Bioprocessing Technology for Production of Biopharmaceuticals and Bioproducts

Written for industrial and academic researchers and development scientists in the life sciences industry, Bioprocessing Technology for Production of Biopharmaceuticals and Bioproducts is a guide to the tools, approaches, and useful developments in bioprocessing. This important guide: Summarizes state-of-the-art bioprocessing methods and reviews applications in life science industries Includes illustrative case studies that review six milestone bio-products * Discuses a wide selection of host strain types and disruptive bioprocess technologies

Autorentext

CLAIRE KOMIVES, PHD, is a Professor in the Chemical and Materials Engineering Department at San Jose State University. Her research interests focus on the development of low cost snake antivenom compounds. WEICHANG ZHOU, PHD, is Chief Technology Officer, Biologics Development & Manufacturing at WuXi Biologics. He has previously worked at Genzyme Corporation and Merck Research Laboratories.

Klappentext
A Guide to the Latest Developments in Bioprocessing that can be Applied in both the Biopharmaceutical and Biochemical Industries Bioprocessing Technology for Production of Biopharmaceuticals and Bioproducts offers a review of some current developments in bioprocessing and puts an emphasis on the effective tools and methods. With contributions from noted experts in the field, the text contains a review of six enabling technologies that cover a broad range of scientific and engineering topics. The authors include information about the history of the process and technologies used in the development of a selection of biopharmaceuticals and bioproducts. As the authors explain, the development of new processing methods and the improvement of traditional reactors and host strains has led to recent advances in manufacturing sciences. Given these new developments, the book explores the engineering methods that can be used for large-scale applications. Bioprocessing Technology for Production of Biopharmaceuticals and Bioproducts contains the information needed to better understand both the reactor technology as well as the characteristics and specific needs of the host strains. Host strains include primarily mammalian cells, and bacteria. This important guide:

• Contains a concise summary of the state-of-the-art bioprocessing methods
• Reviews the broad spectrum of applications to a variety of life science industries
• Includes illustrative case studies that review several milestone bio-products
• Explores of a wide selection of host strain types
• Contains information on disruptive bioprocess technologies Written for industrial and academic researchers and development scientists in the life sciences industry, Bioprocessing Technology for Production of Biopharmaceuticals and Bioproducts is a guide to the tools, approaches, and useful developments in bioprocessing.

Inhalt
List of Contributors xi

Part I Case Study 1

**1 Bacillus and the Story of Protein Secretion and Production 3
**Giulia Barbieri, Anthony Calabria, Gopal Chotani, and Eugenio Ferrari

1.1 Bacillus as a Production Host: Introduction and Historical Account 3

1.2 The Building of a Production Strain: Genetic Tools for B. subtilis Manipulation 5

1.2.1 Promoters 5

1.2.2 Vectors for Building a Production Strain 6

1.2.3 B. subtilis Competent Cell Transformation 7

1.2.4 Protoplasts-Mediated Manipulations 9

1.2.5 Genetics by Electroporation 9

1.3 B. subtilis Secretion Systemand Heterologous Protein Production 9

1.3.1 Bacillus Fermentation and Recovery of Industrial Enzyme 11

1.3.2 Fermentation Stoichiometry 12

1.3.3 Fermentor Kinetics and Outputs 14

1.3.4 Downstream Processing 17

1.4 Summary 21

References 21

**2 New Expression Systems for GPCRs 29
**Dimitra Gialama, Fragiskos N. Kolisis, and Georgios Skretas

2.1 Introduction 29

2.2 Recombinant GPCR Production Traditional Approaches for Achieving High-Level Production 39

2.3 Engineered Expression Systems for GPCR Production 42

2.3.1 Bacteria 42

2.3.2 Yeasts 48

2.3.3 Insect Cells 51

2.3.4 Mammalian Cells 54

2.3.5 Transgenic Animals 54

2.3.6 Cell-Free Systems 56

2.4 Conclusion 57

References 58

**3 Glycosylation 71
**Maureen Spearman, Erika Lattová, Hélène Perreault, andMichael Butler

3.1 Introduction 71

3.2 Types of Glycosylation 72

3.2.1 N-linked Glycans 72

3.2.2 O-linked Glycans 74

3.3 Factors Affecting Glycosylation 76

3.3.1 Nutrient Depletion 76

3.3.2 Fed-batch Cultures and Supplements 79

3.3.3 Specific Culture Supplements 80

3.3.4 Ammonia 82

3.3.5 pH 82

3.3.6 Oxygen 83

3.3.7 Host Cell Systems 83

3.3.8 Other Factors 85

3.4 Modification of Glycosylation 86

3.4.1 siRNA and Gene Knockout/Knockin 86

3.4.2 Glycoprotein Processing Inhibitors and In Vitro Modification of Glycans 88

3.5 Glycosylation Analysis 89

3.5.1 Release of Glycans from Glycoproteins 90

3.5.2 Derivatization of Glycans 91

3.6 Methods of Analysis 91

3.6.1 Lectin Arrays 91

3.6.2 Liquid Chromatography 93

3.6.2.1 HILIC Analysis 93

3.6.2.2 Reversed Phase (RP) and Porous Graphitic Carbon (PGC) Chromatography 95

3.6.2.3 Weak Anion Exchange (WAX) HPLC Analysis 96

3.6.2.4 High pH Anion Exchange Chromatography with Pulsed Amperometric Detection (HPAEC-PAD) 96

3.6.3 Capillary Electrophoresis (CE) 97

3.6.4 Fluorophore-assisted Carbohydrate Electrophoresis (FACE) and CGE-LIF 99

3.6.5 Mass Spectrometry (MS) 100

3.6.5.1 Ionization 100

3.6.5.2 Derivatization Techniques Used for MS Analysis of Glycans 102

3.6.5.3 Fragmentation of Carbohydrates 103

3.7 Conclusion 109

References 109

Part II Bioreactors 131

**4 Bioreactors for StemCell and Mammalian Cell Cultivation 133
**Ana Fernandes-Platzgummer, Sara M. Badenes, Cláudia L. da Silva, and JoaquimM. S. Cabral

4.1 Overview of (Mammalian and Stem) Cell Culture Engineering 133

4.1.1 Cell Products for Therapeutics 134

4.1.2 Cell as a Product: Stem Cells 136

4.2 Bioprocess Characterization 140

4.2.1 Cell Cultivation Methods 140

4.2.2 Cell Metabolism 141

4.2.3 Culture Medium Design 143

4.2.4 Culture Parameters 144

4.2.5 Culture Modes 145

4.3 Cell Culture Systems 147

4.3.1 Static Culture Systems 147

4.3.2 Roller Bottles 150

4.3.3 Spinner Flask 150 4.3.4 Airlift Bioreactor 151</p&g...