Synthetic propeptide design to enhance the secretion of heterologous proteins by Saccharomyces cerevisiae

Saccharomyces cerevisiae is widely used for heterologous protein production but often secretes at comparatively low levels. To improve this, a secretion-enhancing peptide cassette was added to the target protein and the propeptide was optimized using three key parameters:N-glycosylation, net negative charge balance, and glycine-rich flexible linker. Production of the model protein, the human granulocyte colony-stimulating factor, was increased by 190% compared to the original propeptide. This stepwise rational design is a promising strategy applicable to other heterologous proteins. AbstractHeterologous protein production inSaccharomyces cerevisiae is a useful and effective strategy with many advantages, including the secretion of proteins that require posttranslational processing. However, heterologous proteins inS. cerevisiae are often secreted at comparatively low levels. To improve the production of the heterologous protein, human granulocyte colony-stimulating factor (hG-CSF) inS. cerevisiae, a secretion-enhancing peptide cassette including an hIL-1 β-derived propeptide, was added and used as a secretion enhancer to alleviate specific bottlenecks in the yeast secretory pathway. The effects of three key parameters—N-glycosylation, net negative charge balance, and glycine-rich flexible linker —were investigated in batch cultures ofS. cerevisiae. Using a three-stage design involving screening, selection, and optimization, the production and secretion of hG-CSF byS. cer...

Source: MicrobiologyOpen - June 9, 2022 Category: Microbiology Authors: Ji Sung Cho, Hye Ji Oh, Young Eun Jang, Hyun Jin Kim, Areum Kim, Jong ‐Am Song, Eun Jung Lee, Jeewon Lee Tags: ORIGINAL ARTICLE Source Type: research