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Improvement in the secretory expression of recombinant Candida rugosa lipase in Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
34207 45010 2015 7 PDF Available
Title
Improvement in the secretory expression of recombinant Candida rugosa lipase in Pichia pastoris
Abstract

•Two combined processes were developed to improve Lip2 secretion in P. pastoris.•The process includes gene copy amplification and low-temperature culture.•The process results in a maximal 32-fold increase in Lip2 secretion.•The steady-state LIP2 mRNA amount determines the maximal secretion level of Lip2.

The yeast Candida rugosa can secrete a mixture of lipase isoenzymes (Lips), which have been widely applied in industry. Eight Lip genes (LIP1 to LIP8) have been identified and are expressed in Pichia pastoris. However, the expression level was not sufficient for economical industrial application. In this study, two combined processes of antibiotic selection and low-temperature culture efficiently elicited a high-level secretion of recombinant Lip2 in P. pastoris. The LIP2 gene copy number of the Pichia transformants was increased by sequential selections at gradually increasing Zeocin concentrations. After the first selection at 500 μg/mL of Zeocin, three clones (500-clones) with 2.4-fold to 5.8-fold improvement in Lip2 secretion were identified from 105 survival clones through lipase activity screening. Although the maximum number of LIP2 gene copy was four among these three 500-clones, the lipase secretion of the four-copy clone was not higher than that of the three-copy clone. The effects of multiple gene copy number and low culture temperature resulted in a maximal 32-fold increase in Lip2 secretion. This method could be applied to other Lip isoforms to enhance their yields in P. pastoris.

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Keywords
Lipase; Multi-copy strains; Temperature; Secretion; Pichia pastoris; Candida rugosa
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Improvement in the secretory expression of recombinant Candida rugosa lipase in Pichia pastoris
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Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 50, Issue 12, December 2015, Pages 2137–2143
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us