Details

Autor(en) / Beteiligte

• Besada-Lombana, Pamela B.
• Da Silva, Nancy A.

Titel

Engineering the early secretory pathway for increased protein secretion in Saccharomyces cerevisiae

Ist Teil von

• Metabolic engineering, 2019-09, Vol.55, p.142-151

Ort / Verlag

Belgium: Elsevier Inc

Erscheinungsjahr

2019

Quelle

MEDLINE

Beschreibungen/Notizen

• The yeast Saccharomyces cerevisiae is a valuable host for the production of heterologous proteins with a wide array of applications, ranging from cellulose saccharification enzymes to biopharmaceuticals. Efficient protein secretion may be critical for economic viability; however previous efforts have shown limited improvements that are often protein-specific. By enhancing transit through the early secretory pathway, we have successfully improved extracellular levels of three different proteins from variety of origins: a bacterial endoglucanase (CelA), a fungal β-glucosidase (BglI) and a single-chain antibody fragment (4-4-20 scFv). Efficient co-translational translocation into the endoplasmic reticulum (ER) was achieved via secretion peptide engineering and the novel use of a 3′-untranslated region, improving extracellular activity or fluorescence 2.2–5.4-fold. We further optimized the pathway using a variety of new strategies including: i) increasing secretory pathway capacity by expanding the ER, ii) limiting ER-associated degradation, and iii) enhancing exit from the ER. By addressing these additional ER processing steps, extracellular activity/fluorescence increased by 3.5–7.1-fold for the three diverse proteins. The optimal combination of pathway interventions varied, and the highest overall increases ranged from 5.8 to 11-fold. These successful strategies should prove effective for improving the secretion of a wide range of heterologous proteins. •Improved protein secretion by enhancing transit through the early secretory pathway.•Transit improved by targeting ER entry, expansion, retro-translocation, and exit.•Efficient co-translational translocation via novel use of 3′UTR enhanced secretion.•Combined strategies improved secretion up to 11-fold for three heterologous proteins.