Pichia, Optimization of Protein Expression

Introduction Background Strategies for Optimization of Protein Expression Cellular State of the Expression Cassette Site of Integration of the Expression Cassette Methanol Utilization Phenotype, Mut+ or Mut− of the Host Gene Dosage: Exploiting the Clonal Variation of Expression Translational Optimization: 5′ Untranslated Region Translational Optimization: Initiation Codon AUG Context Transcriptional Optimization Product Secretion Choice of the Secretion Signal Product Stabilization by Media Manipulations Protease-Deficient Strains Strategies for Coexpression of Proteins Leucine Zipper Dimerization Motif Mediated Protein Assembly Product Toxicity Strategies for Induction of Protein Expression Fermentation Process Fermentation Equipment Methods for Monitoring the Fermentation Process Continuous Culture of Mut+ and Mut− Pichia on Methanol Fed-Batch Fermentation of Mut+ and Mut− Pichia on Methanol Strategies for Multicycle Fermentation Conclusions and Future Perspectives Media Compositions Stock Solutions Minimal Media Compositions Supplemental Minimal Media Compositions Complex Media Composition Secretion Media Composition Fermentation Media Composition Glossary of P. pastoris Vectors Glossary of P. pastoris Strains Bibliography Keywords: alcohol (methanol) oxidase promoter; clonal variation; expression plasmid; fermentation; methylotrophic yeast; mRNA secondary structure; protease deficient strain; secretion; synthetic gene; transcriptional barrier