Analysis Of Field-Grown Transgenic Switchgrass Lines Shows Stable Increases In Biofuel Yields
Clones of plants representing independent transgenic events and their respective non-transgenic control lines were investigated for biomass yield, carbohydrate composition, and recalcitrance to bioconversion via separate hydrolysis and fermentation to ethanol. Over two consecutive field-growth seasons, most transgenic lines maintain higher glucan and xylan yield at similar, or sometimes better, plant biomass. Transgenic lines targeting cell wall modifications yielded significantly higher bioconversion to ethanol (up to 36% and 21% in year 1 and year 2, respectively).

November 02, 2016

Mean soil organic matter in the upper 0-15 cm (A) and deeper 15-30 cm (B) during the first two growi (...more)

Field Grown Transgenic Switchgrass Has No Negative Affect On Soil Chemistry, Microbiology Or Carbon Storage Potential
BESC has demonstrated that when shoot S/G lignin ratio is decreased in switchgrass, greater yields of biofuel can be produced; understanding agronomic consequences of such changes is important to proving industrial value of engineered biofuel crops. COMT-altered switchgrass appears to be substantially equivalent to non-engineered switchgrass with regards to soil and microbiome properties.

November 01, 2016

Heat map of C. thermocellum transcriptomic response to exogenous acetate. Blue and yellow ind (...more)

Improving Robustness of Engineered C. thermocellum
Understanding and overcoming low robustness in engineered microorganisms will be essential to industrial deployment of these organisms. This study helps reveal mechanisms of electron balancing in C. thermocellum that leads to increased robustness in its potential use in biofuel production.

November 01, 2016

Diagram of electron metabolism in C. thermocellum

Engineering Electron Metabolism to Increase Biofuel Production in Clostridium thermocellum
Engineering electron metabolism is a promising strategy for improving ethanol production in C. thermocellum. A strain with improved ethanol production was also generated by overexpressing the rnfCDGEAB operon.

October 28, 2016

Schematic of reductive C1 metabolism initiated from reversed PFOR and PFL. Carbons in red indicate < (...more)

C. thermocellum Endowed with Ability to Fix CO2
A novel route that C. thermocellum employs to fix CO2 when grown in primarily heterotrophic mode supplemented with sodium bicarbonate was discovered. Critical enzymes responsible for fixing CO2 and channeling the fixed carbon to the C1 metabolic pathway were identified. This research paves the way to future engineering of the Clostridium thermocellum bacterium to utilize cellulose and CO2 simultaneously as a means to improve microbial carbon efficiency and reduce CO2 in the environment.

October 19, 2016

National Bioenergy Day
BESC participated with seven different agencies to provide educational and informative biofuel connected activities and displays for ~200 elementary children during the National Bioenergy Day celebration held at the University of Tennessee (UT) Arboretum in Oak Ridge, Tennessee.

September 30, 2016

GreenWood and ORNL Sign Agreement.
Seated: Brian Stanton, Chief Science Officer, GreenWood Resour (...more)

GreenWood Resources Licenses BESC Invention to Boost Biofuel Yield
GreenWood Resources has licensed a BESC technology based on the discovery of a gene in Populus trichocarpa that makes it easier to convert poplar trees into biofuels. They plan to commercialize the technology to select and breed better varieties of poplar with less lignin content, which simplifies the conversion process and ultimately lowers the overall costs of biofuel production.

September 15, 2016

New Thioacidolysis Method Allows for Improved Throughput in Linking Biomass Recalcitrance with Lignin Structure and Composition
Thioacidolysis is a method used to measure the relative content of coumaryl (H), syringyl (S) and coniferyl (G) alcohol lignin monomers bound by β-O-4 linkages. This new method was developed with significantly higher throughput than traditional techniques and was found to measure statistically similar S/G ratios in lignin from different lignocellulosic biomass types relative to a traditional low-throughput technique. The composition and structure of lignin based on the relative amount of monomers bound by β-O-4 linkages can be more rapidly determined for screening purposes allowing for improved throughput in linking biomass recalcitrance with lignin structure and composition.

September 06, 2016

GEM allows for reaction level investigation

Development Of A Genome-Scale Metabolic Model Of C. Thermocellum Assists In Strain Design For Biofuels And Chemicals
Genome scale modeling is a powerful tool for investigating cellular metabolism. A genome scale model (GEM) of DSM 1313 was constructed and curated for simulation of growth on alternative cellulosic substrates by implementing an adjustable cellulosome. Using the GEM, researchers were able to explore how key redox and bioenergetic reactions change as a function of growth rates and carbon sources in a systematic manner. Use of the model led to a proposed substrate-dependent regulatory mechanism by which ethanol production is enhanced or limited.

August 25, 2016

Heterologous Expression of Xylanases from A. cellulolyticus Enhances Hemicellulolytic Activity of C. bescii
To test whether the addition of A. cellulolyticus xylanases might enhance the hemicellulolytic activity of C. bescii and growth on xylan substrates, the xylanases were expressed in C. bescii using new expression vectors. Expression of the xylanases in C. bescii results in an increase in the activity of the exoproteome on xylan and a dramatic increase in its ability to grow on xylan substrates.

August 10, 2016

Microbial Upgrading of Lignin-Derived Carboxylates to Branched-Chain Esters for Drop-in Biofuels
An innovative biological conversion route was developed for conversion of lignocellulosic biomass or organic wastes into biofuels and high-value chemicals.

August 08, 2016

Cellulose in THF-Water

HPC Simulation of Co-Solvent Pretreatment of Biomass Confirmed by Experiment
Previous BESC work has shown that reaction of biomass in aqueous tetrahydrofuran (THF) in CELF (Co-solvent Enhanced Lignocellulosic Fractionation) pretreatment is highly effective at delignification and deconstruction of biomass for biofuels and bioproducts. Molecular dynamics simulations were performed of whole cellulose fibers and single cellulose chains in a THF-water co-solvent at various temperatures on the ORNL TITAN supercomputer. The simulations were complemented by THF-water experiments on cellulose, measuring its solubilization and imaging the product residue.

August 05, 2016

Creative Discovery Museum Receives Pinnacle Award for Distance Learning
The Creative Discovery Museum (CDM) has been awarded a Center for Interactive Learning and Collaboration (CILC) Pinnacle Award for 2015 – 2016 based on recipient evaluations. The BESC-sponsored CDM "Distance Learning Biofuels" program is a 50-minute lesson designed for students in grades 4 – 8. A museum educator from CDM provides the lesson via video conferencing. The program provides the opportunity for students to learn about innovative research taking place in the fields of biofuels and alternative energy.

July 27, 2016

One-hour hydrolysis rate versus cumulative cellulose conversion for uninterrupted and restarted hydr (...more)

Cellulose DP And Accessibility Contribute To Cellulase Enzyme Action On Biomass Substrate
The enzymatic hydrolysis of lignocellulosic biomass proceeds initially at a fast rate followed by a rapid decrease in the conversion rates. Degree of polymerization (DP) analysis during hydrolysis suggested a synergistic action of endo- and exo-glucanases that contribute to the occurrence of a peeling off mechanism. Cellulose accessibility analysis showed that limited accessible surface area of cellulose is probably not a major limiting factor that causes the decline of hydrolysis rate in its late stage. Restart hydrolysis experiment suggested that enzyme related factors such as enzyme inactivation or steric hindrance of enzymes should be responsible to the reduction in hydrolysis rate given the large size of cellulase enzymes.

June 21, 2016

Dr. Liao received his Bachelor degree at the National Taiwan University and his Doctorate at the University of Wisconsin-Madison

James Liao Appointed To Serve As Academia Sinica President
Dr. James Liao has been named as the 11th president of Academia Sinica — the top academic research institution in Taiwan. Academia Sinica supports research activities in a wide variety of disciplines, ranging from mathematical and physical sciences, to life sciences and to humanities and social sciences. Dr. Liao serves as the BESC Activity Lead for Butanol Production by CBP.

June 20, 2016

Isolation and characterization of M. truncatula mutant seeds lacking physical dormancy.

Discovery Of A Novel Molecular Mechanism Controlling Seed Physical Dormancy
Physical seed dormancy exists widely in higher plants, but its molecular mechanism has remained largely elusive. By screening a large number of Tnt1 retrotransposon-tagged Medicago truncatula lines, we identified non-dormant seed mutants from this model legume species. This study elucidated a novel molecular mechanism of physical dormancy and revealed a new role of class II KNOX genes.

June 20, 2016

Improved Thermophilic Anaerobe Achieves High Biofuel Titers From Wood And Cellulose
This study demonstrates that thermophilic anaerobes are capable of producing ethanol at high yield and at titers greater than 60 g/L from purified substrates. However,challenges were encountered with existing fungal cellulases and inhibitors from pretreatment. The high titers and yields observed support the feasibility of using engineered thermophiles for industrial ethanol production if challenges associated with pretreatment inhibitors can be avoided.

June 20, 2016

Relation between viscosity and shear rate of poplar lignin at 170 °C

Syringyl/Guaiacyl (S/G) Ratio In Lignin May Influence Quality Of Carbon Fiber Produced From Lignins
Current lignin-based carbon fiber using low-cost lignin precursors does not possess the mechanical properties required for many structural applications. Lignins with various syringyl/guaiacyl (S/G) ratios were isolated from Populus genotypes and subjected to rheological treatment at various temperatures and cooling conditions. Organosolv lignins extracted from genotypes with lower S/G ratios exhibited higher viscosity values and molecular weights with larger amounts of condensed structures following rheology treatment at relatively lower temperature; higher viscosity and molecular weight is known to provide for higher quality carbon fiber.

June 07, 2016

Simultaneous Achievement Of High Ethanol Yield And Titer In Clostridium thermocellum
Adaptive evolution successfully improved the ethanol yield and the titer from a rationally engineered CBP strain. This evolved strain has the highest ethanol yield and titer reported to date for C. thermocellum, and is an important step in the development of this microbe for industrial applications.

June 01, 2016

Workshop Summary: Lessons From Industry In Commercializing Biotechnology
This article summarizes lessons learned drawn from the "Technology Challenges and Opportunities in Commercializing Industrial Biotechnology Workshop," September 28-29, 2015, San Diego.

May 09, 2016

A Bifunctional Ammonia-Lyase Is Critical In Grass Cell Wall Biosynthesis Of Lignin
Using phylogenetic, transcriptomic and in vitro biochemical analyses, a single homotetrameric cytosolic bifunctional ammonia-lyase (PTAL) was identified among eight BdPAL enzymes in the model grass species Brachypodium distachyon. 13C stable isotopic labelling experiments along with BdPTAL1-down-regulation in transgenic plants showed that the TAL activity of BdPTAL1 can provide nearly half of the total lignin deposited in grasses, with a significant preference for S-lignin and wall-bound coumarate biosynthesis.

April 29, 2016

Ball-milled whole cell wall poplar samples in different NMR solvents.

Improved Whole Cell Wall NMR Measures Carbohydrate And Lignin Structures Without Derivatization
A new NMR solvent system composed of dimethylsulfoxide (DMSO-d6) and hexamethylphosphoramide (HMPA-d18) was developed to understand plant cell wall structures with minimal deconstruction and modification of biomass. The proposed bi-solvent system does not require derivatization or chemical modification, thus it facilitates easier sample preparation and less signal overlapping of correlation peaks compared to the traditional methods.

April 28, 2016

Reduction of lignin content.

Identification And Overexpression Of Knotted1-Like Transcription Factor For Lignocellulosic Feedstock Improvement
Using transgenic approach, a switchgrass Knotted1 transcription factor, PvKN1 was overexpressed in switchgrass and transgenic plants were characterized for recalcitrance and growth-related traits. PvKN1 overexpression resulted in reduced lignin deposition/biomass recalcitrance and improved sugar release efficiency without affecting various growth parameters such as tiller height, tiller number, fresh and dry biomass.

April 28, 2016

BRC Directors

Intellectual Property Impact of the BRCs
The Bioenergy Science Centers (BRCs) have generated over 500 invention disclosures, 328 patent applications (of which 45 have been awarded) which have led to 133 licenses or options with industry.

April 26, 2016

QTL for lignin content and glucose release

New Genetic Markers Identified For Switchgrass Cell Wall Composition And Conversion
A full-sib switchgrass population was developed by crossing a lowland genotype (AP13) to an upland genotype (VS16). QTL associated with the traits of interest were identified. Significant variation for glucose, xylose and total sugar release, and lignin content (19-27%) in the population; a unit increase in lignin content reduced total sugar release by an average of 10 mg g-1; and QTL analysis detected nine genomic regions underlying sugar release and 14 for lignin content.

April 26, 2016

adhE expression and ethanol production with old vs. new plasmids

Development Of Improved Expression Plasmid In Clostridium thermocellum
Clostridium thermocellum is a promising candidate for ethanol production from cellulose biomass yet requires metabolic engineering to further improve ethanol yield. An expression plasmid was redesigned to improve transformation efficiency and structural stability. The new expression plasmid had both significantly higher transformation efficiency than its predecessor (~2x more), and was also able to more robustly express genes of interest.

April 26, 2016

Structural Diversity Of Xylans Measured In Cell Walls Of Monocots
This study provides a new perspective on the diversity, function and evolution of monocot cell walls, enabling the manipulation of xylan structure and development of new technologies for the conversion of biomass from this important family of plants to yield commercially valuable and sustainable products.

April 25, 2016

BRCs Well Represented at Symposium on Biotechnology for Fuels and Chemicals
The Symposium on Biotechnology for Fuels and Chemicals (SBFC) was held April 25-28, 2016, in Baltimore, Maryland. As part of the overall symposium the combined Bioenergy Research Centers (BRCs) presented thirty-seven talks and twenty posters representing 25% of the talks and 10% of the posters.

April 18, 2016

Photo of the field in June 2013.

Lignification And Recalcitrance Affected By Age And Season In Field-Grown COMT-Downregulated Switchgrass
Previous field and greenhouse studies have shown that COMT-downregulation in switchgrass decreases lignin content and reduces the S/G ratio, thereby lowering recalcitrance and improving ethanol yields. Trends in lignification and recalcitrance differ between growing seasons. Third-year results showed an increase in lignification and decrease in sugar release across the season. S/G ratios increased over time in non-transgenic control lines, but were relatively stable in COMT-downregulated lines. During early (elongation) growth stages, cell wall chemistry was similar between COMT-downregulated and control lines. After reaching the reproductive growth stage in July, COMT-downregulated lines maintained reduced lignin content, decreased S/G ratios, and improved sugar release through December. No differences in rust susceptibility were observed.

April 08, 2016

Dr. Willington Muchero, lead ORNL inventor

GreenWood Resources, Inc. Licenses BESC Invention For Lignin Regulator
Studies on the natural variation in lignin content in poplar trees identified a gene with major effects on lignin biosynthesis, the key polymer hindering efficient biomass digestibility. GreenWood Resources, Inc. has nonexclusively licensed this jointly-owned BESC invention and will continue to partner with ORNL as they complete field studies.

April 01, 2016

BioEnergy Science Center (BESC) Website Receives A "Distinguished Technical Communication" Award
The Washington-Baltimore Society for Technical Communication (STC) recognized the BioEnergy Science Center (BESC) website with the "Distinguished Technical Communication" award in April 2016.

March 30, 2016

Overall schemes of fuels from sunlight and atmospheric carbon.

Review: Using Microorganisms For The Production Of Next-Generation Biofuels
A review on how microorganisms can be exploited for the production of next-generation biofuels: a) based on the ability of bacteria and fungi to use lignocellulose; b) via direct CO2 conversion using microalgae; c) lithoautotrophs driven by solar electricity; or d) relying on the capacity of microorganisms to use methane generated from landfill.

February 05, 2016

TEM micrographs of Avicel particles after incubation with Clostridium thermocellum

Discovery Of New Enzyme System Sheds Further Light On High Cellulolytic Performance Of C. thermocellum
C. thermocellum utilizes not only all the cellulase degradation mechanisms known today (cellulosomes and free enzymes), but also a new category of scaffolded enzymes not attached to the cell. This discovery explains C. thermocellum's superior performance on biomass. These scaffoldins were found to be essential for the well known digestion mechanism, i.e., the defibrillation of plant cell walls.

February 04, 2016

CBP conversion of untreated Populus with high S/G ratio of lignin yielded significantly better hydrolysis and fermentation products.

Contribution Of Lignin Composition And Structure To Improvements In Bioprocessing Of Populus Using CBP
Populus variants with low and high syringyl to guaiacyl (S/G) ratios were hydrolyzed and fermented with Clostridium thermocellum under CBP conditions, and compared to conventional fungal enzymes and yeast. The composition and structure of cellulose and lignin in the biomass was correlated with the bioconversion results. Samples with a S/G ratio of 2:1 were converted more rapidly and to a greater extent than biomass with a 1:2 S/G ratio. The CBP microbe hydrolyzed biomass two to three times more readily than commercial fungal enzymes in either scenario. Lignin polymer size (lignin Mw) was found to be lower for the low S/G biomass.

January 12, 2016

Solubilization of washed mid-season switchgrass by various biocatalysts.

Optimizing Biocatalyst-Feedstock Combinations to Achieve High Solubilization with Minimal Pretreatment
This study carried out the most comprehensive controlled comparison to date of lignocellulosic solubilization by various biocatalysts, and report initial exploration of enhancing microbial solubilization of cellulosic biomass via mechanical disruption (cotreatment). Trends drawn from time-course and end-point data for six conversion systems and three substrates were: greater-than-expected differences in the effectiveness of various biocatalysts; lower yields and greater dependence on particle size for Populus as compared to green or senescent switchgrass; equal fractional solubilization of glucan and xylan with no biological solubilization of the non-carbohydrate fraction of biomass; and two-fold increase in solubilization via mechanical disruption with greater benefits on partially-fermented feedstock.

January 06, 2016

Hemicellulose and Lignin

Review: How Plant Cell Wall NMR Provides Insights Into Biomass Chemistry and Structures
The structure of the plant cell wall and recalcitrance are intimately tied together such that fundamental breakthroughs in recalcitrance have driven the science of biomass characterization. Work from BESC and other energy centers has dramatically enhanced the resolution and productivity of Nuclear Magnetic Resonance (NMR) analysis of biomass polymers including lignin, cellulose and hemicellulose. Whole cell NMR analysis, as reviewed in this article, provides a relatively facile means of characterizing changes in lignin and hemicellulose that contribute to reduced recalcitrance in pretreated biomass and/or transgenic or natural variant plant species.

December 31, 2015

Comparison of cell wall (1) and growth (2) traits between T2-generation transgenic (+) and non-transgenic null-segregant (-) hybrid families.

Successful Transfer of Transgenic Switchgrass Low-Recalcitrance Traits into High-Yielding Field Variety
This study evaluated the feasibility of transferring the low-recalcitrance traits associated with a transgenic switchgrass line into high-yielding field varieties in an attempt to improve growth-related traits and to preserve the transgenic benefit. Our results provide insights into the possible improvement of switchgrass productivity via biotechnology paired with plant breeding.

December 31, 2015

GPC Curves

BESC Biomass Improvements In Lignin Extended From Grasses To Eucalyptus
Lignin reduction through breeding and genetic modification has the potential to reduce costs in biomass processing. Previous work demonstrated that both C3′H and C4H down-regulated lines had lower S/G ratios, lignin and recalcitrance. Both lines have significantly higher extractability under NaOH base extraction, indicating altered cell wall construction. Although the C3′H contained more H lignin than the C4H and control samples, only slight reductions in average molecular weights of the lignin and few changes in the lignin subunit were observed.

December 29, 2015

Preferential expression of FPGS1 in the vascular tissues of Arabidopsis.

Reducing Recalcitrance In Plants By Down-Regulating A Gene Involved In C1 Metabolism
Folylpolyglutamate synthetase (FPGS) genes and their corresponding mutants were characterized in Arabidopsis using cellular, biochemical, genomic and molecular approaches. Down-regulation of a single FPGS isoform leads to reduced lignin and altered hemicellulose in plants and therefore should be explored further for the improvement of lignocellulosic feedstocks.

December 26, 2015

Root enhancement in winter wheat due to S. vermifera colonization in vitro.

Review: Potential Impact of S. vermifera, A Unique Root Symbiont, On Agronomics of Biofeedstocks
Research performed with orchid mycorrhizal fungus Sebacina vermifera clearly indicates significant growth-promoting abilities in a remarkably broad spectrum of plants, including switchgrass, that rivals or even surpasses that of arbuscular mycorrhizae. Sebacinoid fungi should be considered as a previously hidden, but amenable and effective microbial tool for enhancing plant productivity and stress tolerance which has been validated in switchgrass, an important biofeedstock for biofuels.

December 14, 2015

System-wide Redox Stress Response: red indicates decreased transcription, (#) is the number of genes in the pathway decreased.

Towards Understanding Gene Systems Mediating Redox Homeostasis and Energy Metabolism in C. thermocellum
Current moderate ethanol yields and productivities as well as growth inhibition impede industrial deployment of this bacterium for commodity fuel production. Redox imbalances may contribute to incomplete substrate utilization, limit bioproductivity, and direct fermentation products to undesirable overflow metabolites. The redox systems identified are active and fungible in C. thermocellum upon introduction of a redox stress, and considered a link between redox stress and the occurrence of ethanol yield-limiting, unwanted metabolic processes. This expression study informs ongoing metabolic engineering efforts to increase bioproductivity in C. thermocellum.

December 10, 2015

Commercial Licensing of a BESC Global Genetic Regulator of Aromatic Biosynthesis
BESC has been studying SNPs and key compositional and conversion phenotypes in the natural variation population in Populus using GWAS (genome-wide association studies). A study of rare SNPs identified an unusual paralog in Populus which had taken on new regulator functions. Patent applications have been filed. Follow-on work is testing the utility in other plant species. The technology is being licensed to two companies who plan to commercialize the technology: GreenWood Resources and Forest Genetics International. Increasing biofuel yield or forage digestibility can lead to reductions in land use for these crops and thus contributing to long term sustainability.

October 26, 2015

Integrating mRNA and Protein Sequence Data to Enable Detection and Quantitative Profiling of Natural Protein Sequence Variants in Populus
In this study, we used next-generation RNA-sequencing data to generate genotype-specific protein databases, which allowed the identification and quantification of sequence variations at the protein level. A variety of natural sequence protein variants were quantified in two Populus trichocarpa genotypes. In addition to expected neutral single amino acid polymorphisms, investigators characterized polymorphisms predicted to be non-neutral and located in regions of the genome predicted to have undergone recent positive and/or divergent selection and therefore represent a candidate list of protein variants relevant to plant adaptability.

October 16, 2015

Direct Catalytic Upgrading of Ethanol is Efficient and Works Through a Hydrocarbon Pool Mechanism
Experiments with C2H5OD and in situ DRIFTS (diffuse reflectance FT infrared spectroscopy) suggest that most of the products come from the hydrocarbon pool type mechanism and endothermic dehydration step is not necessary. This mechanism supports the slightly exothermic nature of the reactions and potentially improves the process economics.

October 13, 2015

Metabolic Model Used to Predict and Remove Redox Bottlenecks in C. thermocellum Biofuel Production
Metabolic engineering strategies have been used to enhance ethanol production in C. thermocellum, a highly effective cellulose degrader. A central metabolic model has been constructed and validated for C. thermocellum which can expedite the process of designing and testing metabolic engineering strategies. Using the model for experimental design, investigators were able to highlight key overexpression targets for metabolic engineering such as RNF, NFN, or AdhE and to predict genotypes which may enhance ethanol yield.

October 13, 2015

Effect of glucan and cellulase loadings on ethanol yield for CELF-prepared corn stover

CELF Pretreatment Boosts Biofuel Yields and Titers
Novel Co-solvent Enhanced Lignocellulosic Fractionation (CELF) pretreatment has shown to be highly effective in achieving near theoretical sugars yields for corn stover. Earlier preliminary data also showed that CELF performs much better than dilute acid (DA) pretreatment; however, the effect of solids concentrations and enzyme loadings on ethanol yields and titers was not known. The results reported show that CELF pretreatment makes possible high ethanol yields and titers, and most importantly, at low enzyme loadings.

October 12, 2015

Molecular Dynamics Simulation Reveals Molecular Basis of Highly Efficient Lignin Solubilization
Previous BESC work has shown that pretreatment of biomass with a tetrahydrofuran (THF)–water cosolvent is highly effective at extracting lignin from biomass and achieving high yields of fermentable sugars from cellulose. However, the molecular mechanism of this effect is not known. In most aqueous-based pretreatments, lignin is not removed entirely from biomass because it aggregates onto the cellulose surface, blocking enzymatic access to cellulose. Under theta solvent conditions, polymers do not aggregate, thus providing a mechanism for the observed lignin solubilization that facilitates unfettered access of cellulolytic enzymes to cellulose. This mechanism may provide insights to improve this or other novel pretreatments.

September 22, 2015

Computational Inference of the Structure and Regulation of the Lignin Pathway in Switchgrass, Panicum virgatum
Recalcitrance is partially due to the heterologous polymer lignin. The biosynthetic pathway leading to monolignols, and thus to lignin, is not completely known in switchgrass, and difficulties associated with in vivo measurements of its intermediates pose a challenge for a true understanding of the functioning of the pathway. However, such an understanding is a prerequisite for manipulations of transgenics toward altered lignin properties and a reduction in recalcitrance. The computational analysis suggests regulatory mechanisms of product inhibition and enzyme competition, which are well known in biochemistry, but so far had not been reported in switchgrass. Our deepened understanding of the pathway allows us to make prediction regarding combined knock-downs that are likely to alter lignin properties significantly.

September 01, 2015

Highest Yield to Date of Ethanol from Engineered Clostridium thermocellum
C. thermocellum is one of the fastest growing anaerobic organisms on crystalline cellulose. Metabolic engineering is required to increase ethanol yield by eliminating pathways to other fermentation products. Effectively eliminated lactate, acetate, formate, and most H2 production. Ethanol yield doubled relative to the wild type, both on model substrates and dilute-acid pretreated plant biomass. Allowed for higher titer ethanol production from cellulose in the absence of pH control. Highest ethanol yield achieved to date in C. thermocellum. Created platform strain for further metabolic engineering for consolidated bioprocessing.

September 01, 2015

Engineering of a Thermophilic, Cellulolytic Microbe to Produce an Advanced Biofuel
Production of isobutanol from cellulose in minimal medium achieved 5.4 g/L titer and roughly 41% of theoretical yield. This result successfully demonstrates the advantages of using a consolidates bioprocessing approach to produce an advanced biofuel. This is a measureable and significant step towards commercially relevant titers and yields. Isobutanol can be utilized as a "drop-in" biofuel, as a substrate for upgrading into hydrocarbon fuels, or as a feedstock for other bioproducts.

August 29, 2015

Mechanism for expression of xylanases induced by extracellular xylan via sig16

New Insights into Regulation of Cellulosomal Xylanase Genes in Clostridium thermocellum
Clostridium thermocellum is regarded as a model cellulolytic thermophile, and can grow well on un-pretreated lignocellulosic feedstock, yet it does not utilize C5 sugars (e.g., xylose, arabinose) despite including xylanases in its cellulosome. While unable to utilize C5 sugars C. thermocellum has a set of sigma factor regulated xylanases that, when expressed in response to biomass, can increase the amount of solubilized carbohydrate including glucan.

August 29, 2015

BESC co-inventors Sara Jawdy and Lee Gunter evaluating growth performance of rice plants carrying the lignin reducing-flavonoid enhancing mechanism.

Forage Genetics Plans to Commercialize BESC Invention
The invention provides a genetic mechanism for the reduction of lignin biosynthesis while increasing concentration of desirable flavonoids. Reduced lignin content increases digestibility and nutritional value of animal feedstocks such as alfalfa, corn and sorghum. Forage Genetics plans to evaluate commercial viability of this technology in alfalfa, corn and sorghum forage crops as animal feedstocks.

August 27, 2015

Sugar Release with 17.5mins Pretreatment at 180C

Application of Results from Down-Regulation of Lignin in Populus and Switchgrass Leads to Similar Effects in Eucalyptus
Changes to gene expression in switchgrass and Populus have shown increased sugar release and reduced recalcitrance. This work sought to transfer results to Eucalyptus, a fast-growing warm climate woody biofeedstock also suitable for cellulosic biofuel production. This work extends result that lowering overall lignin content, rather than altering S/G/H ratios, has the largest impact on reducing recalcitrance in eucalyptus.

August 01, 2015

Expression and Secretion of E1 in C. bescii Enhances the Cellulolytic Ability of Its Secretome
The in vitro activity of the C. bescii secretome was significantly increased (17.5% at 65oC) by the addition of E1. The E1 protein acts synergistically with CelA to digest cellulose and this result offers the possibility of adding enzymes into engineered secretomes for improved biomass deconstruction. This work contributes to an understanding of the action and limitations of the CelA enzyme that will enable enzyme engineering to generate novel enzymes mixtures for biomass deconstruction.

July 27, 2015

Discovery of Novel Tungsten-Utilization Pathway in Caldicellulosiruptor bescii
Very few microbes are known to require tungsten for growth. Analysis of the genome sequence of C. bescii has unexpectedly revealed a set of transporter genes that are highly specific for the uptake of tungsten. Although its role within the cell is as yet unknown, the conservation of XOR and associated pyranopterin synthesis genes across the genus Caldicellulosiruptor, suggest that this novel tungstoenzyme serves an important role in the primary metabolism of these cellulolytic species.

July 20, 2015

Catalytic Mechanism and Unique Low pH Optimum of Caldicellulosiruptor bescii Pectate Lyase Explained via Crystallography and Mutagenesis
Pectate lyases are a degradation enzyme for an important if small polymer component of plant biomass. The structural changes upon all mutations are analysed and their effect on activity explained. This paper provide a comprehensive examination on the PL family 3 structure and function with an extensive mutagenesis study.

July 15, 2015

Comparison of MethylC-seq and 4mC-TAB-seq.

Rapid Identification of Restriction Systems that Prevent DNA Transformation of Uncharacterized Bacteria
New 4mC-specific sequencing strategy will help uncover and facilitate investigation of 4mC in bacterial R-M systems and can advance the rapid and efficient genetic engineering of these bacteria in the future.

July 01, 2015

Imaging with HPFM (a,c,e,g) complemented with confocal Raman (b,d,f,h) cross-section of fresh (a,b), (...more)

New Hybrid Microscope Provide Unparalleled Submicron View of Plant Cell Walls
A novel, noninvasive method to explore submicron structures of biomass before and after treatment. Researchers will be able to study samples ranging from naturally occurring biological polymers, tissues and plant cells to engineered nanoparticles and nanostructures.

June 22, 2015

Green: Fungal mycelia Red: Root cell membrane

Novel Delivery System for Root Symbiotic Fungus, Sebacina vermifera, Enhances Biomass yield of Low Lignin Switchgrass Lines
S. vermifera colonization enhanced plant biomass growth by 15-50% independent of delivery method; however, clay-based delivery system increased the percentage of fungal biomass in planta. Root biomass and fungal colonization was significantly improved with clay-based system.

June 19, 2015

Pore size distribution of Populus before and after dilute acid pretreatment measured by Mercury porosimetry.

The Role of Nano-Porosity in Lowering Recalcitrance
Pore size distribution analysis indicated that the most fundamental barrier in terms of the biomass porosity scale for efficient enzymatic hydrolysis is the nano-pore space formed between coated microfibrils, although some of the porous architecture such as the cell lumen and pit could be severely destroyed after pretreatment. Lignin still plays a negative role in the processes of enzymatic hydrolysis by binding to cellulases; but this negative effect could be compensated by the positive effect of cellulose accessibility.

June 08, 2015

Transport energy use by fuel and year, displaced CO2 emissions by fuel and year, and tota (...more)

Making the Case for Why Biofuels are Needed to Achieve Low-Carbon Transport Through 2050 and Beyond
Even with aggressive reductions in travel growth, shifts to mass transport modes, strong efficiency improvements and deep market penetration by electricity and hydrogen, there remains a large demand for dense liquid fuels: 80% of transport energy demand in 2050, and 50% in 2075. If conventional fossil fuels were used to meet this demand, emissions from the transportation sector would exceed the economy-wide emissions consistent with the two degree scenario. It will likely be difficult to achieve a low-carbon transport sector without widespread use of biofuels.

June 01, 2015

Kevin Wenger of Mascoma, a subsidiary of Lallemand, LLC and Paul Gilna of BESC

BESC Partner Mascoma Launches C5 FUEL™ — A Yeast Engineered for Improved Cellulosic Biofuel Production
The new strain of yeast simultaneously yields 97 percent conversion of xylose – the C5 sugar – and glucose and it accomplishes this feat in less than 48 hours. The launch of a commercially available product designed for producers in the cellulosic biofuels market is an important milestone for BESC and the BRC program.

May 29, 2015

Book edited by BESC Scientist on Consolidated Bioprocessing
Mike Himmel has edited a new book on "Direct Microbial Conversion of Biomass to Advanced Biofuels," Elsevier, London/New York, NY. 2015. Several chapters report on BESC work and acknowledge BER support.

May 14, 2015

Perspective: Potential Impact of Modern Biotechnology on Future Supply of Biofuels
This study assesses the feasibility and advantages of near-future (10-year) biotechnological developments for a U.S. biomass-based supply chain for bioenergy production.

April 27, 2015

DOE Bioenergy Research Centers Well Represented at the 37th Symposium on Biotechnology for Fuels and Chemicals — 2015
19% of oral presentations were from BESC, GLBRC and JBEI (26 out of 138) and approximately 27 BRC posters were presented. There were 650 attendees from academia, industry and government.

April 15, 2015

Glucose Regression Model

Pyrolysis Molecular Beam Mass Spectrometry as a High-throughput Method for Determining Biomass Sugar Content
Py-MBMS can be used to estimate sugar content of biomass using regression models. The method is much faster (< 2 min/sample) than traditional hydrolysis followed by HPLC with much less sample preparation. This method introduces a new, high-throughput technique for sugar compositional analysis. The py-MBMS spectra can also be used to estimate lignin and S/G ratio, providing a comprehensive analysis.

April 01, 2015

Dr. Liao received his Bachelor of Science degree at the National Taiwan University and his Doctorate at the University of Wisconsin-Madison

James Liao Elected to the National Academy of Sciences
Dr. Liao's science focuses on developing the biological production of the liquid fuel isobutanol while simultaneously consuming carbon dioxide and developing different methods to create liquid fuels from electricity and from waste proteins; as well as identifying a more efficient way to convert sugars into fuels. Dr. Liao serves as the BESC Activity Lead for Butanol Production by CBP.

March 31, 2015

Wild type cellulase, CbhA; Chimeric cellulase with swapped linker domain.

Improved multi-functional cellulase awarded US Patent
One of the most abundant cellulase components of the CipA scaffold, CbhA, was improved by replacing the Fn3 domains with a thermal stable linker peptide from another cellulolytic microbe. Changing out the linker domain doubled the activity of the CbhA enzyme. U.S. Patent 8993276 (March 31, 2015) was issued for the invention of the new improved cellulase enzyme.

March 21, 2015

New Promoters for Metabolic Engineering of C. thermocellum
Several promoters were identified (eno, cbp, cbp_2, 815, 966, 2638 and 2926) which gave high expression and high enzymatic activity of both reporter genes in C. thermocellum.

March 16, 2015

Release of a genome dataset comprised of results from multiple sequencing modalities
Access to a comprehensive dataset representing a range of sequencing technologies can serve as a benchmark/training dataset to assist others in developing new technologies and computational algorithms.

March 05, 2015

Review: Ethanol Production by Engineered Thermophiles
Four potential pathways from the glycolysis intermediate pyruvate into ethanol have been identified: pyruvate decarboxylase (PDC), pyruvate dehydrogenase (PDH), pyruvate ferredoxin oxidoreductase (PFOR) and pyruvate formate lyase (PFL). Three of these pathways (PDC, PDH and PFOR) have been used in thermophiles for ethanol production at significantly higher yield, and two pathways (PDC and PFOR) resulted in ethanol production at higher titer.

March 01, 2015

Reducing Recalcitrance and Increasing Growth in Populus by Downregulation of Matrix Polysaccharide Biosynthetic Gene
Populus GAUT12.1-Knockdown lines have 3 to 7% increased sugar release, confirming strategy to reduce recalcitrance. Populus GAUT12.1-Knockdown lines have 12 to 52% increase in plant height, 12 to 44% increased stem diameter, and 52 to 117% increased leaf size, demonstrating that plant growth and recalcitrance are linked and can simultaneously be improved. Improved biomass phenotype was accompanied by reduction in pectin and xylan cell wall content, supporting hypothesis that pectin and xylan contribute to a wall infrastructure(s) critical for recalcitrance.

March 01, 2015

Tāpirins, a New Class of Biomolecules, Mediate Attachment of Caldicellulosiruptor Species to Cellulose
Tāpirins represent a new class of proteins used by Caldicellulosiruptor species to attach to insoluble cellulose. The tapirins establish a new paradigm for how cellulolytic bacteria adhere to cellulose.

February 27, 2015

Genomes Sequenced

Review paper summarizes insights from 20 years of bacterial genome sequencing
Second and third generation sequencing have greatly reduced the cost of sequencing and made it accessible to a great many more individual investigators. Sequencing of bacterial genome sequences is now a standard procedure and the information from tens of thousands of bacterial genomes has had a major impact on the view of the bacterial world.

February 21, 2015

Ten initial BESC Populus TOP lines identified with attractive biofuel traits in conversion and growth
Simultaneous comparison of Populus TOP lines for enhanced growth and biofuel traits will allow BESC researchers to identify the best among them to further evaluate in field studies (e.g., to determine sustainability traits) in preparation for application and licensing with commercial partners.

February 11, 2015

Novel Pretreatment Greatly Reduces Enzyme Loadings and Increases Biofuel Yields
Co-solvent Enhanced Lignocellulosic Fractionation (CELF) pretreatment achieved about a 10-fold reduction in enzyme loadings compared to traditional DA pretreatment. CELF also removed a majority of biomass lignin (over 90% in wood) that could be precipitated as a high purity product suitable for conversion to high value chemicals and materials.

February 01, 2015

Glycome Profiling of AFEX™ Pretreated Biomass Provides Insights into Plant Cell Wall Structure
AFEX™ pretreatment, in general, causes loosening of non-cellulosic glycans, especially xylans, in diverse biomass types resulting in a reduced recalcitrance. These effects of AFEX™ vary significantly depending on the type of biomass. Monitoring changes in cell wall glycan compositions and their relative extractability for untreated and pretreated plant biomass provide improved understanding of variations in structure and composition of plant cell walls that relate to the role(s) of matrix-polysaccharides in recalcitrance.

January 30, 2015

Production of lignofuels and electrofuels by extremely thermophilic microbes
Caldicellulosiruptor and Pyrococcus sp. could be potentially metabolically engineered to produce advanced biofuels compatible with existing infrastructure, generating so-called lignofuels and electrofuels, respectively. Thermophilic biomass degradation is more efficient, and thermophilic fuel synthesis affords the opportunity for simultaneous distillation of volatile products.

January 30, 2015

Novel approach to blocking H2 production increases Clostridium thermocellum ethanol yield
Approach allowed four hydrogenases to be simultaneously inactivated using only two mutations on the chromosome. Highest C. thermocellum ethanol yield reported so far shows promise for further yield improvement. Provides insight into redox balancing in C. thermocellum. Mutation demonstrates ethanol tolerance in C. thermocellum is more complicated than previously thought.

January 25, 2015

Field experiment in the second (2013) growing season. Photographed August 1 2013

Field performance of modified switchgrass demonstrates a 'Goldilocks Effect'
This study showed that too high PvMYB4 transgene expression is fatal to plants in the field, whereas too low expression shows no difference from controls; but when optimal expression is achieved the altered switchgrass produces higher biomass and biofuel yields. These gains represent the potential for a doubling of biofuel production per hectare over conventional feedstocks. This is the highest potential gain reported to date from any field-grown modified feedstock.

January 23, 2015

A four-year-old P. trichocarpa in a common garden

Rapid gene discovery and validation using the BESC Populus association population
Complementary QTL and association mapping are powerful tools for rapid gene discovery with no a priori candidate gene selection. This proof of concept in a perennial organism opens up opportunities for discovery of novel genetic determinants of economically important but complex traits in plants.

January 11, 2015

BESC Novel Approach to Bioenergy Outreach and Education
BESC and the Creative Discovery Museum have developed and implemented a novel 'hub and spoke' model by working with science museums and centers around the country to deliver impactful science outreach in biofuels. The program continues to move toward becoming self-sustaining with over 75% of the hands-on interactions being supported by local schools and museums.

January 01, 2015

BioEnergy Research Vol. 8(3), 2015.

BioEnergy Research Special Issue Features DOE Bioenergy Research Centers
Sixteen articles are being released in a special issue devoted entirely to the work of the three DOE Bioenergy Research Centers including an overview of the three Centers and articles on feedstock sustainability, pretreatment and fermentation.

December 30, 2014

Data from M. Nar and N. D’Souza, University of North Texas

Discovery of C-lignin shows promise as feedstock for carbon fiber
The higher purity of C-lignin over Kraft fibers and PAN-based fibers is expected to translate into higher mechanical stiffness, thermal and electrical conductivity.

December 30, 2014


BIOFUELS lessons on Outreach site
BESC has an ongoing outreach effort with the Creative Discovery Museum (CDM) of Chattanooga, Tennessee. targeting 4th through 8th grades and their families. This successful program has reached over 115,000 students and parents to date through the Farming For Fuels curriculum and activities.

December 16, 2014

Identification of key transcription factor that controls chlorophyll degradation and leaf senescence
This represents the first confirmation of NAP in positive regulation of ABA biosynthesis in Arabidopsis, and thus a key factor in induction of chlorophyll degradation genes and plant senescence.

December 16, 2014

Young, middle-aged, and senescent tillers

Switchgrass transcription factors linked to nutrient efficiency in senescence
This study confirms that PvNAC1 and PvNAC2 play conserved roles in leaf senescence in switchgrass and hold promise for improving nutrient use efficiency in switchgrass through genetic manipulation or breeding.

December 03, 2014

Toward improving tolerance of thermophilic microorganisms to pretreatment inhibitors
A heat-stable furfural reductase has been identified in T. pseudethanolicus 39E for inhibitor detoxification. It promises rational improvement in tolerance and has already shown promising results when expressed in the cellulolytic thermophile, C. bescii (in preparation).

December 01, 2014

The thermodynamic origin of the reduced lignin-hemicellulose association is found to be a more favor (...more)

ORNL TITAN supercomputer used to determine molecular basis of reduced recalcitrance
Reduced lignin-hemicellulose association increases the cell wall porosity in the mutant plants, thus explaining their easier deconstruction to biofuels. This work illustrates how MD simulations can be a useful tool for predicting and explaining the effects of changes in plant genotypes on cell wall phenotypes.

November 25, 2014

Simons' stain dye adsorption diagram for NaOH-, Ca(OH)2-, and ammonia-pretreated Populus.

Characterization of various alkaline pretreatment methods on cellulose structure and accessibility (A joint BESC/ORNL Biofuels SFA effort)
The study identifies important cellulose structural features and relevant parameters related to biomass recalcitrance and the changes that occur in cellulose structure and accessibility upon a variety of low-cost and mild alkaline treatments and determines some of the key factors responsible for biomass recalcitrance.

November 11, 2014

Synthetic AOR/AdhA Pathway for Bioalcohol Production from Sugar (top) or from Organic Acids with H2 or CO2

Development of a synthetic metabolic pathway for sugar to bioalcohol production in thermophilic microorganisms
The AOR/AdhA pathway is a new energy-conserving, redox-balanced route for sugar to ethanol conversion that could potentially be used in any CBP microorganism. Carbon monoxide and hydrogen gas (syngas) can be used as the driving force for bioalcohol production from a range of aliphatic and aromatic aldehydes, and is a potentially game-changing strategy for syngas fermentation.

November 02, 2014

Expression patterns of the three putative C20 GA2ox genes among tissues and developmental stages in switchgrass as determined by zRT-PCR.

Overexpression of gibberellin 2-oxidases leads to reduced recalcitrance in switchgrass
Results suggest that overexpression of GA2ox genes in switchgrass is a feasible strategy to improve plant architecture and reduce biomass recalcitrance for biofuel.

October 10, 2014

Symposium on Biotechnology for Fuels and Chemicals and Annual World Congress on Industrial Biotechnology
Bioenergy Research Centers Update and DOE Bioenergy Research Centers Achievements

October 10, 2014

BioEnergy Science Center 2014 Science Retreat Chattanooga, Tennessee

BioEnergy Science Center 2014 Science Retreat Chattanooga, Tennessee
BioEnergy Science Center 2014 Science Retreat Chattanooga, Tennessee

October 07, 2014

Morphology and cell wall composition of Populus deltoides plants overexpressing UGPase relative to (...more)

Overexpression of key cellulose biosynthesis precursor enzyme, UGPase, reveals altered sugar and secondary metabolism in Populus
This study provides evidence that manipulation of sugar metabolizing enzymes, such as UGPase, can have qualitative and quantitative effects on secondary metabolic pathways of shikimate, phenylpropanoid, and lignin biosynthesis. A cross-collaborative BESC effort is investigating the substrate preferences and reaction kinetics of UGPase to understand the observed pleiotropic phenotype. In integration with assessments of effects of other gene sequence and expression variations, our studies are yielding critical insights into biomass formation in Populus.

October 01, 2014

New Insights into Biomass Deconstruction from Deletion of Genes in Caldicellulosiruptor
Three papers report the first in vivo analyses of genes predicted to be important for plant biomass deconstruction in the context of the entire suite of enzymes in Caldi. This demonstrates the potential of identifying and engineering enzymes to improve both substrate utilization and the range of substrates utilized. This is an example of using microbial deconstruction capabilities to probe important plant cell wall structures such as pectin.

September 14, 2014

MALDI-TOF MS of the products in the absence and presence of IRX10-L, consistent with the sequential (...more)

Discovery of two key enzymes in xylan synthesis and acetylation in plant cell walls
Two key enzymes for two critical process in xylan (and secondary plant cell wall) synthesis have now been identified, purified and confirmed. These finding will accelerate our understanding and ability to manipulate plant cell wall structures for advanced renewable feedstocks for conversion into sugars and fuels or into valuable products such as biomaterials.

September 02, 2014

Process scheme for a closed-loop biorefinery using ILs derived from lignocellulosic biomass

Ionic Liquids (ILs) derived from biomass – provide an effective biomass pretreatment
Deriving ILs from lignocellulosic biomass shows significant potential for the realization of a “closed-loop” process for future lignocellulosic biorefineries and has far-reaching economic impacts for other IL-based conversion technology currently using ILs synthesized from petroleum sources.

August 24, 2014

Locations of plantations (yellow diamonds) and tree collections. Colors indicate genetically differentiated groups.

Genome-wide adaptive variation in Populus trees revealed by massive population genomic analysis
This study is the first to explore the genomic legacy of selection across an entire tree genome and highlight both the wide range of selection pressures as well as the climatic influence on phenological systems and growth. This study provides new insights into mechanisms of adaptive evolution, and provides potential targets for genetic improvement for biofuels production as well as the management of natural populations in the face of environmental change.

August 16, 2014

Linville, et al., “Transcriptomic analysis of Clostridium thermocellum Populus hydrolysate-tolerant (...more)

Clostridium thermocellum develops increased cellular efficiency to tolerate Populus hydrolysate
A conceptual model of the improved mutant strain was developed, which provides a summary of the major mutations and related changes in gene expression or pathway activities. These changes resulted in an overall increase in cellular efficiency of the hydrolysate-tolerant organism by increased overall energy production and conversion rather than specific tolerance genes. In contrast, the wild type response to inhibition appears as decreased growth and increased expression of stress response genes. These results suggest new global strategies for engineering the development of increased tolerance to biomass hydrolysates in microbial processes used for biofuel production.

August 05, 2014

Gene co-expression networks for Arabidopsis PrR1 and PrR2. (A) PrR1 is co-expressed with many known (...more)

New relationship between lignan biosynthesis and lignin distribution in secondary cell wall structure in Arabidopsis
Provides new evidence for a relationship between lignan synthesis and the role of lignin in secondary plant cell wall structure, which may shed light on evolution of plant secondary wall structure. Further analysis of this relationship may provide strategies for reducing recalcitrance of lignocellulosic biomass while enhancing plant defense.

June 03, 2014

16S rRNA tree of the 40 Clostridium genomes: biomass degraders (red),animal pathogens (blue), other (black)

Comparative Analysis of 40 Genomes provides new insights into biomass degradation by Clostridia
New insights provided by this analysis could provide useful information to mechanistic studies of biomass degradation and the range of metabolic pathways in Clostridia. This range of pathways may aid to designing or improving more efficient biomass degraders. Note: Over 100 clostridial genomes have now been sequenced (2014); therefore these types of analyses will need to be repeated on a regular basis as more genomes become available.

June 02, 2014

Lactate, acetate and ethanol production by wild-type (I), ldh- (J), and ldh-AdhE+ (K) strains of C bescii grown on non-pretreated switchgrass

Direct conversion of biomass to biofuel without pretreatment using engineered Caldicellulosiruptor bescii
This work reports the direct conversion of plant biomass to a fuel without pre-treatment which is the major economic barrier to commercialization It is also the first demonstration of metabolic engineering of an extreme thermophilic organism for conversion of lignocellulosic biomass to a liquid fuel

May 22, 2014

Conversion (mg/g glucan loaded) for C. thermocellum mutant M1570 and wild-type DSM 1313 strains on b (...more)

Improved production from modified switchgrass when combined with an engineered C. thermocellum strain
This is the first report of the use of a microbe engineered to produce increased amounts of a biofuel on a bioenergy feedstock that has been modified for the same purpose. The results demonstrate the potential additive advantages from combining a modified feedstock with an engineered consolidated bioprocessing microorganism.

May 15, 2014

Ragauskas, et al., “Lignin valorization: improving lignin processing in the biorefinergy,” Science, 2014

Lignin Valorization: Improving Lignin Processing in the Biorefinery is outlined in Science
This perspective review offers a rationale for optimism that more value can be derived from lignin within a biorefinery due to current and project research

May 13, 2014

Glucose yields in solution following enzymatic hydrolysis of dilute acid (DA), ammonia fiber expansi (...more)

Comparison of Enzymatic Reactivity of Corn Stover Solids
Initial hydrolysis yields were highest for substrates with greater lignin removal, a greater degree of change in cellulose crystallinity, and high enzyme accessibility. Final glucose yields could not be clearly related to concentrations of xylooligomers released from xylan during hydrolysis. Overall, none of these factors could completely account for differences in enzymatic digestion performance of solids produced by AFEX, DA, and IL pretreatments.

April 30, 2014

Lin, et al., “Isobutanol production at elevated temperatures in thermophilic Geobacillus thermoglucosidasius,” Metabolic Engineering 24, 1-8, 2014

Microbial production of isobutanol at elevated temperatures
First demonstration of isobutanol production on a recombinant bacterium at elevated temperature

April 21, 2014

University of Tennessee switchgrass

Field study of COMT transgenic switchgrass
The lignin-modified transgenic switchgrass had similar gains in sugar release (up to 34% higher) and biofuel production (up to 28% higher) as observed when comparing transgenic and control plants grown in the greenhouse. The greenhouse-grown experimental plant tissue was live and green, compared with the senesced, brown tissue harvested at the end of the growing season from the field experiments. The COMT2 transgenic event yielded over 50% more liters of ethanol per hectare compared with the control, which demonstrates that transgenic feedstocks could make important improvements in lignocellulosic biofuels.

March 21, 2014

Brown, S. D. et al., “Comparison of single-molecule sequencing and hybrid approaches for finishing t (...more)

Genomic analysis of the industrially relevant Clostridium autoethanogenum genome reveals potential role for CRISPR systems
This study demonstrates value of PacBio single molecule sequencing in producing single contig finished genomes. Potential associations between plasmid content and CRISPR systems may have implications for historical industrial scale Acetone-Butanol-Ethanol (ABE) fermentation failures and future large scale bacterial fermentations.

March 03, 2014

Dr. Arthur J. Ragauskas

Art Ragauskas Named Fourteenth University of Tennessee Oak Ridge National Laboratory Governor’s Chair
Dr. Arthur J. Ragauskas will serve as Governor’s Chair for Biorefining, based in the University of Tennessee’s (UT) Department of Chemical and Biomolecular Engineering with a complementary appointment in the UT Institute of Agriculture’s Department of Forestry, Wildlife, and Fisheries. He will serve in the Oak Ridge National Laboratory (ORNL) Energy and Environmental Sciences Directorate, Biosciences Division, and as a member of the Department of Energy’s (DOE) BioEnergy Science Center (BESC).

March 01, 2014

Creative Discovery Museum

BESC Communicates to Science Museum Community on Farming for Fuels Outreach Program
Farming for Fuels has grown into a program with a national presence and is currently being used in 20 science centers and museums in 12 states; over 80,000 people across the United States have now participated in the program. The journal in which this article appears was circulated to nearly 600 science-technology museums in over 40 countries, greatly increasing the awareness of this program across the museum educational community.

January 31, 2014

BESC Inventions, Licenses and Patents Awarded

BESC – Awarded Patents – FY2014
BESC – Awarded Patents – FY2014

December 30, 2013

Chemical Factors that Control Lignin Polymerization
The work provides a coherent framework for understanding the propensity of different terminal subunits in lignin to undergo growth or termination.

December 20, 2013

Cellulase Diversity – CelA is a highly active and stable cellulase with a Novel Digestion Mechanism
CelA and multi-functional cellulases represent a distinct paradigm for cellulose digestion with high activity created by combining modules separated by long linker peptides on the same gene product.

November 30, 2013

Genomics approach to deciphering lignin biosynthesis in switchgrass (Panicum virgatum L.)
This work provides a blueprint and new resources for the systematic genome-wide study of the monolignol pathway in switchgrass as well as other C4 monocots.

October 10, 2013

BioEnergy Science Center Achievements
Dr. Xing Xu; Dr. Art Ragauskas; Dr. James (Jim) Liao; Dr. Richard Dixon; Dr Steve Brown; Dr. Michael Udvardi

September 12, 2013

Robust RNA-Seq and DNA Microarray Comparison for C. thermocellum Fermenting Biomass
High degree of concordance in gene expression measurements between transcriptomic platforms. Appropriate normalization of RNA-Seq data is essential with a scaling factor based on the geometric mean of the RNA-Seq data most closely aligning with the microarray analyses of the same samples in this study.

June 01, 2013

Progress in Development of Caldicellulosiruptor as a Consolidated Bioprocessing (CBP) Platform
New capabilities in metabolic pathway engineering combined with intrinsic utilization of lignocellulosic materials position these thermophilic cellulolytic organisms to provide a new paradigm for consolidated bioprocessing of fuels and other products from biomass. Recent development of methods for genetic manipulation has set the stage for rational engineering for improved biofuel production.

May 01, 2013

Caldicellulosiruptor bescii (C. bescii) degrades lignin in switchgrass as well as cellulose and hemicellulose
Chemical and thermal pretreatment strategies for conventional cellulosic biofuels production contribute a significant cost overhead to the overall process. This unexpected result of simultaneous lignin and carbohydrate solubilization by C. bescii bodes well for complete industrial conversion by extremely thermophilic microbes of biomass that requires limited or no chemical pretreatment.

April 08, 2013

Fungal cellulases and complexed cellulosomal enzymes exhibit different yet synergistic mechanisms in cellulose deconstruction
New understanding of mechanistic differences between these important enzymatic paradigms highlights new opportunities for combining these two systems for enhanced performance, and allows for development of an optimum strategy through enzyme engineering that employs both approaches.

April 01, 2013

Overexpression of lignin transcription factor in switchgrass significantly increases biofuel yields
Results have demonstrated that overexpression of PvMYB4, a general transcriptional repressor of the phenylpropanoid/lignin biosynthesis pathway, can lead to a very high yield ethanol production through dramatic reduction of recalcitrance. MYB4-OX switchgrass is an excellent model system for understanding recalcitrance, and provides new germplasm for developing switchgrass cultivars as biomass feedstocks for biofuel production.

January 05, 2013

Development of an integrated transcript sequence database and a gene expression atlas for gene discovery and analysis in switchgrass (Panicum virgatum L.)
Switchgrass research will be enhanced by this webserver. A site for these data sets provides a solid foundation for functional genomics and breeding in switchgrass and provides annotations that helps users formulate hypotheses about gene function. The server handles RNA seq data also, which means that it can serve as a ‘one-stop-shop’ for switchgrass transcriptomics.

January 01, 2013

New plant structure (APAP1) discovery may lead to improved biofuel processing
APAP1 is contrary to prevailing cell wall models that depict separate protein, pectin and hemicellulose polysaccharide networks. The modified sugar composition and increased extractability of pectin and xylan immunoreactive epitopes in APAP1 mutant aerial biomass support a key role for the APAP1 proteoglycan in plant wall architecture and function. The existence of APAP1 in plant walls has significant consequences for understanding of plant wall architecture and function, and potentially for engineering plant cell walls for improved agronomic and renewable biomaterial uses. Pectin, a primary cell wall component, is important in secondary cell wall structure.

November 23, 2012

Nanoscale Architecture of Plant Cell Walls Determines Their Accessibility and Digestibility by Enzymes
Plant cell walls are complex nanocomposites containing networks of cellulose fibrils and complex “matrixing” polymers. There is little agreement about which plant cell wall features most affect digestibility by microbes and cellulolytic enzymes. The overall performance of biomass saccharification may be attributed to the synergistic action of many complementary enzymes, which makes it difficult to study one factor at a time. Traditional solution methods have suffered from the classical ensemble average limitation presented by analysis of these mixtures of complex biomass, and the bulk data measurements gathered are therefore sometimes inconclusive and in part contradictory. Our imaging data support the hypothesis that ideal pretreatments should maximize lignin removal and minimize polysaccharide modification, thereby retaining the essentially native microfibrillar structure and improving accessibility.

October 16, 2012

New Developments in High-performance Proteomics Identify Important Networks and Pathways in Populus
This large scale dataset features the first systems-wide survey of protein expression across different Populus organs and strongly testifies to the credibility of the approach; instead of quantitatively comparing a few proteins, a systems view of all the changes associated with a given cellular perturbation can now be made. Most large-scale proteome analyses with Populus are performed on isolated organs, this method combines state-of-the-art sample preparation methodology to generate a high-coverage proteome atlas of the principal organ types from Populus.

October 10, 2012

Methods in Molecular Biology
Of the twenty-one chapters in Volume 908 of “Methods in Molecular Biology” released in August 2012, thirteen chapters covered the science of BESC.

October 10, 2012

Industrial Biotechnology Special Issue on Advances in Biomass Characterization
The August 2012 issue of Industrial Biotechnology will feature five articles by BESC scientists and one article featuring JBEI research. The featured cover art highlights the work of BESC researchers Marcus Foston and Arthur J. Ragauskas at the Georgia Institute of Technology.

September 28, 2012

3-Dimensional Chemical Imaging of Populus Tension Wood by Time-of-Flight Secondary Ion Mass Spectrometry
This demonstrates the usefulness of 3D TOF-SIMS to the study of biomass recalcitrance by using tension wood samples as a model substrate to investigate how chemical spatial and lateral heterogeneity affects recalcitrance. Spatial distribution of major cell wall components in biomass is critical to bioconversion and can provide useful information in the future in determining the effects of pretreatment or genetic variation.

September 28, 2012

Down-Regulation of the Caffeic acid O-methyltransferase Gene in Switchgrass Reveals a Novel Monolignol Analog
The more facile breakdown of cell walls of COMT-deficient plants is associated with increased concentrations of phenolic acid and aldehyde inhibitors of microbial fermentation, and a monolignol analog of sinapyl alcohol that is not integrated into the cell walls.

August 22, 2012

BESC Reports Successful Genetic Manipulation of Caldicellulosiruptor
This technology is being established in BESC partner labs and will allow the metabolic engineering of these bacteria for the direct conversion of lignocellulose to biofuels such as ethanol and butanol.

August 14, 2012

Discovery of plant glucuronoxylan synthesis gene lays groundwork for improved biofuel processing
The ability to selectively manipulate polysaccharide methylation may provide new opportunities to modulate biopolymer interactions in the plant cell wall of bioenergy crops such as switchgrass. This work extends the portfolio of BESC structural targets that can be modified either alone or in combination to increase the economic value of lignocellulosic biomass.

July 23, 2012

BESC Reports Clostridium thermocellum Ethanol Stress Responses
Largest responses were related to nitrogen uptake and metabolism Likely important for redirecting the cells physiology to overcome inhibition and allow growth to resume. Study suggests possible avenues for metabolic engineering and provides comprehensive, integrated systems biology datasets Useful for future metabolic modeling, synthetic biology and strain development endeavors.

May 29, 2012

dbCAN: a web resource for automated carbohydrate-active enzymes (CAZymes) annotation
Developed a community web resource, dbCAN (http://www.bioenergycenter.org/besc/redirect/dbcan.html), to provide a capability for automated CAZyme signature domain-based annotation for any given protein dataset (e.g. proteins from a newly sequenced genome). Defined a signature domain and constructed a hidden Markov model (HMM) for every CAZyme family, which is not publically available through any existing tools.

May 25, 2012

Caldicellulosiruptor pan genomes and proteomes reveal thermophilic deconstruction of plant biomass
Celluloytic ability is linked to modular, multi-domain enzymes (not cellulosomes). Weakly cellulolytic species lost this ability through deletion of a key genomic locus. The strongly cellulolytic Caldicellulosiruptor species possess novel cellulose-binding adhesins. Biogeography influences phylogeny and synteny among co-located species, but does not predict the ability to hydrolyze crystalline cellulose.

May 25, 2012

BESC Reports Genome Sequences for Clostridium thermocellum
Access to these genome sequences will help shed light on the mechanism of action of the cellulosome and cellulose-degrading capabilities in these and other cellulolytic organisms.

May 15, 2012

Genome for the Model Plant Setaria

Sequencing and Analysis of a Reference Genome for the Model Plant Setaria
The high quality whole genome assembly of the diploid S. italica is proving valuable as a framework in assisting with the genome assembly of other polyploid sequenced grasses, notably switchgrass. Comparison between the genomes of S. italica and S. viridis, switchgrass, sorghum and rice have revealed both shared and unique properties of the independent adaptations of several grasses to life on earth. Phylogenetic analysis of five nuclear genes shows that millet and switchgrass lineages underwent independent polyploidization events, both of which occurred after the divergence of Panicum from Setaria. The completed genome sequence enables the further development of Setaria as a model plant, improves our understanding of cell wall composition, plant structure and development and traits that are pertinent to the development of biofuel crops.

May 01, 2012

Metagenome of Microbial Community Decomposing Wood Biomass
Imagery shows differential digestion of cellulose over lignin under anaerobic conditions. Identified Magnetospirillum-like bacteria as dominant community members—possible role of de-toxification. Distinct differences between decay community and other cellulose-digesting microbiomes.

April 04, 2012

High-Performance Computing Delineates Chemistry of Lignin Synthesis
The synthesis of lignin involves oxidative radical-radical coupling of monolignols. In a BESC collaboration, we used density functional theory to characterize these reactions.

January 17, 2012

Unusual “C-lignin” polymer discovered in certain plant seeds
This discovery shows the natural capability that native plants have in producing specific lignin polymers. It supports the radical coupling polymerization hypothesis. This lignin may be new source for carbon fiber manufacturing Selected as the cover story for an upcoming issue in the Proceedings of the National Academy of Sciences and as an “editor’s choice” story in an upcoming volume of Science magazine

January 11, 2012

BESC Knowledge Base (BESCKB)
This integration is the key to gain comprehensive, systems level understanding of cellular processes involved in plant biomass formation, degradation and biofuel production. Data available through BESCKB include genomic, pathways, Omics data, resequencing data, and various assay results related to biomass properties, structure and composition. Tools available through knowledgebase include annotation, visualization, comparative analysis, integration, data mining, and various navigation tools.

November 21, 2011

A fundamental discovery by University of Georgia scientists Melani Atmodjo (left) and Debra Mohnen ( (...more)

The University of Georgia (UGA) discovery changes the way scientists think about how plant cell walls are made and holds implications for biofuels
Wood and grasses, sources of lignocellulosic biomass, are enriched in the tough secondary wall – this new discovery provides a means for modifying pectin to improve the availability of carbon reserves in the plant cell wall for renewable sources of energy.

October 20, 2011

BESC investigates tension wood formation as a model for studies of reduced recalcitrance
Tension wood is formed in response to tension stress on upper side of bent stems and is characterized by many desirable feedstock properties. BESC researchers undertook a first-of-its-kind study of tension wood formation in Populus integrating phenotyping techniques such as CARS, 13C CP/MAS, py-MBMS and GPC and correlating results to recalcitrance assays. Tension wood samples were found to contain higher cellulose and lower lignin levels as well as increased % crystallinity, % total and accessible fibril surfaces, cell wall thickness and xylem cell number. Combinatorial effects of increased accessible specific surface area, localized lignin down-regulation and chemical alteration along with increased accumulation of cellulose was found to result in significantly increased enzymatic hydrolysis yields and reduced biomass recalcitrance. The present study in integration with molecular assessments is yielding new gene targets for further evaluation in the design of suitably tailored bioenergy feedstocks.

October 10, 2011

BESC Website Renovation
The BESC website (bioenergycenter.org) recently underwent a renovation to facilitate a more user friendly interface which includes several new features to highlight the science of BESC. News - highlights of recent media exposure Publications – up to date front page information on current and recently published BESC publications Partners – expanded information on BESC inventions and industrial opportunities Who’s Who – pictorial comprehensive listing of BESC management, scientists, SAB and BOD members Research – expanded sections on Biomass Formation, Deconstruction & Conversion and Characterization & Modeling science areas Video – new video interviews of BESC scientists Featured Scientist – monthly feature section on a BESC scientist Newsletter – sign up capability for web users to receive the BESC newsletter electronically or to request information Expanded and revised navigation tools to aid web users in locating just the right information they may be seeking about BESC

September 11, 2011

Salicylic acid mediates the reduced growth phenotype of plants with strong reductions in lignin content
Blocking formation of SA by a genetic knock-out (sid2) restores growth to low lignin (HCT-RNAi) Arabidopsis plants while maintaining low lignin levels. Therefore, high SA rather than low lignin is the reason for the growth defects, indicating that it is possible to engineer plants with very low lignin and good biomass yield.

August 16, 2011

Single Microbial Gene Linked to Increased Ethanol Tolerance
A mutated alcohol dehydrogenase (AdhE) with altered co-factor specificity was shown to enhance ethanol tolerance in Clostridium thermocellum, a candidate consolidated bioprocessing microbe. The simplicity of the genetic basis for this ethanol-tolerant phenotype informs rational engineering of mutant microbial strains for cellulosic ethanol production.

July 18, 2011

2011 BESC Science Retreat
The BioEnergy Science Center held their 5th Annual Science Retreat in Chattanooga, Tennessee, on July 18th - 21st 2011. The retreat was attended by over 250 members of BESC, including researchers, post-docs, students, commercialization council members, industrial affiliates, science advisory board members and members of the board of directors.

July 12, 2011

Co-Hydrolysis of Pretreated Populus Slurries to Support Development of a High-Throughput Pretreatment System
The co-hydrolysis approach can be confidently applied in high-throughput systems to screen for low recalcitrant plants, optimize enzyme formulations, determine whether lower severity pretreatments can achieve high sugar yields, and facilitate identification of enzymes that withstand inhibitors from pretreatment.

June 14, 2011

Genetic Response during Cellulose Fermentation by Clostridium thermocellum Elucidated
Under limited carbon situations growth slows to decrease metabolic burden and C. thermocellum alters gene expression to modulate the composition of cellulosomes which are released into the environment.

June 11, 2011

BIO International Convention—BESC Highlights
At the annual biotechnology gathering, BESC met with Congressional staffers and federal agency representatives, as well as state and industry representatives, to brief them on the progress of research.

June 01, 2011

Representation of a cellulosome from C thermocellum: Scaffoldin (blue). Enzymes in gray.

Self-Assembly of the Cellulosome Enzyme Complex Shows Larger Sub-units Bind More Rapidly
This study can be used to produce designer cellulosomes with enhanced or decreased binding affinity for the scaffoldin to produce improved catalysts. Demonstrates that numerical modeling can be applied successfully to large, complex biological systems.

May 16, 2011

The mathematical model allowed the inference of metabolic channels as well as a significant regulatory mechanism controlling the pathway.

Mathematical Modeling Elucidates Control of Lignin Biosynthesis
BESC researchers developed a novel model combining static flux-based analysis with Monte Carlo simulation to identify control points in the lignin biosynthetic pathway. The model, built upon experimentally generated S/G monolignol data from stem internodes of transgenic alfalfa (Medicago sativa L.) lines, predicted new metabolic channels and novel regulatory control schemes which then were confirmed experimentally (e.g., salicylic acid).

May 16, 2011

Mathematical Modeling Elucidates Control of Lignin Biosynthesis
Ratios of lignin monomers can be a significant contributor to recalcitrance of biomass to cellulosic biofuel conversion. BESC researchers developed a novel model combining static flux-based analysis with Monte Carlo simulation to identify control points in the lignin biosynthetic pathway. The model, built upon experimentally generated S/G monolignol data from stem internodes of transgenic alfalfa (Medicago sativa L.) lines, predicted new metabolic channels and novel regulatory control schemes which then were confirmed experimentally (e.g., salicylic acid). Such wall biosynthesis pathway models are indispensable tools in the rational design of feedstock crops for efficient biofuels production.

May 02, 2011

BESC Well Represented at the Symposium on Biotechnology for Fuels and Chemicals—2011
BESC researchers presented 15 posters and 9 presentations at the Symposium.

May 01, 2011

BESC Compiles Biomass Characterization Technique Reference
This includes brief descriptions of the methods, sample preparation, and the contact person.

April 15, 2011

Quantitative Proteomics Reveal the Secreted, Highly Thermostable Cellulases of Two Extremely Thermophilic Bacteria
53-57 proteins changed in abundance during the course of cellulose fermentations, including glycosidases, pectate lyases and extracellular binding proteins. Caldicellulosiruptor spp. secrete a small number of highly abundant multidomain, multifunctional glycosidases with carbohydrate binding domains. Glycosidase mixtures efficiently degraded crystalline cellulose at 85°C. Glycosidase domains and architecture vary between the two closely related species.

April 01, 2011

Microbe engineered to produce isobutanol directly from cellulose
BESC researchers engineered a native cellulose-degrading microbe, Clostridium cellulolyticum, to produce isobutanol. Demonstrating the ability to combine CBP (consolidated bioprocessing) with production of next generation biofuels.

March 01, 2011

Lignin content and composition in natural Populus variants affects sugar release
The largest recalcitrance studies of Populus natural variants. Strong negative correlation between lignin content and glucose release does not apply to biomass samples with an S/G ratio > 2.0. Certain natural variants yielded unusually high sugar yields with no pretreatment. New insight for the rational design of biomass with reduced recalcitrance and efficient biofuels production.

March 01, 2011

Genetic manipulation of lignin improves biofuel production from switchgrass
Down-regulation of a single gene reduces recalcitrance with no apparent growth defects and leads to: Increase in ethanol production by over one third. Reduction in needed severity of pre-treatment. Evidence that biofuel processing costs can be reduced by at least 20% with 300-400% lower enzyme costs.

January 01, 2011

Complete Genome Sequence for Five Anaerobic, Extremely Thermophilic, Plant Biomass-Degrading Caldicellulosiruptor Species
Whole genome sequences (WGS) now available for eight geographically diverse species. Includes three hemicellulolytic but less cellulolytic species.

January 01, 2011

Snapshot of the 2 lignin-cellulose models at the end of the Molecular Dynamics simulations

Lignin Aggregation and Precipitation onto Cellulose: A Molecular Dynamics Study
Biomass recalcitrance after acid pretreatment: lignin aggregation and reprecipitation onto cellulose. Lignin Forms Larger Interface (closer affinity) with Crystalline Cellulose vs. Semi-Crystalline.

December 30, 2010

Davison Awarded ORNL Science Communicator of the Year
UT-Battelle has given the Science Communicator Award for 2010 to Brian H. Davison, Ph.D. "For his outstanding efforts in communicating the science of biofuels research—both to scientific audiences and to the public"

October 22, 2010

Phylogenetic, Microbiological, and Glycoside Hydrolase Diversities within the Extremely Thermophilic, Plant Biomass-Degrading Genus Caldicellulosiruptor
Preliminary genome sequencing data highlighted diverse glycoside hydrolase (GH) inventories. Each Caldicellulosiruptor species is capable of deconstructing lignocellulosic biomass in unique and complementary manner.

October 22, 2010

Multiple Members of the Genera Caldicellulosiruptor are Now Sequenced
Extremely thermophilic, gram+ bacteria capable of fermenting cellulose.

October 01, 2010

Mutation of Key TF Increases Pith Cell Wall Thickness
The discovery of negative regulators of secondary wall formation in pith opens up the possibility of significantly increasing the mass of fermentable cell wall components in bioenergy crops.

June 29, 2010

BESC and Harvard Scientists Report Label-Free, Real-Time Coherent Raman Scattering Microscopy for Chemical Imaging of Biomass
Coherent Raman Scattering (CRS) Microscopy has been developed and applied for biomass imaging at NREL in collaboration with Harvard University, and this unique tool is powerful in imaging complex biological systems in situ, specifically in identifying recalcitrant features evolving in biomass with reference to their processing to biofuels.

June 01, 2010

Phylogenomic map (7a) and resulting reconstruction of networks identified by this approach (7b).

UTK-ORNL Scientists Report the Identification of Bacterial Signal Transduction Networks from Sequence Information
Probes the origins of bacterial chemotactic (navigation) response as they developed from simple signal transduction systems that control transcription. This study has revealed more than a dozen of versions of this navigation system and assigned hundreds of bacterial species to each of them.

April 01, 2010

nhaA gene overexpression confers tolerance to inhibitor acetate

Gene Discovery is a Step to Cost-Competitive Cellulosic Biofuel
Overexpression of a sodium-proton antiporter gene (nhaA) identified at ORNL confers enhanced tolerance of acetate salts (a potential inhibitor of biofuels production) in multiple microorganisms that can be employed for biofuel production, thus moving cellulosic biofuel closer to commercial viability.

April 01, 2010

A Paradigm for Strain Improvement Breaks Down Barriers for Biofuel Production
The application of classical and systems biology tools is a paradigm for industrial strain improvement. Evolved acetate resistance Z. mobilis strains resequenced. Identification and overexpression of Na+/H+ antiporter genes confers enhanced tolerance to acetate salts in yeast and Z. mobilis.

January 11, 2010

Surface Chemistry of Pretreated Poplar Changes Measured by ToF-SIMS
The relative content of xylan after DAP increased by 30% on the surface, while the relative concentration of xylose by bulk carbohydrate analysis decreased 10-fold in comparison with untreated poplar wood.

January 01, 2010

Small Scale and Automatable Compositional Analysis of Biomass
Method is based on NREL wet chemistry procedure at 1/100x. Downscaled method produces statistically identical carbohydrate results to conventional wet chemistry procedures.

December 20, 2009

New Capabilities for the Atomic Force Microscope
The dramatic increase in the information extracted from a single run provides an unlimited number of usable signals which are suitable for high resolution imaging in liquid, gases and vacuum.

December 20, 2009

R&D 100 Award for the Development of Mode-Synthesizing Atomic Force Microscope (MSAFM)
MSAFM offers a unique way of exploring the physical properties of a large variety of samples, from biological tissues to nanofabricated systems. The system can attain atomic resolution. It retains all the advantages of an AFM but possesses a new powerful feature for exploring the interior of samples.

December 18, 2009

New Isolate from YNP Displays Rapid Hydrolysis of Plant Biomass at High Temperatures
A novel isolate from the genus Caldicellulosiruptor was isolated and displayed rapid growth on crystalline cellulose and pretreated switchgrass at 80°C.

June 01, 2009

Mycorrhizae inoculation increases switchgrass biomass under drought
Symbiotic microbes hold great potential for bioenergy crop production, or any agronomic crop, grown under a low-input regime.

BioEnergy Science Center one of three DOE Bioenergy Research Centers established by the U.S. Department of Energy.