Download or read book Studies of Thermobifida Fusca Cellulases Cel9A and Cel48A Using a Novel Kinetic Modeling Approach for the Enzymatic Digestion of Cellulose written by Maxim Kostylev and published by . This book was released on 2013 with total page 137 pages. Available in PDF, EPUB and Kindle. Book excerpt: Lignocellulosic biomass is a potential source of sustainable transportation fuels, but efficient enzymatic saccharification of cellulose is a key challenge in its utilization. Enzymatic digestion of cellulose is a complex, heterogeneous process. An enzyme must repeatedly take multiple steps to hydrolyze the substrate to product. In addition, most bulk cellulose contains crystalline, semi-crystalline, and amorphous fractions, whose ratio likely changes during digestion. Access to substrate, rather than hydrolysis, is widely accepted to be the rate limiting step, as we showed experimentally using Cel9A, and the rate of digestion rapidly and continuously drops off as digestion proceeds. Predictive kinetic models typically incorporate different parameters and constants to account for the possible factors responsible for such behavior. Cellulose hydrolysis by individual cellulases and their mixtures can be modeled with a simple two-parameter model based on a modified classical kinetics scheme. Analogous to a fractal kinetics approach, the specific activity constant is replaced with a time-dependent activity coefficient in order to account for the continuous decrease in the digestion rate. The parameter that quantifies the time dependence of the digestion rate is an intrinsic constant for a given cellulase or mixture on a given substrate. The developed kinetic model was utilized in studies aimed to understand the function of aromatic residues located near the active site tunnel entrance of Cel48A and to develop a distinct model of synergistic cooperation between Cel48A and Cel9A. Additional experiments were carried out to establish experimentally the catalytic base in family 48 glycoside hydrolases.
Download or read book Mechanistic Studies of Thermobifida Fusca Exocellulase Cel6B written by Thu Van Vuong and published by . This book was released on 2010 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Understanding the mechanism by which cellulases catalyze cellulose hydrolysis can greatly contribute to the development of biofuels. The thermophilic bacterium Thermobifida fusca, a major degrader of plant cell walls in certain environments, secretes seven different cellulases including exocellulase Cel6B. This cellulase acts by an inverting mechanism; however, its catalytic acid and base residues had not been identified. Biochemical approaches confirmed D274 to be the catalytic acid residue. A single catalytic base residue could not be determined, as sodium azide assays showed no activity rescue for any single mutations of candidate residues. However, a double mutation of D226A and S232A knocked out enzymatic activity and its activity was partially rescued by sodium azide. We therefore propose a novel hydrolysis mechanism for T. fusca Cel6B involving a proton-transferring network to carry out the catalytic base function. T. fusca exocellulase Cel6B was also engineered to gain knowledge on the relationship between processivity and synergism as these properties are important for hydrolyzing crystalline cellulose. Mutations of several residues in the active site tunnel of Cel6B gave higher processivity. This improvement was confirmed by two assays: the ratio of soluble/insoluble reducing sugars as well as the ratio of oligosaccharide products. Surprisingly, the mutant enzyme, which has the highest processivity, showed the least synergism in mixtures with endocellulases, suggesting that improving exocellulase processivity might not always be an effective strategy for producing improved cellulase mixtures for biomass conversion. The highly processive Cel6B mutant enzymes were successfully fluorescently labeled, so these species can be used to visualize binding and track their movement on cellulose. The catalytic domains of Cel6B was found to bind non-productively to other polysaccharides; therefore, the balance between specific binding and non-specific adsorption should be always considered when engineering cellulases for hydrolyzing complex substrates. Using immuno-precipitation, Cel6B was demonstrated to contribute greatly to the hydrolysis of crystalline cellulose by T. fusca.
Download or read book Studies of Thermobifida Fusca Lytic Polysaccharide Monooxygenases AA10A and AA10B for Cleavage of Recalcitrant Cellulose written by Nathan Kruer-Zerhusen and published by . This book was released on 2016 with total page 274 pages. Available in PDF, EPUB and Kindle. Book excerpt: Cellulose recalcitrance remains a barrier to producing sustainable fuels and chemicals from renewable plant biomass. The cellulolytic bacterium Thermobifida fusca secretes lytic polysaccharide monooxygenases (LPMOs) to contribute to cellulose breakdown. LPMOs use an oxidative mechanism to synergize with hydrolytic cellulases. LPMOs may play an important role in overcoming cellulose recalcitrance, the physical properties that slow cellulose breakdown. Understanding LPMO structure and function will inform the optimization of industrial cellulase mixtures for use in biomass saccharification. The goal of my work is to explore the relationship between LPMO structure and oxidative cleavage using site-directed mutagenesis, and to determine if the mechanism of LPMO-cellulase synergism relies on changes to cellulose crystallinity. Mutations to residues on the AA10A binding surface change both activity and binding properties, confirming the histidine brace for chelation of copper to be essential for activity. In addition to surface residues that mediate substrate interaction, the conserved cellulose binding domain was shown to be critical for activity of TfAA10B on crystalline cellulose. Kinetics of LPMO mutants were measured by performing secondary hydrolysis to simplify the product distribution to monomeric products, then quantifying using a rapid high performance liquid chromatography (HPLC) approach. The role of LPMOs was initially thought to be amorphogensis, a disruption of crystalline cellulose structure, induced by oxidative cleavages of surface chains. This proposed amorphogenesis model of LPMO action and cellulase synergism was explored using Fourier transform infrared spectroscopy (FTIR). FTIR can penetrate residual substrates to detect changes in multiple measures of crystallinity caused by digestion. While past reports had observed significant changes to cellulose crystallinity imparted by cellulases, these changes were not reproduced after optimization of the measurement approach. Similarly, LPMOs did not induce changes to cellulose crystallinity during oxidative cleavage, suggesting an alternative mechanism of LPMO synergism with cellulases.
Download or read book Biofuels written by Jonathan R. Mielenz and published by Humana Press. This book was released on 2012-03-07 with total page 294 pages. Available in PDF, EPUB and Kindle. Book excerpt: This editor believes the public finally understands that bioenergy should be a critical new 39 component of our world’s future energy supply. While this new enlightenment might have 40 come from bioenergy’s environmentally friendly attributes and its renewable and sustain- 41 able nature, the reality is that the primary driver has been both the high cost of energy, 42 led recently by petroleum products, and the unique climatic changes such as local weather 43 events and reports of melting of ancient glaciers. I believe that much of the public under- 44 stands the primary cause of these changes is the growing demand for fossil fuels for heat- 45 ing and transportation needs driven by both population increases and dramatic economic 46 development around the globe. This increased demand, as well as global politics, has 47 impacted the costs for these energy products. At the same time, increased use of fossil fuels 48 has been liberating unprecedented levels of CO and other greenhouse gasses (GHGs) 49 2 that must be impacting the global climate. As a result, the world is looking for alternative 50 solutions. Renewable energy is one of the leading options as long as it is pursued in a sus- 51 tainable fashion. Of course, renewable energy has numerous sources, including wind, solar, 52 geothermal, ocean currents/tides, and bio-based energy. The latter category of renewable 53 energy is centered primarily on use of green plant matter, i. e.
Download or read book Aqueous Pretreatment of Plant Biomass for Biological and Chemical Conversion to Fuels and Chemicals written by Charles E. Wyman and published by John Wiley & Sons. This book was released on 2013-03-27 with total page 597 pages. Available in PDF, EPUB and Kindle. Book excerpt: Plant biomass is attracting increasing attention as a sustainable resource for large-scale production of renewable fuels and chemicals. However, in order to successfully compete with petroleum, it is vital that biomass conversion processes are designed to minimize costs and maximize yields. Advances in pretreatment technology are critical in order to develop high-yielding, cost-competitive routes to renewable fuels and chemicals. Aqueous Pretreatment of Plant Biomass for Biological and Chemical Conversion to Fuels and Chemicals presents a comprehensive overview of the currently available aqueous pretreatment technologies for cellulosic biomass, highlighting the fundamental chemistry and biology of each method, key attributes and limitations, and opportunities for future advances. Topics covered include: • The importance of biomass conversion to fuels • The role of pretreatment in biological and chemical conversion of biomass • Composition and structure of biomass, and recalcitrance to conversion • Fundamentals of biomass pretreatment at low, neutral and high pH • Ionic liquid and organosolv pretreatments to fractionate biomass • Comparative data for application of leading pretreatments and effect of enzyme formulations • Physical and chemical features of pretreated biomass • Economics of pretreatment for biological processing • Methods of analysis and enzymatic conversion of biomass streams • Experimental pretreatment systems from multiwell plates to pilot plant operations This comprehensive reference book provides an authoritative source of information on the pretreatment of cellulosic biomass to aid those experienced in the field to access the most current information on the topic. It will also be invaluable to those entering the growing field of biomass conversion.
Download or read book Synthetic Metabolic Pathways written by Michael Krogh Jensen and published by Humana Press. This book was released on 2017-11-24 with total page 354 pages. Available in PDF, EPUB and Kindle. Book excerpt: This volume outlines key steps associated with the design, building, and testing of synthetic metabolic pathways for optimal cell factory performance and robustness, and illustrates how data-driven learning from these steps can be used for rational cost-effective engineering of cell factories with improved performance. Chapters are divided into four sections focusing on the four steps of the iterative design-build-test-learn cycle related to modern cell factory engineering. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Synthetic Metabolic Pathways: Methods and Protocols aims to ensure successful results in the further study of this vital field.
Download or read book Biomass Recalcitrance written by Michael Himmel and published by Wiley-Blackwell. This book was released on 2008-06-23 with total page 552 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book examines the connection between biomass structure, ultrastructure, and composition, to resistance to enzymatic deconstruction, with the aim of discovering new cost-effective technologies for biorefineries. It contains chapters on topics extending from the highest levels of biorefinery design and biomass life-cycle analysis, to detailed aspects of plant cell wall structure, chemical treatments, enzymatic hydrolysis, and product fermentation options."--Pub. desc.
Download or read book Specificity of Proteolysis written by Borivoj Keil and published by Springer Science & Business Media. This book was released on 2012-12-06 with total page 343 pages. Available in PDF, EPUB and Kindle. Book excerpt: Specificity of Proteolysis presents a survey and conclusions on the action or proteinases - enzymes which are cleaving proteins or peptides. The specificity of proteinases which is determined as the sequence of amino acids at the cleavage site of a substrate, is an important criteria to choose an enzyme as tool in protein research. Whenever one is looking for an enzyme to act at a defined site or to give defined cleavage products one will find comprehensive information in this work. Comprehensive information about more than 280 endopeptidases which are based on the database LYSIS including a calculation program to determine cleavage sites, is given in the book.
Download or read book Biomass as a Sustainable Energy Source for the Future written by Wiebren de Jong and published by John Wiley & Sons. This book was released on 2014-11-03 with total page 600 pages. Available in PDF, EPUB and Kindle. Book excerpt: Focusing on the conversion of biomass into gas or liquid fuels the book covers physical pre-treatment technologies, thermal, chemical and biochemical conversion technologies • Details the latest biomass characterization techniques • Explains the biochemical and thermochemical conversion processes • Discusses the development of integrated biorefineries, which are similar to petroleum refineries in concept, covering such topics as reactor configurations and downstream processing • Describes how to mitigate the environmental risks when using biomass as fuel • Includes many problems, small projects, sample calculations and industrial application examples
Download or read book Forest Products Biotechnology written by Alan Bruce and published by CRC Press. This book was released on 2002-09-10 with total page 337 pages. Available in PDF, EPUB and Kindle. Book excerpt: Industries are developing radical, new biotechnology processes to expand and develop their range of products that originate from the world's forests. As a result of the growing understanding of the process involved, biotechnology is also helping reduce any adverse impact on the environment.; This book presents a review of specialist research directed towards efficient and environmentally sensitive use of forests. An introductory chapter explaining the structure and anatomy of wood is followed by a chapter-by-chapter review of the most current developments on individual topics associated with a wide range of forest products such as timber, trees, pulp and paper, drugs and valuable chemicals. In addition, chapters focus on the ways of resolving some of the environmental problems faced by these industries.
Download or read book Trichoderma Reesei Cellulases written by C. P. Kubicek and published by . This book was released on 1990 with total page 228 pages. Available in PDF, EPUB and Kindle. Book excerpt: The proceedings of a symposium on Trichoderma cellulases held at the Technical University of Vienna September 14-16, 1989, attended by 100 scientists representing both academic institutions and industrial research laboratories.
Download or read book Encyclopedia of Food Science written by Johann Wells and published by . This book was released on 2015-01-09 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book provides all the basic and advanced concepts of fruit science. It includes different aspects of food science, such as medical problems associated with the food consumption of some specific types. With the ever growing need of food in order to satisfy the population, food scientists and specialists have compiled their knowledge of food science in this book.
Download or read book Proteases in Human Diseases written by Sajal Chakraborti and published by Springer. This book was released on 2017-07-13 with total page 516 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book bridges the gap between fundamental research and biomedical and pharmacological applications on proteases. It represents a comprehensive overview of the multifaceted field of proteases in cellular environment and highlights the recently elucidated functions of complex proteolytic systems in different diseases. Several established investigators have elucidated the crucial role of proteases in biological processes, including how proteolytic function and regulation can be combined to develop new strategies of therapeutic interventions. Proteases form one of the largest and most diverse families of enzymes known. It is now clear that proteases are involved in every aspect of life functions of an organism. Under physiological conditions, proteases are regulated by their endogenous inhibitors; however, when the activity of proteases is not regulated appropriately, disease processes can result in. So, there is absolute need for a stringent control of proteolytic activities in cells and tissues. Dysregulation of proteases may cause derangement of cellular signalling network resulting in different pathophysiological conditions such as vascular remodelling, atherosclerotic plaque progression, ulcer and rheumatoid arthritis, Alzheimer disease, cancer metastasis, tumor progression and inflammation. Additionally, many infective microorganisms require proteases for replication or use proteases as virulence factors, which have facilitated the development of protease-targeted therapies for a variety of parasitic diseases.
