Establishing cell suitability for high-level production of licorice triterpenoids in yeast  

作　　者：Wentao Sun Shengtong Wan Chuyan Liu Ruwen Wang Haocheng Zhang Lei Qin Runming Wang Bo Lv Chun Li 

机构地区：[1]Department of Chemical Engineering,Tsinghua University,Beijing 100084,China [2]Key Laboratory of Medical Molecule Science and Pharmaceutics Engineering,Ministry of Industry and Information Technology,Institute of Biochemical Engineering,School of Chemistry and Chemical Engineering,Beijing Institute of Technology,Beijing 100081,China [3]The University of Chicago,Chicago,IL 60637,USA [4]Key Lab for Industrial Biocatalysis,Ministry of Education,Tsinghua University,Beijing 100084,China [5]Center for Synthetic and Systems Biology,Tsinghua University,Beijing 100084,China [6]Institute of Biopharmaceutical and Health Engineering,Tsinghua Shenzhen International Graduate School,Tsinghua University,Shenzhen 518055,China

出　　处：《Acta Pharmaceutica Sinica B》2024年第9期4134-4148,共15页药学学报（英文版）

基　　金：grants from the National Key Research and Development Program of China(2021YFC2100800);the National Natural Science Foundation of China(No.22108154,No.22138006,No.32171430,No.22278240);the Natural Science Foundation of Beijing Municipality(M21010,China);the Shuimu Tsinghua Scholar Program(2020SM097,China).

摘　　要：Yeast has been an indispensable host for synthesizing complex plant-derived naturalcompounds, yet the yields remained largely constrained. This limitation mainly arises from overlookingthe importance of cell and pathway suitability during the optimization of enzymes and pathways. Herein,beyond conventional enzyme engineering, we dissected metabolic suitability with a framework forsimultaneously augmenting cofactors and carbon flux to enhance the biosynthesis of heterogenoustriterpenoids. We further developed phospholipid microenvironment engineering strategies, dramaticallyimproving yeast’s suitability for the high performance of endoplasmic reticulum (ER)-localized, ratelimitingplant P450s. Combining metabolic and microenvironment suitability by manipulating only threegenes, NHMGR (NADH-dependent HMG-CoA reductase), SIP4 (a DNA-binding transcription factor)andGPP1 (Glycerol-1-phosphate phosphohydrolase 1), we enabled the high-level production of 4.92 g/L rarelicorice triterpenoids derived from consecutive oxidation of b-amyrin by two P450 enzymes afterfermentation optimization. This production holds substantial commercial value, highlighting the criticalrole of establishing cell suitability in enhancing triterpenoid biosynthesis and offering a versatileframework applicable to various plant natural product biosynthetic pathways.

关 键 词：Cell suitability Phospholipid microenvironment Cytochrome P450 TRITERPENOIDS Saccharomyces cerevisiae 

分 类 号：R28[医药卫生—中药学]