Engineering protein folding and translocation improves heterologous protein secretion in Saccharomyces cerevisiae

• Published in: Biotechnology and bioengineering Vol. 112; no. 9; pp. 1872 - 1882
• Main Authors: Tang, Hongting, Bao, Xiaoming, Shen, Yu, Song, Meihui, Wang, Shenghuan, Wang, Chengqiang, Hou, Jin
• Format: Journal Article
• Language: English
• Published: United States Blackwell Publishing Ltd 01.09.2015; Wiley Subscription Services, Inc
• Subjects: Bioengineering; Biotechnology; chaperones; Endoglucanase; Endoplasmic reticulum; Enzyme kinetics; Folding; heterologous protein secretion; Pathways; Protein Engineering - methods; Protein expression; Protein Folding; protein translocation; Proteins; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Saccharmyces cerevisiae; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Secretions; Signal Transduction; Yeast; chaperones; heterologous protein secretion; Saccharmyces cerevisiae; protein translocation
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.25596

ABSTRACT Saccharomyces cerevisiae is widely used as a producer of heterologous proteins of medical and industrial interest. Numerous efforts have been made to overcome bottlenecks in protein expression and secretion. However, the effect of engineering protein translocation to heterologous protein secretion has not been studied extensively in S. cerevisiae. In this work, we confirmed that heterologous protein expression in S. cerevisiae induced the unfolded protein response (UPR). To enhance protein folding capacity, the endoplasmic reticulum (ER) chaperone protein BiP and the disulfide isomerase Pdi1p were each over‐expressed, and the secretion of three heterologous proteins, β‐glucosidase, endoglucanase, and α‐amylase, was improved. The impact of engineering key translocation components was also studied. The over‐expression of co‐translational translocation components Srp14p and Srp54p enhanced the secretion of three heterologous proteins (β‐glucosidase, endoglucanase, and α‐amylase), but over‐expressing the cytosolic chaperone Ssa1p (involved in post‐translational translocation) only enhanced the secretion of β‐glucosidase. By engineering both co‐translational translocation and protein folding, we obtained strains with β‐glucosidase, endoglucanase, and α‐amylase activities increased by 72%, 60%, and 103% compared to the controls. Our results show that protein translocation may be a limiting factor for heterologous protein production. Biotechnol. Bioeng. 2015;112: 1872–1882. © 2015 Wiley Periodicals, Inc. Engineering protein folding and translocation can increase heterologous proteins secretion in Saccharomyces cerevisiae. Over‐expression of co‐translational components Srp14p and Srp54p enhanced the secretion of three heterologous proteins (β‐glucosidase, endoglucanase and α‐amylase), and engineering post‐translational translocation components only improved the secretion of β‐glucosidase and endogenous invertase. It indicates different proteins may translocate through different pathways. This study demonstrates that modifying the protein translocation pathway is a promising secretory pathway engineering approach for recombinant proteins production.