Biocatalysis

Andreas S. Bommarius obtained his diploma in Chemistry from the Technical Universtiy in Munich, Germany, and, in 1989, his PhD in Chemical Engineering from Massachusetts Institute of Technology (MIT) in Cambridge, MA,USA. In 1990, he joined Degussa as head of the Biocatalysis Laboratory and pilot plant. Since 2000 he is affiliated at Georgia Institute of Technology in Atlanta, Georgia, USA, as professor in the Schools of Chemical and Biomolecular Engineering and Chemistry and Biochemistry. His main research interests are novel redox enzyme systems, stability of protein biocatalysts, as well as novel catalysts through directed evolution. Bettina R. Riebel obtained her diploma in Biology at the University of Cologne, Germany, and her PhD in Biochemistry at the University of Dusseldorf, Germany. She then occupied postdoctoral associate positions with Roche Diagnostics (then Boehringer Mannheim) in Penzberg and at the Research Center Julich. As of 2000, she is part of the Institute of Pathology in the School of Medicine at Emory University in Atlanta, Georgia, USA, where her main research interests are in host-pathogen relationships, and specifically in tyrosine kinases and cell signaling.

• Preface vAcknowledgments vii1 Introduction to Biocatalysis 11.1 Overview:The Status of Biocatalysis at the Turn of the 21st Century 21.2 Characteristics of Biocatalysis as a Technology 61.3 Current Penetration of Biocatalysis 111.4 The Breadth of Biocatalysis 142 Characterization of a (Bio-)catalyst 192.1 Characterization of Enzyme Catalysis 202.2 Sources and Reasons for the Activity of Enzymes as Catalysts 232.3 Performance Criteria for Catalysts, Processes, and Process Routes 303 Isolation and Preparation of Microorganisms 433.1 Introduction 443.2 Screening of New Enzyme Activities 463.3 Strain Development 483.4 Extremophiles 523.5 Rapid Screening of Biocatalysts 564 Molecular Biology Tools for Biocatalysis 614.1 Molecular Biology Basics: DNA versus Protein Level 624.2 DNA Isolation and Purification 654.3 Gene Isolation, Detection, and Verification 674.4 Cloning Techniques 774.5 (Over)expression of an Enzyme Function in a Host 815 Enzyme Reaction Engineering 915.1 Kinetic Modeling: Rationale and Purpose 925.2 The Ideal World: Ideal Kinetics and Ideal Reactors 945.3 Enzymes with Unfavorable Binding: Inhibition 975.4 Reactor Engineering 1055.5 Enzyme Reactions with Incomplete Mass Transfer: Influence of Immobilization 1135.6 Enzymes with Incomplete Stability: Deactivation Kinetics 1195.7 Enzymes with Incomplete Selectivity: E-Value and its Optimization 1266 Applications of Enzymes as Bulk Actives: Detergents, Textiles, Pulp and Paper, Animal Feed 1356.1 Application of Enzymes in Laundry Detergents 1366.2 Enzymes in the Textile Industry: Stone-washed Denims, Shiny Cotton Surfaces 1406.3 Enzymes in the Pulp and Paper Industry: Bleaching of Pulp with Xylanases or Laccases 1456.4 Phytase for Animal Feed: Utilization of Phosphorus 1527 Application of Enzymes as Catalysts: Basic Chemicals, Fine Chemicals, Food, Crop Protection, Bulk Pharmaceuticals 1597.1 Enzymes as Catalysts in Processes towards Basic Chemicals 1607.2 Enzymes as Catalysts in the Fine Chemicals Industry 1707.3 Enzymes as Catalysts in the Food Industry 1877.4 Enzymes as Catalysts towards Crop Protection Chemicals 1957.5 Enzymes for Large-Scale Pharma Intermediates 1978 Biotechnological Processing Steps for Enzyme Manufacture 2098.1 Introduction to Protein Isolation and Purification 2108.2 Basics of Fermentation 2128.3 Fermentation and its Main Challenge: Transfer of Oxygen 2188.4 Downstream Processing: Crude Purification of Proteins 2238.5 Downstream Processing: Concentration and Purification of Proteins 2318.6 Examples of Biocatalyst Purification 2379 Methods for the Investigation of Proteins 2439.1 Relevance of Enzyme Mechanism 2449.2 Experimental Methods for the Investigation of an Enzyme Mechanism 2459.3 Methods of Enzyme Determination 2539.4 Enzymatic Mechanisms: General Acid–Base Catalysis 2589.5 Nucleophilic Catalysis 2619.6 Electrophilic catalysis 26910 Protein Engineering 28110.1 Introduction: Elements of Protein Engineering 28210.2 Methods of Protein Engineering 28310.3 Glucose (Xylose) Isomerase (GI) and Glycoamylase: Enhancement of Thermostability 28910.4 Enhancement of Stability of Proteases against Oxidation and Thermal Deactivation 29310.5 Creating New Enzymes with Protein Engineering 29510.6 Dehydrogenases, Changing Cofactor Specificity 29810.7 Oxygenases 30010.8 Change of Enantioselectivity with Site-Specific Mutagenesis 30210.9 Techniques Bridging Different Protein Engineering Techniques 30311 Applications of Recombinant DNA Technology: Directed Evolution 30911.1 Background of Evolvability of Proteins 31011.2 Process steps in Directed Evolution: Creating Diversity and Checking for Hits 31411.3 Experimental Protocols for Directed Evolution 31911.4 Successful Examples of the Application of Directed Evolution 32511.5 Comparison of Directed Evolution Techniques 33112 Biocatalysis in Non-conventional Media 33912.1 Enzymes in Organic Solvents 34012.2 Evidence for the Perceived Advantages of Biocatalysts in Organic Media 34112.3 State of Knowledge of Functioning of Enzymes in Solvents 34412.4 Optimal Handling of Enzymes in Organic Solvents 35112.5 Novel Reaction Media for Biocatalytic Transformations 35512.6 Solvent as a Parameter for Reaction Optimization (“Medium Engineering”) 36613 Pharmaceutical Applications of Biocatalysis 37313.1 Enzyme Inhibition for the Fight against Disease 37413.2 Enzyme Cascades and Biology of Diseases 38013.3 Pharmaceutical Applications of Biocatalysis 39313.4 Applications of Specific Biocatalytic Reactions in Pharma 40214 Bioinformatics 41314.1 Starting Point: from Consequence (Function) to Sequence 41414.2 Bioinformatics: What is it, Why do we Need it, and Why Now? (NCBI Homepage) 41514.3 Tools of Bioinformatics: Databases, Alignments, Structural Mapping 41814.4 Applied Bioinformatics Tools, with Examples 42214.5 Bioinformatics for Structural Information on Enzymes 42914.6 Conclusion and Outlook 43115 Systems Biology for Biocatalysis 43315.1 Introduction to Systems Biology 43415.2 Genomics, Proteomics, and other -omics 43515.3 Technologies for Systems Biology 43815.4 Metabolic Engineering 44916 Evolution of Biocatalytic Function 45716.1 Introduction 45816.2 Search Characteristics for Relatedness in Proteins 46116.3 Evolution of New Function in Nature 46616.4 α/β-Barrel Proteins as a Model for the Investigation of Evolution 47417 Stability of Proteins 48717.1 Summary: Protein Folding, First-Order Decay, Arrhenius Law 48817.2 Two-State Model: Thermodynamic Stability of Proteins (Unfolding) 49117.3 Three-State Model: Lumry–Eyring Equation 49317.4 Four-State Model: Protein Aggregation 49617.5 Causes of Instability of Proteins: ∆G < 0, γ(t), A 50117.6 Biotechnological Relevance of Protein Folding: Inclusion Bodies 50517.7 Summary: Stabilization of Proteins 50618 Artificial Enzymes 51118.1 Catalytic Antibodies 51218.2 Other Proteinaceous Catalysts: Ribozymes and Enzyme Mimics 52118.3 Design of Novel Enzyme Activity: Enzyme Models (Synzymes) 52318.4 Heterogenized/Immobilized Chiral Chemical Catalysts 52618.5 Tandem Enzyme Organometallic Catalysts 53219 Design of Biocatalytic Processes 53919.1 Design of Enzyme Processes: High-Fructose Corn Syrup (HFCS) 54019.2 Processing of Fine Chemicals or Pharmaceutical Intermediates in an Enzyme Membrane Reactor 54919.3 Production of Enantiomerically Pure Hydrophobic Alcohols: Comparison of Different Process Routes and Reactor Configurations 55620 Comparison of Biological and Chemical Catalysts for Novel Processes 56920.1 Criteria for the Judgment of (Bio-)catalytic Processes 57020.2 Position of Biocatalysis in Comparison to Chemical Catalysts for Novel Processes 57520.3 Pathway Engineering through Metabolic Engineering 586Index 593

"...the book is excellent and could be read cover to cover or used for reference and I strongly recommend it to anyone interested in the field of biocatalysis whether they be graduate students just entering the field or more experienced practitioners."Organic Process Research & Development"This guidebook is warmly recommended to scientists in academia, industry and authorities engaged in biocatalysis because one unique feature of primary importance is that the interdisciplinary fields of chemistry, biology and bioengineering receive equal attention, thus, addressing both practioners and students from all three areas."AFS"I strongly recommend it to advanced students, experienced chemists, biologists, and engineers interested in or actively working in biocatalysis, as it is a rich source of information and offers an excellent opportunity to get an insight into neighbouring areas of expertise." Angewandte Chemie I.E."The content of the book is excellent...the book is excellent and could be read cover to cover or used for reference and I strongly recommend it to anyone interested in the field of biocatalysis..."Organic Process Research & Development"With their book "Biocatalysis" Bommarius and Riebel successfully bridge the gap between textbooks and original research papers. The book is recommended for advanced students, experienced chemists and engineers interested in the area of industrial biocatalysis."CHEMBIOCHEM"...a timely and detailed summary of the important recent developments in the rapidly moving fields pertinent to harnessing the efficiency and specificity of biological catalysts. It will definitely assist research scientists' efforts to improve their own chemical transformations."Synthesis "…this is an excellent guide that should be read by everyone in the field." (Journal of Natural Products, January 2005) "The book nicely brings together the essentials of biocatalysis including reactions, the products and processes that utilize the methodology, and techniques for improving biocatalysts." (Catalyst, November 2004)