Author: Paul William Wolski
Publisher:
ISBN:
Category :
Languages : en
Pages : 71
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Abstract Engineering and Identification of Ionic Liquid-Tolerant Cellulases for Biofuels Production by Paul William Wolski Doctor of Philosophy in Comparative Biochemistry University of California, Berkeley Professor Douglas S. Clark, Chair Cellulose for biofuels production presents both a great opportunity, in that cellulose is the world's most abundant source of organic material, but also a great challenge in that cellulosic material is highly crystalline and very recalcitrant to degradation. Cellulose is composed of glucose, and this glucose can serve as fermentation feedstock to biofuels in processes that are very well developed. This glucose could also be converted to hydrocarbons similar to diesel for easier commercial adoption. Before cellulase enzymes can degrade the cellulosic biomass, the biomass generally must be preteated to make the cellulose more accessible to the enzymes (i.e. less crystalline). Typically this involves acid or base treatment that only moderately affects the cellulose. Ionic liquids (ILs), which are organic salts that are liquid at or near room temperature, have the ability to dissolve cellulose by disrupting the hydrogen-bonding network that makes cellulose so strong. Ideally the cellulase enzymes would be active against dissolved cellulose. The theme of this research has been to combine the pretreatment step with the enzymatic hydrolysis step. The design of the research was as follows: Identify ionic liquids that can dissolve cellulose, while still supporting enzymatic activity and enzymes from nature that can withstand high ionic liquid concentrations. Then, use directed evolution to enhance the ionic liquid tolerance of cellulases and screen for variants that were indeed more IL-tolerant. Additionally, it was of interest to determine what happens to the enzymes, when inactivated by the ionic liquid. Do they unfold? Does the ionic liquid block the active site? All of these main objectives were achieved, to varying degrees, in this work. First, using GFP as a reporter protein for quickly measuring protein stability by GFP fluorescence, we identified the ionic liquid 1,3-Dimethylimidazolium dimethylphosphate (Mmim DMP) to support greater cellulase activity than other ionic liquids, including the more commonly used 1-Ethyl-3-methylimidazolium (Emim) acetate. Then, we found cellulases from hyperthermophiles, such as Pyrococcus furiosus, to be more stable in aqueous ionic liquid than cellulases from mesophilic organisms. A cellulase from this organism was active in up to 70% (w/w) Mmim DMP. Using DNA shuffling we generated a library of chimeric cellulase (cellobiohydrolase I or Cel7A) genes from several homologous genes. After screening a library of over 1200 variants, we identified two variants that were more stable than the native enzyme from Talaromyces emersonii. However, the degree of increase in stability was much less after both the wild type and variant enzyme were treated with exogenous glutamine cyclase to convert the N terminal glutamine to pyroglutamate. This post-translational modification occurs in T. emersonii. However, when the wild type and variant enzymes were expressed in Saccharomyces cerevisiae, this modification did not occur, and was confirmed via differential scanning calorimetry. We also used differential scanning calorimetry to determine that the ionic liquid Mmim DMP lowers the melting temperature of the enzyme, in some cases, below the assay temperature. For this reason, we concluded that the ionic liquid, in conjunction with the assay temperature, is working to inactivate the cellulase via global unfolding of the enzyme. In this work we showed that although enzymatic hydrolysis against dissolved cellulose, was not achieved, we successfully enhanced ionic liquid tolerance of cellulases via directed evolution and by selection of cellulases from extremophiles. We demonstrated a strong correlation between ionic liquid tolerance and thermotolerance. Finally, we confirmed that in directed evolution the winning variants are based directly off the screen used. In this case, evolving cellulases in a system that does not perform the native post-translational modification will not necessarily produce the same results in one that does. A future study using the same screening system thus should involve either the exogenous addition of the glutamine cyclase enzyme or endogenous production of the glutamine cyclase to make the screen as close to the production of the native host as possible.
Author: Pedro Lozano
Publisher: Academic Press
ISBN: 032391425X
Category : Science
Languages : en
Pages : 562
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Book Description
Biocatalysis in Green Solvents offers a pragmatic overview and instruction in biocatalysis and enzymology of green solvents for sustainable industries and medicine, running from concept to application. Here, international experts in the field discuss structure-function relationships of enzymes in ionic liquids (ILs) and examine how enzymes act as selective catalysts for fine biochemical synthesis in non-aqueous environments. Several integral green biochemical processes of biocatalytic transformation and pure product separation are described in detail. Application focused chapters discuss the role of biocatalysis in creating and implementing deep eutectic solvents, biomass derived solvents, sub and supercritical fluids, carbon dioxide biphasic systems, and enzymatic membrane reactors, as well as applying these biocatalytic processes in drug discovery and production. Examines the structure-function relationships of enzymes in ionic liquids and biocatalytic processes for various applications across industry and medicine Includes clear instruction in core green biochemical processes—of (bio)catalytic transformation—and pure product separation Features chapter contributions from international experts across academia and industry
Author:
Publisher:
ISBN:
Category :
Languages : en
Pages :
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Some ionic liquids (ILs) have been shown to be very effective solvents for biomass pretreatment. It is known that some ILs can have a strong inhibitory effect on fungal cellulases, making the digestion of cellulose inefficient in the presence of ILs. The identification of IL-tolerant enzymes that could be produced as a cellulase cocktail would reduce the costs and water use requirements of the IL pretreatment process. Due to their adaptation to high salinity environments, halophilic enzymes are hypothesized to be good candidates for screening and identifying IL-resistant cellulases. Using a genome-based approach, we have identified and characterized a halophilic cellulase (Hu-CBH1) from the halophilic archaeon, Halorhabdus utahensis. Hu-CBH1 is present in a gene cluster containing multiple putative cellulolytic enzymes. Sequence and theoretical structure analysis indicate that Hu-CBH1 is highly enriched with negatively charged acidic amino acids on the surface, which may form a solvation shell that may stabilize the enzyme, through interaction with salt ions and/or water molecules. Hu-CBH1 is a heat tolerant haloalkaliphilic cellulase and is active in salt concentrations up to 5 M NaCl. In high salt buffer, Hu-CBH1 can tolerate alkali (pH 11.5) conditions and, more importantly, is tolerant to high levels (20percent w/w) of ILs, including 1-allyl-3-methylimidazolium chloride ([Amim]Cl). Interestingly, the tolerances to heat, alkali and ILs are found to be salt-dependent, suggesting that the enzyme is stabilized by the presence of salt. Our results indicate that halophilic enzymes are good candidates for the screening of IL-tolerant cellulolytic enzymes.
Author: Neha Srivastava
Publisher: Elsevier
ISBN: 0444642242
Category : Technology & Engineering
Languages : en
Pages : 314
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Book Description
New and Future Developments in Microbial Biotechnology and Bioengineering: From Cellulose to Cellulase: Strategies to Improve Biofuel Production outlines new methods for the industrial production of the cellulose enzyme. The book compares the various processes for the production of biofuels, including the cost of cellulose production and availability. Biofuels are considered to be the main alternatives to fossil fuels in reducing environmental pollution and climate change. Currently, all existing biofuel production is suffering because of the high costs of production processes. As a result, cost effective practical implementation is needed to make this a viable energy alternative. Introduces new and innovative strategies for cellulase enzyme production at industrial scale Provides sustainable approaches to produce cellulase at low cost Covers all aspect and possible factors for economical, low cost, cellulase mediated biofuels production
Author: Geoffrey M. Gadd
Publisher: Academic Press
ISBN: 012820706X
Category : Science
Languages : en
Pages : 180
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Book Description
Advances in Applied Microbiology, Volume 111, continues the comprehensive reach of this widely read and authoritative review source in microbiology. Users will find invaluable references and information on a variety of areas relating to the topic. Contains contributions from leading authorities in the field Informs and updates on all the latest developments in the field of microbiology Includes discussions on the role of specific molecules in pathogen life stages and interactions, and much more
Author: Surajit Mondal
Publisher: John Wiley & Sons
ISBN: 1394174780
Category : Science
Languages : en
Pages : 470
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Book Description
CLEAN and RENEWABLE ENERGY PRODUCTION According to the World Renewable Energy Council (WREC), by the year 2100, the world’s population will increase to 12 billion and the worldwide energy demand will increase steeply to about five times the present scenario. Researchers are striving to find alternative forms of energy, and this quest is strongly forced by the increasing worry over climate change and planetary heating. Among the diverse varieties of alternative energy sources, biomass has the singular advantage of being carbon neutral. The carbon that is discharged to the atmosphere during its exercise is read back during the utilization of biomass resources for energy output. Currently, biomass provides approximately 13% of the world’s primary energy supply and more than 75% of global renewable energy. Indeed, it is estimated that bioenergy could contribute 25–33% of the global energy supply by 2050. Continued adoption of biomass will require efficient conversion rates and avoidance of competition with food and fibers. This book focuses on the recent practices in clean energy and renewable energy. The contributors highlight how newer technologies are reducing the dependency on non-renewable resources, benefiting the researchers who are working in the area of clean and renewable energy production. This new volume will also benefit mechanical engineers, electrical engineers, and bioengineers as they will be updated with the recent work progressing all over the globe. It will benefit the professionals working in the renewable energy sector such as solar, wind, hydrothermal, hydrogen, and bioenergy, including professors, research scholars, industry professionals, and students working in this field.
Author: Telma Encarnação
Publisher: Springer Nature
ISBN: 3031172264
Category : Science
Languages : en
Pages : 273
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Book Description
Marine environments represent an underexplored source for numerous biotechnological applications. Of particular interest are organisms that can provide various valuable molecules and are potential candidates for bioremediation strategies. Fungi, algae, bacteria, yeasts, and sponges are some unique resources in marine ecosystems. But these must be preserved and protected from irreversible damage. Sustainable exploitation through farming systems is the alternative to prevent pressure on harvesting wild marine organisms. Written by an international team of experts, this book provides a broad overview of the possible approaches and technologies that can be applied in bioremediation processes and the possibilities to add value to the biomass produced. It provides a comprehensive state-of-the-art of current research and practice in bioremediation technology and bio-based materials. New processing technologies, and recent technical advances in molecular biology such as gene mining, omics techniques, and metabolic engineering are highlighted. The exciting possibilities that artificial intelligence can bring to the future of the biotechnology industry are also approached. The multidisciplinary nature of this book makes it of interest to a wide range of readers, including researchers, students, consulting professionals, engineers, governmental entities, and institutions working in environmental biotechnology, pollution control and prevention, and chemical processes.
Author: Nicholas Korres
Publisher: Routledge
ISBN: 1136489649
Category : Technology & Engineering
Languages : en
Pages : 473
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Book Description
Interest in anaerobic digestion (AD), the process of energy production through the production of biogas, has increased rapidly in recent years. Agricultural and other organic waste are important substrates that can be treated by AD. This book is one of the first to provide a broad introduction to anaerobic digestion and its potential to turn agricultural crops or crop residues, animal and other organic waste, into biomethane. The substrates used can include any non-woody materials, including grass and maize silage, seaweeds, municipal and industrial wastes. These are all systematically reviewed in terms of their suitability from a biological, technical and economic perspective. In the past the technical competence and high capital investment required for industrial-scale anaerobic digesters has limited their uptake, but the authors show that recent advances have made smaller-scale systems more viable through a greater understanding of optimising bacterial metabolism and productivity. Broader issues such as life cycle assessment and energy policies to promote AD are also discussed.
Author: João Lucio de Azevedo
Publisher: Springer
ISBN: 3319558048
Category : Science
Languages : en
Pages : 441
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Book Description
This book addresses the diversity of tropical microorganisms and its applications in agriculture, renewable energy production and environmental protection. It covers several tropical habitats such as rain forests, mangroves, sea and river waters and describes how microorganisms isolated from these regions can be used to control insects and plant diseases, to improve sugar cane and biofuels production among other applications. The book also aims to bring researchers’ attention to the potential of tropical microorganisms for biotechnological purposes, an area that is still far from being well explored.
Author: Ligia R. Rodrigues
Publisher: Frontiers Media SA
ISBN: 2889740374
Category : Science
Languages : en
Pages : 161
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