Stepwise Optimization of Inducible Expression System for the Functional Secretion of Horseradish Peroxidase in <i>Saccharomyces cerevisiae</i>
Xinrui Zhao, Haibo Yu, Qingfeng Liang, Jingwen Zhou, Jianghua Li, Guocheng Du, Jian Chen
Abstract
Horseradish peroxidase (HRP) is a plant-derived glycoprotein that can be developed as a food additive to cross-link proteins or biopolymers. Although Saccharomyces cerevisiae has advantages in the production of food-grade HRP, the low expressional level and inefficient secretion hindered its application values. After comparing the effects of constitutive and inducible expression on cell growth, the strength of HRP expression was roughly tuned by replacing core regions of the promoter in the GAL80 -knockout strain and further finely tuned by terminator screening. Additionally, the most suitable signal peptide was selected, and the pre-peptide with pro-peptides was modified to balance the transport of HRP in the endoplasmic reticulum. The extracellular HRP activity of the best strain reached 13 506 U/L at the fermenter level, 330-fold higher than the previous result of 41 U/L in S. cerevisiae . The strategy can be applied to alleviate the inhibition of cell growth caused by the expression of toxic proteins and improve their secretion.