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Comparative expression of lipase CAL-A in the yeasts Saccharomyces cerevisiae, Kluyveromyces lactis and Hansenula polymorpha to investigate a possible host influence

Paper ID Volume ID Publish Year Pages File Format Full-Text
23013 43409 2014 11 PDF Available
Title
Comparative expression of lipase CAL-A in the yeasts Saccharomyces cerevisiae, Kluyveromyces lactis and Hansenula polymorpha to investigate a possible host influence
Abstract

•One optimized gene sequence of CAL-A was used for the expression in three yeast hosts.•N-Deglycosylation (non-denaturing or tunicamycin-blocked) reduced CAL-A activity.•The thermostability and substrate profiles vary depending on the host.•The protein host influences the enzyme properties.

Yeasts are an attractive expression platform, as they combine the ease of handling with the eukaryotic ability to process the produced protein. Important aspects of eukaryotic protein expression are posttranslational modifications, which can be required for functional expression of the protein of interest and can only be performed by eukaryotes. Each organism has its own modification pattern: for instance, the same protein produced in different hosts is subjected to various glycosylation pathways. It is amenable that these kinds of modifications can have an influence on the biochemical properties of the protein. To verify this thesis, the well-studied lipase CAL-A from Candida antarctica was chosen as a model enzyme. The codon bias of the gene sequence was uniformly optimized and expressed in three industrially relevant yeast hosts: Saccharomyces cerevisiae, Kluyveromyces lactis, and Hansenula polymorpha. The capacity of the expression systems to produce the enzyme was analyzed, as well as the biochemical properties of the produced and purified CAL-A. All hosts produced active enzyme; however, significant differences in the obtained yield were observed. H. polymorpha appeared to be the most productive host with a tenfold increase in productivity in comparison to S. cerevisiae. Studies on thermostability and activity of the purified enzymes towards various substrates showed a significant impact of the host on the biochemical properties of the produced protein. The most thermostable CAL-A from K. lactis retained 70% of its activity after incubation at 60 °C, in comparison to 45% remaining activity for the enzyme purified from S. cerevisiae. In the screenings with different substrates, a fourfold increase in activity between the enzymes from H. polymorpha and S. cerevisiae was found. Altogether, we herein exemplify how the selection of the host even within one taxonomic family (Saccharomycetaceae) significantly affects the produced enzyme's characteristics.

Keywords
Candida antarctica lipase A; Comparative expression; Yeast expression systems; Posttranslational modifications; Host influence
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Comparative expression of lipase CAL-A in the yeasts Saccharomyces cerevisiae, Kluyveromyces lactis and Hansenula polymorpha to investigate a possible host influence
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 191, 10 December 2014, Pages 176–186
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