Blocking endocytotic mechanisms to improve heterologous protein titers in Saccharomyces cerevisiae

William A. Rodríguez-Limas, Victoria Tannenbaum, Keith Edward Jaggard Tyo*

*Corresponding author for this work

Research output: Contribution to journalArticle

13 Scopus citations

Abstract

Saccharomyces cerevisiae is a useful platform for protein production of biopharmaceuticals and industrial enzymes. To date, substantial effort has focused on alleviating several bottlenecks in expression and the secretory pathway. Recently, it has been shown that highly active endocytosis could decrease the overall protein titer in the supernatant. In this study, we block endocytosis and trafficking to the vacuole using a modified TEV Protease-Mediated Induction of Protein Instability (mTIPI) system to disrupt the endocytotic and vacuolar complexes. We report that conditional knock-down of endocytosis gene Rvs161 improved the concentration of α-amylase in supernatant of S. cerevisiae cultures by 63.7% compared to controls. By adaptive evolution, we obtained knock-down mutants in Rvs161 and End3 genes with 2-fold and 3-fold α-amylase concentrations compared to controls that were not evolved. Our study demonstrates that genetic blocking of endocytotic mechanisms can improve heterologous protein production in S. cerevisiae. This result is likely generalizable to other eukaryotic secretion hosts.

Original languageEnglish (US)
Pages (from-to)376-385
Number of pages10
JournalBiotechnology and Bioengineering
Volume112
Issue number2
DOIs
StatePublished - Jan 1 2015

Keywords

  • Conditional mutants
  • Endocytosis
  • Protein production
  • TIPI system
  • Yeast

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

Fingerprint Dive into the research topics of 'Blocking endocytotic mechanisms to improve heterologous protein titers in Saccharomyces cerevisiae'. Together they form a unique fingerprint.

  • Cite this