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작성일 : 13-10-11 10:54

(2014, IF=1.809) Effects of signal sequences and folding accessory proteins on extracellular expression of carboxypeptidase Y in recombinant Saccharomyces cerevisiae

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http://link.springer.com/article/10.1007%2Fs00449-013-1078-0 [479]

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

So-Yeon Shin1,†, Yi-Hyun Bae2,†, Sun-Ki Kim1, Yeong-Je Seong3, Suk-Hwan Choi4, Kyoung Heon Kim5, Yong-Cheol Park3,** and Jin-Ho Seo1,2,*

Bioprocess and Biosystems Engineering, Accepted 37(6):1065-1071. 2014.06.01

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 PRC1gene in recombinant S. cerevisiae, the proCPY gene was combined with the gene coding for a signal sequence of S. cerevisiae mating factor α (MFα), invertase (SUC2), or Kluyveromyces marxianus inulinase (INU1). Among the three constructs, the MFα 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-MFαCPY. 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.

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