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MODERN BIOTECHNOLOGY free pdf ebook download
Shubham Goyal

MODERN BIOTECHNOLOGY free pdf ebook download

Shubham Goyal | 04-Mar-2016 |
MODERN BIOTECHNOLOGY , Biotechnology , New Biotechnology , Bioproducts and Biofuels , Microbial Fermentations , Modeling and Simulation , Aerobic Bioreactors , Enzymes , Enzyme Kinetics , Metabolism , Biological Energetics , Metabolic Pathways , Genetic Engineering: DNA , RNA , Genes , Metabolic Engineering , Genomes and Genomics ,

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Preface xv
Acknowledgments xvii
List of Illustrations xix
1 Biotechnology 1
Introduction 1
The Directed Manipulation of Genes Distinguishes the New
Biotechnology from Prior Biotechnology 2
Growth of the New Biotechnology Industry Depends on
Venture Capital 3
Submerged Fermentations Are the Industry’s Bioprocessing
Cornerstone 10
Oil Prices Affect Parts of the Fermentation Industry 10
Growth of the Antibiotic/Pharmaceutical Industry 11
The Existence of Antibiotics Was Recognized in 1877 11
Penicillin Was the First Antibiotic Suitable for Human
Systemic Use 12
Genesis of the Antibiotic Industry 12
Other Antibiotics Were Quickly Discovered after the
Introduction of Penicillin 13
Discovery and Scaleup Are Synergistic in the Development of
Pharmaceutical Products 15
Success of the Pharmaceutical Industry in Research, Development,
and Engineering Contributed to Rapid Growth but Also
Resulted in Challenges 15
Growth of the Amino Acid/Acidulant Fermentation Industry 16
Production of Monosodium Glutamate (MSG) via Fermentation 17
The Impact of Glutamic Acid Bacteria on Monosodium
Glutamate Cost Was Dramatic 17
Auxotrophic and Regulatory Mutants Enabled Production of
Other Amino Acids 17
Prices and Volumes Are Inversely Related 19
Biochemical Engineers Have a Key Function in All Aspects of
the Development Process for Microbial Fermentation 21
References 22
Homework Problems 24
2 New Biotechnology 27
Introduction 27
Growth of the Biopharmaceutical Industry 28
The Biopharmaceutical Industry Is in the Early Part of Its
Life Cycle 31
Discovery of Type II Restriction Endonucleases Opened a New
Era in Biotechnology 33
The Polymerase Chain Reaction (PCR) Is an Enzyme-Mediated,
In Vitro Amplifi cation of DNA 33
Impacts of the New Biotechnology on Biopharmaceuticals, Genomics,
Plant Biotechnology, and Bioproducts 34
Biotechnology Developments Have Accelerated Biological
Research 35
Drug Discovery Has Benefi ted from Biotechnology Research
Tools 36
The Fusing of Mouse Spleen Cells with T Cells Facilitated
Production of Antibodies 36
Regulatory Issues Add to the Time Required to Bring a New
Product to Market 36
New Biotechnology Methods Enable Rapid Identifi cation of
Genes and Their Protein Products 39
Genomics Is the Scientifi c Discipline of Mapping, Sequencing, and
Analyzing Genomes 39
Products from the New Plant Biotechnology Are Changing the
Structure of Large Companies that Sell Agricultural Chemicals 42
Bioproducts from Genetically Engineered Microorganisms
Will Become Economically Important to the
Fermentation Industry 43
References 45
Homework Problems 47
3 Bioproducts and Biofuels 49
Introduction 49
Biocatalysis and the Growth of Industrial Enzymes 49
Glucose Isomerase Catalyzed the Birth of a New Process for
Sugar Production from Corn 51
Identifi cation of a Thermally Stable Glucose Isomerase and an
Inexpensive Inducer Was Needed for an Industrial Process 53
The Demand for High-Fructose Corn Syrup (HFCS) Resulted
in Large-Scale Use of Immobilized Enzymes and Liquid
Chromatography 53
Rapid Growth of HFCS Market Share Was Enabled by Large-
Scale Liquid Chromatography and Propelled by Record-High
Sugar Prices 55
Biocatalysts Are Used in Fine-Chemical Manufacture 56
Growth of Renewable Resources as a Source of Specialty Products
and Industrial Chemicals 58
A Wide Range of Technologies Are Needed to Reduce Costs
for Converting Cellulosic Substrates to Value-Added
Bioproducts and Biofuels 59
Renewable Resources Are a Source of Natural Plant Chemicals 63
Bioseparations Are Important to the Extraction, Recovery, and
Purifi cation of Plant-Derived Products 64
Bioprocess Engineering and Economics 65
Bioseparations and Bioprocess Engineering 66
References 67
Homework Problems 71
4 Microbial Fermentations 73
Introduction 73
Fermentation Methods 75
Fermentations Are Carried Out in Flasks, Glass Vessels,
and Specially Designed Stainless-Steel Tanks 75
Microbial Culture Composition and Classifi cation 78
Microbial Cells: Prokaryotes versus Eukaryotes 78
Classifi cation of Microorganisms Are Based on Kingdoms 81
Prokaryotes Are Important Industrial Microorganisms 81
Eukaryotes Are Used Industrially to Produce Ethanol,
Antibiotics, and Biotherapeutic Proteins 82
Wild-Type Organisms and Growth Requirements in
Microbial Culture 83
Wild-Type Organisms Find Broad Industrial Use 83
Microbial Culture Requires that Energy and All Components
Needed for Cell Growth Be Provided 86
Media Components and Their Functions (Complex and
Defi ned Media) 86
Carbon Sources Provide Energy, and Sometimes
Provide Oxygen 86
Complex Media Have a Known Basic Composition but a
Chemical Composition that Is Not Completely Defi ned 89
Industrial Fermentation Broths May Have a High Initial Carbon
(Sugar) Content (Ethanol Fermentation Example) 91
The Accumulation of Fermentation Products Is Proportional to
Cell Mass in the Bioreactor 92
A Microbial Fermentation Is Characterized by Distinct Phases
of Growth 93
Expressions for Cell Growth Rate Are Based on Doubling
Time 94
Products of Microbial Culture Are Classifi ed According to
Their Energy Metabolism (Types I, II, and III Fermentations) 96
Product Yields Are Calculated from the Stoichiometry of
Biological Reactions (Yield Coeffi cients) 102
The Embden–Meyerhof Glycolysis and Citric Acid Cycles Are
Regulated by the Relative Balance of ATP, ADP, and AMP
in the Cell 104
References 105
Homework Problems 108
5 Modeling and Simulation 111
Introduction 111
The Runge–Kutta Method 112
Simpson’s Rule 112
Fourth-Order Runge–Kutta Method 113
Ordinary Differential Equations (ODEs) 115
Runge–Kutta Technique Requires that Higher-Order Equations
Be Reduced to First-Order ODEs to Obtain Their Solution 115
Systems of First-Order ODEs Are Represented in Vector Form 116
Kinetics of Cell Growth 117
Ks Represents Substrate Concentration at Which the Specifi c
Growth Rate Is Half Its Maximum 120
Simulation of a Batch Ethanol Fermentation 122
Ethanol Case Study 123
Luedeking–Piret Model 127
Continuous Stirred-Tank Bioreactor 128
Batch Fermentor versus Chemostat 132
References 133
Homework Problems 135
6 Aerobic Bioreactors 141
Introduction 141
Fermentation Process 144
Fermentation of Xylose to 2,3-Butanediol by Klebsiella
oxytoca Is Aerated but Oxygen-Limited 144
Oxygen Transfer from Air Bubble to Liquid Is Controlled by
Liquid-Side Mass Transfer 153
Chapter 6 Appendix: Excel Program for Integration of
Simultaneous Differential Equations 159
References 161
Homework Problems 162
7 Enzymes 165
Introduction 165
Enzymes and Systems Biology 165
Industrial Enzymes 166
Enzymes: In Vivo and In Vitro 167
Fundamental Properties of Enzymes 169
Classifi cation of Enzymes 170
Sales and Applications of Immobilized Enzymes 172
Assaying Enzymatic Activity 173
Enzyme Assays 181
Batch Reactions 187
Thermal Enzyme Deactivation 187
References 192
Homework Problems 195
8 Enzyme Kinetics 199
Introduction 199
Initial Rate versus Integrated Rate Equations 200
Obtaining Constants from Initial Rate Data Is an Iterative
Process 204
Batch Enzyme Reactions: Irreversible Product Formation
(No Inhibition) 207
Rapid Equilibrium Approach Enables Rapid Formulation of
an Enzyme Kinetic Equation 207
The Pseudo-Steady-State Method Requires More Effort to Obtain
the Hart Equation but Is Necessary for Reversible Reactions 209
Irreversible Product Formation in the Presence of Inhibitors
and Activators 210
Inhibition 212
Competitive Inhibition 213
Uncompetitive Inhibition 214
(Classical) Noncompetitive Inhibition 216
Substrate Inhibition 217
Example of Reversible Reactions 220
Coenzymes and Cofactors Interact in a Reversible Manner 223
King–Altman Method 225
Immobilized Enzyme 234
Online Databases of Enzyme Kinetic Constants 236
References 237
Homework Problems 238
9 Metabolism 243
Introduction 243
Aerobic and Anaerobic Metabolism 245
Glycolysis Is the Oxidation of Glucose in the Absence of Oxygen 245
Oxidation Is Catalyzed by Oxidases in the Presence of O2,
and by Dehydrogenases in the Absence of O2 246
A Membrane Bioreactor Couples Reduction and Oxidation
Reactions (R-Mandelic Acid Example) 247
Three Stages of Catabolism Generate Energy, Intermediate
Molecules, and Waste Products 248
The Glycolysis Pathway Utilizes Glucose in Both Presence
(Aerobic) and Absence (Anaerobic) of O2 to Produce Pyruvate 249
Glycolysis Is Initiated by Transfer of a High-Energy Phosphate
Group to Glucose 250
Products of Anaerobic Metabolism Are Secreted or Processed
by Cells to Allow Continuous Metabolism of Glucose by
Glycolysis 253
Other Metabolic Pathways Utilize Glucose Under Anaerobic
Conditions (Pentose Phosphate, Entner–Doudoroff, and
Hexose Monophosphate Shunt Pathways) 255
Knowledge of Anaerobic Metabolism Enables Calculation of
Theoretical Yields of Products Derived from Glucose 257
Economics Favor the Glycolytic Pathway for Obtaining
Oxygenated Chemicals from Renewable Resources 258
Citric Acid Cycle and Aerobic Metabolism 259
Respiration Is the Aerobic Oxidation of Glucose and Other
Carbon-Based Food Sources (Citric Acid Cycle) 260
The Availability of Oxygen, under Aerobic Conditions,
Enables Microorganisms to Utilize Pyruvate via the Citric
Acid Cycle 260
The Citric Acid Cycle Generates Precursors for Biosynthesis of
Amino Acids and Commercially Important Fermentation
Products 264
Glucose Is Transformed to Commercially Valuable Products via
Fermentation Processes: A Summary 264
Essential Amino Acids Not Synthesized by Microorganisms
Must Be Provided as Nutrients (Auxotrophs) 267
The Utilization of Fats in Animals Occurs by a Non–
Tricarboxylic Acid (TCA) Cycle Mechanism 267
Some Bacteria and Molds Can Grow on Hydrocarbons or
Methanol in Aerated Fermentations (Single-Cell Protein
Case Study) 269
Extremophiles: Microorganisms that Do Not Require Glucose,
Utilize H2, and Grow at 80–100 °C and 200 atm Have
Industrial Uses 270
The Terminology for Microbial Culture Is Inexact: “Fermentation”
Refers to Both Aerobic and Anaerobic Conditions While
“Respiration” Can Denote Anaerobic Metabolism 271
Metabolism and Biological Energetics 272
References 272
Homework Problems 273
10 Biological Energetics 277
Introduction 277
Redox Potential and Gibbs Free Energy in Biochemical Reactions 277
Heat: Byproduct of Metabolism 286
References 292
Homework Problems 293
11 Metabolic Pathways 295
Introduction 295
Living Organisms Control Metabolic Pathways at Strategic and
Operational Levels 296
Auxotrophs Are Nutritionally Defi cient Microorganisms that
Enhance Product Yields in Controlled Fermentations (Relief
of Feedback Inhibition and Depression) 296
Both Branched and Unbranched Pathways Cause Feedback
Inhibition and Repression (Purine Nucleotide Example) 299
The Accumulation of an End Metabolite in a Branched Pathway
Requires a Strategy Different from that for the Accumulation
of an Intermediate Metabolite 301
Amino Acids 305
The Formulation of Animal Feed Rations with Exogeneous
Amino Acids Is a Major Market for Amino Acids 306
Microbial Strain Discovery, Mutation, Screening, and
Development Facilitated Introduction of Industrial, Aerated
Fermentations for Amino Acid Production by Corynbacterium
glutamicum 308
Overproduction of Glutamate by C. glutamicum Depends
on an Increase in Bacterial Membrane Permeability
(Biotin-Defi cient Mutant) 309
A Threonine and Methionine Auxotroph of C. glutamicum Avoids
Concerted Feedback Inhibition and Enables Industrial Lysine
Fermentations 310
Cell (Protoplast) Fusion Is a Method for Breeding Amino Acid
Producers that Incorporate Superior Characteristics of Each
Parent (Lysine Fermentation) 312
Amino Acid Fermentations Represent Mature Technologies 313
Antibiotics 314
Secondary Metabolites Formed During Idiophase Are Subject
to Catabolite Repression and Feedback Regulation
(Penicillin and Streptomycin) 314
The Production of Antibiotics Was Viewed as a Mature
Field Until Antibiotic-Resistant Bacteria Began to Appear 317
Bacteria Retain Antibiotic Resistance Even When
Use of the Antibiotic Has Ceased for Thousands
of Generations 318
Antibiotic Resistance Involves Many Genes
(Vancomycin Example) 318
References 320
Homework Problems 323
12 Genetic Engineering: DNA, RNA, and Genes 331
Introduction 331
DNA and RNA 332
DNA Is a Double-Stranded Polymer of the Nucleotides:
Thymine, Adenine, Cytosine, and Guanine 332
The Information Contained in DNA Is Huge 332
Genes Are Nucleotide Sequences that Contain the Information
Required for the Cell to Make Proteins 333
Transcription Is a Process Whereby Specifi c Regions of the
DNA (Genes) Serve as a Template to Synthesize Another
Nucleotide, Ribonucleic Acid (RNA) 333
Chromosomal DNA in a Prokaryote (Bacterium) Is
Anchored to the Cell’s Membrane While Plasmids Are in
the Cytoplasm 333
Chromosomal DNA in a Eukaryote (Yeast, Animal or Plant
Cells) Is Contained in the Nucleus 334
Microorganisms Carry Genes in Plasmids Consisting of Shorter
Lengths of Circular, Extrachromosomal DNA 334
Restriction Enzymes Enable Directed In Vitro Cleavage of
DNA 337
Different Type II Restriction Enzymes Give Different Patterns
of Cleavage and Different Single-Stranded Terminal
Sequences 339
DNA Ligase Covalently Joins the Ends of DNA Fragments 341
DNA Fragments and Genes of ≤150 Nucleotides Can Be
Chemically Synthesized if the Nucleotide Sequence Has Been
Predetermined 342
Protein Sequences Can Be Deduced and Genes Synthesized
on the Basis of Complementary DNA Obtained from
Messenger RNA 343
Genes and Proteins 344
Selectable Markers Are Genes that Facilitate Identifi cation of
Transformed Cells that Contain Recombinant DNA 344
A Second Protein Fused to the Protein Product Is Needed to
Protect the Product from Proteolysis (β-Gal-Somatostatin
Fusion Protein Example) 346
Recovery of Protein Product from Fusion Protein Requires
Correct Selection of Amino Acid that Links the Two Proteins
(Met Linker) 347
Chemical Modifi cation and Enzyme Hydrolysis Recover an
Active Molecule Containing Met Residues from a Fusion
Protein (β-Endorphin Example) 347
Metabolic Engineering Differs from Genetic Engineering by
the Nature of the End Product 348
References 349
Homework Problems 350
13 Metabolic Engineering 355
Introduction 355
Building Blocks 359
l-Threonine-Overproducing Strains of E. coli K-12 359
Genetically Altered Brevibacterium lactoferrin Has Yielded
Improved Amino Acid–Producing Strains 360
Metabolic Engineering May Catalyze Development of New
Processes for Manufacture of Oxygenated Chemicals 362
Gene Chips Enable Examination of Glycolytic and Citric
Acid Cycle Pathways in Yeast at a Genomic Level
(Yeast Genome Microarray Case Study) 362
The Fermentation of Pentoses to Ethanol Is a Goal of
Metabolic Engineering (Recombinant Bacteria and
Yeast Examples) 364
Metabolic Engineering for a 1,3-Propanediol-Producing
Organism to Obtain Monomer for Polyester Manufacture 370
Redirection of Cellular Metabolism to Overproduce an
Enzyme Catalyst Results in an Industrial Process for
Acrylamide Production (Yamada–Nitto Process) 373
References 377
Homework Problems 379
14 Genomes and Genomics 385
Introduction 385
Human Genome Project 385
Deriving Commercial Potential from Information Contained
in Genomes 388
The Genome for E. coli Consists of 4288 Genes that Code
for Proteins 390
DNA Sequencing Is Based on Electrophoretic Separations
of Defi ned DNA Fragments 391
Sequence-Tagged Sites (STSs) Determined from
Complementary DNA (cDNA) Give Locations of Genes 394
Single-Nucleotide Polymorphisms (SNPs) Are Stable Mutations
Distributed throughout the Genome that Locate Genes More
Effi ciently than Do STSs 394
Gene Chip Probe Array 398
Polymerase Chain Reaction (PCR) 401
The Polymerase Chain Reaction Enables DNA to Be
Copied In Vitro 402
Thermally Tolerant DNA Polymerase from Thermus
aquaticus Facilitates Automation of PCR 403
Only the 5'-Terminal Primer Sequence Is Needed to Amplify
the DNA by PCR 404
The Sensitivity of PCR Can Be a Source of Signifi cant
Experimental Error 405
Applications of PCR Range from Obtaining Fragments of
Human DNA for Sequencing to Detecting Genes Associated
with Diseases 405

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