title: In vitro reconstitution of the cyclosporine specific P450 hydroxylases using heterologous redox partner proteins.
abstract: The cytochrome P450 enzymes (CYPs) CYP-sb21 from Sebekia benihana and CYP-pa1 from Pseudonocardia autotrophica are able to hydroxylate the immunosuppressant cyclosporin A (CsA) in a regioselective manner, giving rise to the production of two hair-stimulating agents (with dramatically attenuated immunosuppressant activity), γ-hydroxy-N-methyl-L-Leu4-CsA (CsA-4-OH) and γ-hydroxy-N-methyl-L-Leu9-CsA (CsA-9-OH). Recently, the in vitro activity of CYP-sb21 was identified using several surrogate redox partner proteins. Herein, we reconstituted the in vitro activity of CYP-pa1 for the first time via a similar strategy. Moreover, the supporting activities of a set of ferredoxin (Fdx)/ferredoxin reductase (FdR) pairs from the cyanobacterium Synechococcus elongatus PCC 7942 were comparatively analyzed to identify the optimal redox systems for these two CsA hydroxylases. The results suggest the great value of cyanobacterial redox partner proteins for both academic research and industrial application of P450 biocatalysts.
authors.affiliation: College of Chemical Engineering, Qingdao University of Science and Technology, Qingdao, Shandong, China.
authors.forename: Yue
authors.initials: Y
authors.lastname: Sun
authors.affiliation: Shandong Provincial Key Laboratory of Synthetic Biology, CAS Key Laboratory of Biofuels, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong, China.
authors.forename: Li
authors.initials: L
authors.lastname: Ma
authors.affiliation: Shandong Provincial Key Laboratory of Synthetic Biology, CAS Key Laboratory of Biofuels, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong, China.
authors.forename: Dongfei
authors.initials: D
authors.lastname: Han
authors.affiliation: Shandong Provincial Key Laboratory of Synthetic Biology, CAS Key Laboratory of Biofuels, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Du
authors.affiliation: Shandong Provincial Key Laboratory of Synthetic Biology, CAS Key Laboratory of Biofuels, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong, China.
authors.forename: Fengxia
authors.initials: F
authors.lastname: Qi
authors.affiliation: Shandong Provincial Key Laboratory of Synthetic Biology, CAS Key Laboratory of Biofuels, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Shandong Provincial Key Laboratory of Synthetic Biology, CAS Key Laboratory of Biofuels, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong, China.
authors.forename: Jingran
authors.initials: J
authors.lastname: Sun
authors.affiliation: College of Chemical Engineering, Qingdao University of Science and Technology, Qingdao, Shandong, China.
authors.forename: Shan
authors.initials: S
authors.lastname: Huang
authors.affiliation: Department of Biological Engineering, Inha University, Incheon, South Korea. eungsoo@inha.ac.kr.
authors.forename: Eung-Soo
authors.initials: ES
authors.lastname: Kim
authors.affiliation: Shandong Provincial Key Laboratory of Synthetic Biology, CAS Key Laboratory of Biofuels, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong, China. lishengying@qibebt.ac.cn.
authors.forename: Shengying
authors.initials: S
authors.lastname: Li
mesh_terms: D000192:Actinomycetales / Q000145:classification / Q000235:genetics*; D016572:Cyclosporine / Q000737:chemistry*; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D007166:Immunosuppressive Agents / Q000737:chemistry; D010084:Oxidation-Reduction; D017422:Sequence Analysis, DNA
chemical_list: D004269:DNA, Bacterial; D007166:Immunosuppressive Agents; D016572:Cyclosporine; D003577:Cytochrome P-450 Enzyme System
keywords: Cyclosporin A; Hair-stimulating agent; P450 hydroxylase; Redox partner proteins; Synechococcus elongatus