Tunnel engineering for modulating the substrate preference in cytochrome P450BsβHI

Bioresources and Bioprocessing, vol. 8, pp. 26

Abstract

Abstract An active site is normally located inside enzymes, hence substrates should go through a tunnel to access the active site. Tunnel engineering is a powerful strategy for refining the catalytic properties of enzymes. Here, P450 Bsβ HI (Q85H/V170I) derived from hydroxylase P450 Bsβ from Bacillus subtilis was chosen as the study model, which is reported as a potential decarboxylase. However, this enzyme showed low decarboxylase activity towards long-chain fatty acids. Here, a tunnel engineering campaign was performed for modulating the substrate preference and improving the decarboxylation activity of P450 Bsβ HI. The finally obtained BsβHI-F79A variant had a 15.2-fold improved conversion for palmitic acid; BsβHI-F173V variant had a 3.9-fold improved conversion for pentadecanoic acid. The study demonstrates how the substrate preference can be modulated by tunnel engineering strategy.

Authors 10

1. Shuaiqi Meng Aachen Institute of Biotechnology

   Beijing University of Chemical Technology · RWTH Aachen University

   Affiliation as printed

   Beijing Bioprocess Key Laboratory, Beijing University of Chemical Technology, Beijing, 100029, People's Republic of China

   Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, 52074, Aachen, Germany

2. Zhongyu Li

   Beijing University of Chemical Technology

   Affiliation as printed

   Beijing Bioprocess Key Laboratory, Beijing University of Chemical Technology, Beijing, 100029, People's Republic of China

3. Ulrich Schwaneberg Aachen DWI - Leibniz Institute for Interactive Materials Institute of Biotechnology

   DWI – Leibniz Institute for Interactive Materials · RWTH Aachen University

   Affiliation as printed

   DWI-Leibniz Institute for Interactive Materials, Forckenbeckstraße 50, 52074, Aachen, Germany

   Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, 52074, Aachen, Germany

4. Yu Ji Aachen Institute of Biotechnology

   RWTH Aachen University

   Affiliation as printed

   Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, 52074, Aachen, Germany

5. Mehdi D. Davari Aachen Institute of Biotechnology

   RWTH Aachen University

   Affiliation as printed

   Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, 52074, Aachen, Germany

6. Fang Wang

   Beijing University of Chemical Technology

   Affiliation as printed

   Beijing Bioprocess Key Laboratory, Beijing University of Chemical Technology, Beijing, 100029, People's Republic of China

7. Meng Wang

   Beijing University of Chemical Technology

   Affiliation as printed

   Beijing Bioprocess Key Laboratory, Beijing University of Chemical Technology, Beijing, 100029, People's Republic of China

8. Meng Qin

   Beijing University of Chemical Technology

   Affiliation as printed

   Beijing Bioprocess Key Laboratory, Beijing University of Chemical Technology, Beijing, 100029, People's Republic of China

9. Kaili Nie

   Beijing University of Chemical Technology

   Affiliation as printed

   Beijing Bioprocess Key Laboratory, Beijing University of Chemical Technology, Beijing, 100029, People's Republic of China

10. Luo Liu corresponding

    Beijing University of Chemical Technology

    Affiliation as printed

    Beijing Bioprocess Key Laboratory, Beijing University of Chemical Technology, Beijing, 100029, People's Republic of China. liuluo@mail.buct.edu.cn

    Beijing Bioprocess Key Laboratory, Beijing University of Chemical Technology, Beijing, 100029, People's Republic of China