Transporter Engineering for Cellobiose Fermentation Under Lower PH Conditions by Engineered Saccharomyces Cerevisiae

The aim of this study was to engineer cellodextrin transporter 2 (CDT-2) from Neurospora crassa for improved cellobiose fermentation under lower pH conditions by Saccharomyces cerevisiae. Through directed evolution, a mutant CDT-2 capable of facilitating cellobiose fermentation under lower pH conditions was obtained. Specifically, a library of CDT-2 mutants with GFP fusion was screened by flow cytometry and then serial subcultured to isolate a CDT-2 mutant capable of transporting cellobiose under acidic conditions. The engineered S. cerevisiae expressing the isolated mutant CDT-2 (I96N/T487A) produced ethanol with a specific cellobiose consumption rate of 0.069g/gcell/h, which was 51% and 55% higher than those of the strains harboring wild-type CDT-1 and CDT-2 in a minimal medium with 2g/L of acetic acid.