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Hynninen A.-P.,National Bioenergy Center | Matthews J.F.,National Renewable Energy Laboratory | Beckham G.T.,National Bioenergy Center | Beckham G.T.,Colorado School of Mines | And 2 more authors.
Journal of Chemical Theory and Computation | Year: 2011

We present a coarse-grain (CG) simulation model for aqueous solutions of β-d-glucose, cellobiose, and cellotetraose, based on atomistic simulation data for each system. In the model, three spherical beads are used to represent glucose, and a single bead is used to represent water. For glucose, the force field is calculated using force matching by minimizing the sum of the square differences between forces calculated from atomistic and CG simulations. For cellobiose and cellotetraose, we use a hybrid method where the nonbonded interactions are obtained using force matching and the bonded interactions are obtained using Boltzmann inversion. We demonstrate excellent agreement in the structural properties between the atomistic simulations and the CG simulations. This model represents the first step in developing a CG force field for cellulose, as it is of significant interest to study cellulose behavior at much longer time and length scales relative to atomistic simulations. © 2011 American Chemical Society. Source


Nimlos M.R.,National Bioenergy Center | Beckham G.T.,National Bioenergy Center | Beckham G.T.,Colorado School of Mines | Matthews J.F.,Biosciences Center | And 3 more authors.
Journal of Biological Chemistry | Year: 2012

Cellulase enzymes often contain carbohydrate-binding modules (CBMs) for binding to cellulose. The mechanisms by which CBMs recognize specific surfaces of cellulose and aid in deconstruction are essential to understand cellulase action. The Family 1 CBM from the Trichoderma reesei Family 7 cellobiohydrolase, Cel7A, is known to selectively bind to hydrophobic surfaces of native cellulose. It is most commonly suggested that three aromatic residues identify the planar binding face of this CBM, but several recent studies have challenged this hypothesis. Here, we use molecular simulation to study the CBM binding orientation and affinity on hydrophilic and hydrophobic cellulose surfaces. Roughly 43 μs of molecular dynamics simulations were conducted, which enables statistically significant observations. We quantify the fractions of the CBMs that detach from crystal surfaces or diffuse to other surfaces, the diffusivity along the hydrophobic surface, and the overall orientation of the CBM on both hydrophobic and hydrophilic faces. The simulations demonstrate that there is a thermodynamic driving force for the Cel7ACBMto bind preferentially to the hydrophobic surface of cellulose relative to hydrophilic surfaces. In addition, the simulations demonstrate that the CBM can diffuse from hydrophilic surfaces to the hydrophobic surface, whereas the reverse transition is not observed. Lastly, our simulations suggest that the flat faces of Family 1 CBMs are the preferred binding surfaces. These results enhance our understanding of how Family 1CBMs interact with and recognize specific cellulose surfaces and provide insights into the initial events of cellulase adsorption and diffusion on cellulose. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc. Source


Linger J.G.,National Bioenergy Center | Adney W.S.,Chemical and Biosciences Center | Darzins A.,National Bioenergy Center | Darzins A.,National Renewable Energy Laboratory
Applied and Environmental Microbiology | Year: 2010

Development of the strategy known as consolidated bioprocessing (CBP) involves the use of a single microorganism to convert pretreated lignocellulosic biomass to ethanol through the simultaneous production of saccharolytic enzymes and fermentation of the liberated monomeric sugars. In this report, the initial steps toward achieving this goal in the fermentation host Zymomonas mobilis were investigated by expressing heterologous cellulases and subsequently examining the potential to secrete these cellulases extracellularly. Numerous strains of Z. mobilis were found to possess endogenous extracellular activities against carboxymethyl cellulose, suggesting that this microorganism may harbor a favorable environment for the production of additional cellulolytic enzymes. The heterologous expression of two cellulolytic enzymes, E1 and GH12 from Acidothermus cellulolyticus, was examined. Both proteins were successfully expressed as soluble, active enzymes in Z. mobilis although to different levels. While the E1 enzyme was less abundantly expressed, the GH12 enzyme comprised as much as 4.6% of the total cell protein. Additionally, fusing predicted secretion signals native to Z. mobilis to the N termini of E1 and GH12 was found to direct the extracellular secretion of significant levels of active E1 and GH12 enzymes. The subcellular localization of the intracellular pools of cellulases revealed that a significant portion of both the E1 and GH12 secretion constructs resided in the periplasmic space. Our results strongly suggest that Z. mobilis is capable of supporting the expression and secretion of high levels of cellulases relevant to biofuel production, thereby serving as a foundation for developing Z. mobilis into a CBP platform organism. © 2010, American Society for Microbiology. Source


Yang S.,National Bioenergy Center | Guarnieri M.T.,National Bioenergy Center | Smolinski S.,National Renewable Energy Laboratory | Ghirardi M.,National Renewable Energy Laboratory | Pienkos P.T.,National Bioenergy Center
Biotechnology for Biofuels | Year: 2013

Background: Microalgae can make a significant contribution towards meeting global renewable energy needs in both carbon-based and hydrogen (H2) biofuel. The development of energy-related products from algae could be accelerated with improvements in systems biology tools, and recent advances in sequencing technology provide a platform for enhanced transcriptomic analyses. However, these techniques are still heavily reliant upon available genomic sequence data. Chlamydomonas moewusii is a unicellular green alga capable of evolving molecular H2 under both dark and light anaerobic conditions, and has high hydrogenase activity that can be rapidly induced. However, to date, there is no systematic investigation of transcriptomic profiling during induction of H2 photoproduction in this organism. Results: In this work, RNA-Seq was applied to investigate transcriptomic profiles during the dark anaerobic induction of H2 photoproduction. 156 million reads generated from 7 samples were then used for de novo assembly after data trimming. BlastX results against NCBI database and Blast2GO results were used to interpret the functions of the assembled 34,136 contigs, which were then used as the reference contigs for RNA-Seq analysis. Our results indicated that more contigs were differentially expressed during the period of early and higher H2 photoproduction, and fewer contigs were differentially expressed when H2-photoproduction rates decreased. In addition, C. moewusii and C. reinhardtii share core functional pathways, and transcripts for H 2 photoproduction and anaerobic metabolite production were identified in both organisms. C. moewusii also possesses similar metabolic flexibility as C. reinhardtii, and the difference between C. moewusii and C. reinhardtii on hydrogenase expression and anaerobic fermentative pathways involved in redox balancing may explain their different profiles of hydrogenase activity and secreted anaerobic metabolites. Conclusions: Herein, we have described a workflow using commercial software to analyze RNA-Seq data without reference genome sequence information, which can be applied to other unsequenced microorganisms. This study provided biological insights into the anaerobic fermentation and H2 photoproduction of C. moewusii, and the first transcriptomic RNA-Seq dataset of C. moewusii generated in this study also offer baseline data for further investigation (e.g. regulatory proteins related to fermentative pathway discussed in this study) of this organism as a H 2-photoproduction strain. © 2013 Yang et al.; licensee BioMed Central Ltd. Source


Habas S.E.,National Bioenergy Center | Baddour F.G.,National Renewable Energy Laboratory | Ruddy D.A.,National Renewable Energy Laboratory | Nash C.P.,National Bioenergy Center | And 4 more authors.
Chemistry of Materials | Year: 2015

Metal phosphides have been identified as a promising class of materials for the catalytic upgrading of bio-oils, which are renewable and potentially inexpensive sources for liquid fuels. Herein, we report the facile synthesis of a series of solid, phase-pure metal phosphide nanoparticles (NPs) (Ni2P, Rh2P, and Pd3P) utilizing commercially available, air-stable metal-phosphine complexes in a one-pot reaction. This single-source molecular precursor route provides an alternative method to access metal phosphide NPs with controlled phases and without the formation of metal NP intermediates that can lead to hollow particles. The formation of the Ni2P NPs was shown to proceed through an amorphous Ni-P intermediate, leading to the desired NP morphology and metal-rich phase. This low-temperature, rapid route to well-defined metal NPs is expected to have broad applicability to a variety of readily available or easily synthesized metal-phosphine complexes with high decomposition temperatures. Hydrodeoxygenation of acetic acid, an abundant bio-oil component, was performed to investigate H2 activation and deoxygenation pathways under conditions that are relevant to ex situ catalytic fast pyrolysis (high temperatures, low pressures, and near-stoichiometric H2 concentrations). The catalytic performance of the silica-supported metal phosphide NPs was compared to the analogous incipient wetness (IW) metal and metal phosphide catalysts over the range 200-500°C. Decarbonylation was the primary pathway for H2 incorporation in the presence of all of the catalysts except NP-Pd3P, which exhibited minimal productive activity, and IW-Ni, which evolved H2. The highly controlled NP-Ni2P and NP-Rh2P catalysts, which were stable under these conditions, behaved comparably to the IW-metal phosphides, with a slight shift to higher product onset temperatures, likely due to the presence of surface ligands. Most importantly, the NP-Ni2P catalyst exhibited H2 activation and incorporation, in contrast to IW-Ni, indicating that the behavior of the metal phosphide is significantly different from that of the parent metal, and more closely resembles that of noble metal catalysts. © 2015 American Chemical Society. Source

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