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KMID : 0545120230330111513
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Journal of Microbiology and Biotechnology
2023 Volume.33 No. 11 p.1513 ~ p.1520
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Modulation of Kex2p Cleavage Site for In Vitro Processing of Recombinant Proteins Produced by Saccharomyces cerevisiae
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Kim Mi-Jin
Park Se-Lin
Kim Seung-Hwa
Park Hyun-Joo
Sung Bong-Hyun
Sohn Jung-Hoon
Bae Jung-Hoon
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Abstract
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Kex2 protease (Kex2p) is a membrane-bound serine protease responsible for the proteolytic maturation of various secretory proteins by cleaving after dibasic residues in the late Golgi network. In this study, we present an application of Kex2p as an alternative endoprotease for the in vitro processing of recombinant fusion proteins produced by the yeast Saccharomyces cerevisiae. The proteins were expressed with a fusion partner connected by a Kex2p cleavage sequence for enhanced expression and easy purification. To avoid in vivo processing of fusion proteins by Kex2p during secretion and to guarantee efficient removal of the fusion partners by in vitro Kex2p processing, P1′, P2′, P4, and P3 sites of Kex2p cleavage sites were elaborately manipulated. The general use of Kex2p in recombinant protein production was confirmed using several recombinant proteins.
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KEYWORD
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Recombinant protein, secretion, Kex2p, in-vitro processing, Saccharomyces cerevisiae
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