Affiliation:
1. Department of Biology and Biological Engineering, Chalmers University of Technology , Kemivägen 10, SE-41296 Gothenburg, Sweden
2. Novo Nordisk Foundation Center for Biosustainability, Chalmers University of Technology , Kemivägen 10, SE-41296 Gothenburg, Sweden
Abstract
Abstract
The rapid expansion of the application of pharmaceutical proteins and industrial enzymes requires robust microbial workhorses for high protein production. The budding yeast Saccharomyces cerevisiae is an attractive cell factory due to its ability to perform eukaryotic post-translational modifications and to secrete proteins. Many strategies have been used to engineer yeast platform strains for higher protein secretion capacity. Herein, we investigated a line of strains that have previously been selected after UV random mutagenesis for improved α-amylase secretion. A total of 42 amino acid altering point mutations identified in this strain line were reintroduced into the parental strain AAC to study their individual effects on protein secretion. These point mutations included missense mutations (amino acid substitution), nonsense mutations (stop codon generation), and frameshift mutations. For comparison, single gene deletions for the corresponding target genes were also performed in this study. A total of 11 point mutations and seven gene deletions were found to effectively improve α-amylase secretion. These targets were involved in several bioprocesses, including cellular stresses, protein degradation, transportation, mRNA processing and export, DNA replication, and repair, which indicates that the improved protein secretion capacity in the evolved strains is the result of the interaction of multiple intracellular processes. Our findings will contribute to the construction of novel cell factories for recombinant protein secretion.
Funder
Swedish Foundation for Strategic Research
VINNOVA
Novo Nordisk Foundation
Publisher
Oxford University Press (OUP)
Subject
Applied Microbiology and Biotechnology,General Medicine,Microbiology
Cited by
7 articles.
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