Engineering Saccharomyces cerevisiae for Hyperproduction of β‑Amyrin by Mitigating the Inhibition\nEffect of Squalene on β‑Amyrin Synthase

The study aims to enhance β-amyrin\nproduction in Saccharomyces cerevisiae by peroxisome compartmentalization.\nFirst, overaccumulated squalene was determined as a key limiting factor\nfor the production of β-amyrin since it could inhibit the activity\nof β-amyrin synthase GgbAs1. Second, to mitigate the inhibition\neffect, the enhanced squalene synthesis pathway was compartmentalized\ninto peroxisomes to insulate overaccumulated squalene from GgbAs1,\nand thus the specific titer of β-amyrin reached 57.8 mg/g dry\ncell weight (DCW), which was 2.6-fold higher than that of the cytosol\nengineering strain. Third, by combining peroxisome compartmentalization\nwith the “push-pull-restrain” strategy (ERG1 and GgbAs1\noverexpression and ERG7 weakening), the production of β-amyrin\nwas further increased to 81.0 mg/g DCW (347.0 mg/L). Finally, through\nfed-batch fermentation in a 5 L fermenter, the titer of β-amyrin\nreached 2.6 g/L, which is the highest reported to date. The study\nprovides a new perspective to engineering yeasts as a platform for\ntriterpene production.