Effects of signal sequences and folding accessory proteins on extracellular expression of carboxypeptidase Y in recombinant Saccharomyces cerevisiae

So Yeon Shin, Yi Hyun Bae, Sun Ki Kim, Yeong Je Seong, Suk Hwan Choi, Kyoung Heon Kim, Yong Cheol Park, Jin Ho Seo

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Carboxypeptidase Y (CPY) is a yeast vacuolar protease with useful applications including C-terminal sequencing of peptides and terminal modification of target proteins. To overexpress CPY with the pro-sequence (proCPY) encoded by the Saccharomyces cerevisiae PRC1 gene in recombinant S. cerevisiae, the proCPY gene was combined with the gene coding for a signal sequence of S. cerevisiae mating factor a (MF a), invertase (SUC2), or Kluyveromyces marxianus inulinase (INU1). Among the three constructs, the MFa signal sequence gave the best specific activity of extracellular CPY. To enhance the CPY expression level, folding accessory proteins of Kar2p, Pdi1p and Ero1p located in the S. cerevisiae endoplasmic reticulum were expressed individually and combinatorially. A single expression of Kar2p led to a 28% enhancement in extracellular CPY activity, relative to the control strain of S. cerevisiae CEN.PK2-1D/p426Gal1-MFa CPY. Coexpression of Kar2p, Pdi1p and Ero1p gave a synergistic effect on CPY expression, of which activity was 1.7 times higher than that of the control strain. This work showed that engineering of signal sequences and protein-folding proteins would be helpful to overexpress yeast proteins of interest.

Original languageEnglish
Pages (from-to)1065-1071
Number of pages7
JournalBioprocess and Biosystems Engineering
Volume37
Issue number6
DOIs
Publication statusPublished - 2014 Jan 1

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ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering

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