Engineering of Yeast Old Yellow Enzyme OYE3 Enables Its Capability Discriminating of (<i>E</i>)-Citral and (<i>Z</i>)-Citral
Tairan Wang,
Ran Wei,
Yingting Feng,
Lijun Jin,
Yunpeng Jia,
Duxia Yang,
Zuonan Liang,
Mengge Han,
Xia Li,
Chenze Lu,
Xiangxian Ying
Affiliations
Tairan Wang
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Ran Wei
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Yingting Feng
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Lijun Jin
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Yunpeng Jia
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Duxia Yang
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Zuonan Liang
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Mengge Han
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Xia Li
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
Chenze Lu
College of Life Sciences, China Jiliang University, Hangzhou 310018, China
Xiangxian Ying
Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, China
The importance of yeast old yellow enzymes is increasingly recognized for direct asymmetric reduction of (E/Z)-citral to (R)-citronellal. As one of the most performing old yellow enzymes, the enzyme OYE3 from Saccharomyces cerevisiae S288C exhibited complementary enantioselectivity for the reduction of (E)-citral and (Z)-citral, resulting in lower e.e. value of (R)-citronellal in the reduction of (E/Z)-citral. To develop a novel approach for the direct synthesis of enantio-pure (R)-citronellal from the reduction of (E/Z)-citral, the enzyme OYE3 was firstly modified by semi-rational design to improve its (R)-enantioselectivity. The OYE3 variants W116A and S296F showed strict (R)-enantioselectivity in the reduction of (E)-citral, and significantly reversed the (S)-enantioselectivity in the reduction of (Z)-citral. Next, the double substitution of OYE3 led to the unique variant S296F/W116G, which exhibited strict (R)-enantioselectivity in the reduction of (E)-citral and (E/Z)-citral, but was not active on (Z)-citral. Relying on its capability discriminating (E)-citral and (Z)-citral, a new cascade reaction catalyzed by the OYE3 variant S296F/W116G and glucose dehydrogenase was developed, providing the enantio-pure (R)-citronellal and the retained (Z)-citral after complete reduction of (E)-citral.
