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Biopharmaceutical Engineering : Focus on Recombinant Proteins

par: Pr. Sonia Adi-Bessalem Dr. Fares Daachi
Disponible
Pages 136
Dimensions 16*23,5
Publication 2025-06-26
Éditeur —
Papier : 950.00 DA Numérique: 475.00 DA

The field of Biopharmaceutical Engineering has seen remarkable advancements over the past decades, with recombinant protein technology standing at the forefront of therapeutic and diagnostic innovation. Biopharmaceuticals derived from recombinant proteins have transformed modern medicine, offering highly specific and effective treatments for a wide range of diseases, including cancer, autoimmune disorders, and genetic conditions. As the demand for biologics and protein-based treatments continues to rise, a comprehensive understanding of the processes involved in their design, expression, and large-scale production becomes essential. This book, Biopharmaceutical Engineering: Focus on Recombinant Proteins, serves as a complete guide to the principles and practices of recombinant protein engineering. It is designed to provide students, researchers, and professionals with the fundamental knowledge necessary to navigate the complexities of bioengineering, from molecular design to industrial production. The chapters cover essential topics such as molecular cloning, transformation and transfection techniques, expression systems, protein purification, and quality control. By the end of this book, readers will have acquired an in-depth understanding of recombinant protein production and its applications in modern healthcare. We hope that this book serves as a valuable resource for those looking to expand their expertise in Biopharmaceutical Engineering and contributes to the advancement of knowledge in this rapidly evolving field.

Afficher la table des matières
preface 03
Table of Contents 05
Introduction to biotechnology 09
Bioengineering of therapeutic biomolecules 11
Processes used in the engineering of therapeutic biomolecules A)Chemical and enzymatic modifications 11
B)Modifications by genetic engineering (Bioengineering of recombinant proteins)
Chapter 1: Bioengineering of recombinant proteins 15
1. Molecular cloning 17
1.1.The molecular tools of cloning
1.1.1. Restriction enzymes 18
1.1.2. DNA-cutting enzymes (Nuclease) 19
1.1.3. Enzymes removing or adding phosphate groups.
1.1.4. Enzymes copying nucleic acids: DNA polymerase – Reverse retrotranscriptase – RNA polymerase
2. Some molecular biology methods 22
2.1. Amplification of specific nucleic acids 22
2.1.1. Polymerase chain reaction (PCR) 22
2.1.2. PCR-directed mutagenesis 23
2.1.3. rt-pcr 24
2.1.4. Real-time PCR 25
2.1.5. The operating conditions of PCR 27
2.1.6. The limits of the PCR technique 29
2.2. dna library 29
2.2.1. genomic dna library 29
2.2.2. cdna Library 30
2.3. Library screening: Selection of specific nucleic acids 30
2.3.1. Antibiotic resistance screening: 30
2.3.2. Hybridization screening 31
2.3.3. Immunological detection 32
Summative evaluation 34
Chapter 2: Expression Systems of recombinant protein (Vectors+ Promoters)
1. Properties of vectors 39
2. Types of vectors (Clone/Expression)
2.1. plasmids 40
2.2. Vectors for Yeast 44
2.2.1. yip (yeast integrating plasmid):
2.2.2. yep (yeast episomal plasmid)
2.2.3. ycp (yeast centromere plasmid):
2.3. Vectors for mammalian cells
2.4. Virus as vectors: 49
3. Types of promoters used 40
Summative evaluation 53
Chapter 3: Transformation and transfection: Methods For Incorporation Of Expression Vectors Into Host Cells
1. chemical methods 58
2. Physical methods 59
2.1. Electroporation (electro permeabilization) 59
2.2. Biolistic method: bombardment of cells with DNA-coated beads. 59
3. Biological methods (INFECTION) 59
Summative evaluation 61
Chapter 4: Expression of recombinant proteins 65
1. Expression in the prokaryotic system
1.1. Escherichia coli 68
1.2. Other bacteria used in protein production: Bacillus, Streptomyces
2. Expression in eukaryotic system
2.1. yeasts 69
2.2. mammalian cells 71
2.3. the baculovirus-insect cell system 68
2.4. Transgenic plants 73
Summative evaluation 75
Chapter 5: different cellular compartments for recombinant
protein expression 79
1. inclusion bodies 80
1.1. Denaturation (Solubilization) and Renaturation of Proteins in Vitro
1.2. Minimizing in vivo aggregation
2. Cytoplasm-soluble proteins 81
3. Expression in the periplasm (4% of bacterial proteins) 81
Summative evaluation 84
Chapter 6: Production of recombinant proteins: bioreactor 87
1. Fermentation and scale-up stage
2. Different fermentation process 89
2.1. the batch process or batch fermentation
2.2. the Fed-batch process or fed batch fermentation
2.3. The continuous cultivation process or Chemostat. 88
Summative evaluation 90
Chapter 7: Extraction, purification and characterization of
recombinant proteins 93
1. Protein isolation (extraction) 94
2. General purification/characterization strategy 94
3. Protein purification using fusion proteins for labeling: tags and
proteases 94
Summative evaluation 100
Chapter 8: Structural and functional characterization of the
recombinant protein 103
1. Structural Features of Recombinant Proteins 104
1.1. physico-chemical analysis: 104
1.2. sequence determination 104
1.3. N- and C-Terminal Sequencing: 105
2. post-translational modification (ptm) control 106
2.1. Common Types of ptms and Their Detection Methods 106
2.2. Conformation Control and Aggregate Analysis 107
3. Functional Characterization of Recombinant Proteins 108
3.1. enzyme activity assays 108
3.2. cellular response assays 108
3.3. ligand binding studies 108
3.4. in vivo functional studies 109
3.5. Comparison with Reference Proteins 109
Summative evaluation
Chapter 9: The development of the biomedicine 115
1. Clinical trials 116
1.1. Phase I: Safety Assessment
1.2. Phase II: Beneficial effects
1.3. Phase III: Therapeutic effects on a larger population
1.4. Phase IV: Phase de pharmacovigilance
2. Marketing authorization and regulatory aspects 116
Summative evaluation 118
Chapter 10: Annexes 121
Conclusion 122
References 123
Glossary
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