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Enhancement of lipase r27RCL production in Pichia pastoris by regulating gene dosage and co-expression with chaperone protein disulfide isomerase

Paper ID Volume ID Publish Year Pages File Format Full-Text
17075 42638 2013 6 PDF Available
Title
Enhancement of lipase r27RCL production in Pichia pastoris by regulating gene dosage and co-expression with chaperone protein disulfide isomerase
Abstract

•A series of strains with different gene dosage were generated in Muts phenotype.•Co-expression with PDI significantly improved the protein quantity and quality.•The highest enzyme activity was achieved by the five copy strain.•This research aids understanding the secretory pathway in Pichia pastoris.

Pichia pastoris has been successfully used in the production of many secreted and intracellular recombinant proteins, but there is still a large room of improvement for this expression system. Two factors drastically influence the lipase r27RCL production from Rhizopus chinensis CCTCC M201021, which are gene dosage and protein folding in the endoplasmic reticulum (ER). Regarding the effect of gene dosage, the enzyme activity for recombinant strain with three copies lipase gene was 1.95-fold higher than that for recombinant strain with only one copy lipase gene. In addition, the lipase production was further improved by co-expression with chaperone PDI involved in the disulfide bond formation in the ER. Overall, the maximum enzyme activity reached 355 U/mL by the recombinant strain with one copy chaperone gene PDI plus five copies lipase gene proRCL in shaking flasks, which was 2.74-fold higher than that for the control strain with only one copy lipase gene. Overall, co-expression with PDI vastly increased the capacity for processing proteins of ER in P. pastoris.

Keywords
Lipase; Gene dosage; Chaperone PDI; Pichia pastoris
First Page Preview
Enhancement of lipase r27RCL production in Pichia pastoris by regulating gene dosage and co-expression with chaperone protein disulfide isomerase
Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 53, Issues 6–7, 10 December 2013, Pages 438–443
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering