6M4S

Crystal Structure Analysis of the cytochrome P450 CYP-Sb21

Summary for 6M4S

• Entry DOI: 10.2210/pdb6m4s/pdb
• Descriptor: Cytochrome P450 hydroxylase sb21, GLYCEROL, (4S)-2-METHYL-2,4-PENTANEDIOL, ... (6 entities in total)
• Functional Keywords: cytochrome p450, cyp-sb21, oxidoreductase
• Biological source: Nonomuraea dietziae
• Total number of polymer chains: 1
• Total formula weight: 47280.32

Authors

Li, F.W.,Li, S.Y. (deposition date: 2020-03-09, release date: 2021-02-03, Last modification date: 2023-11-29)

Primary citation

Li, F.,Ma, L.,Zhang, X.,Chen, J.,Qi, F.,Huang, Y.,Qu, Z.,Yao, L.,Zhang, W.,Kim, E.S.,Li, S.
Structure-guided manipulation of the regioselectivity of the cyclosporine A hydroxylase CYP-sb21 from Sebekia benihana .
Synth Syst Biotechnol, 5:236-243, 2020

PubMed Abstract: The cytochrome P450 enzyme CYP-sb21 from the rare actinomycete is capable of hydroxylating the immunosuppressive drug molecule cyclosporine A (CsA) primarily at the 4th N-methyl leucine (MeLeu), giving rise to -hydroxy--methyl-l-Leu-CsA (CsA-4-OH). This oxidative modification of CsA leads to dramatically reduced immunosuppressive activity while retaining the hair growth-promoting side-effect, thus demonstrating great application potential in both pharmaceutical and cosmetic industries. However, this P450 enzyme also hydroxylates CsA at the unwanted position of the 9th -methyl leucine (MeLeu), indicating that the regioselectivity needs to be improved for the development of CsA-4-OH into a commercial hair growth stimulator. Herein, we report the crystal structure of CYP-sb21 in its substrate-free form at 1.85 Å. Together with sequence and 3D structure comparisons, Autodock-based substrate docking, molecular dynamics (MD) simulation, and site-directed mutagenesis, we identified a number of key residues including R294, E264, and M179 that can improve catalytic efficiency or change the regioselectivity of CYP-sb21 towards CsA, setting the stage for better enzymatic preparation of CsA-4-OH. This study also provides new insights into the substrate recognition and binding mechanism of P450 enzymes that accommodate bulky substrates.

PubMed: 32775708
DOI: 10.1016/j.synbio.2020.07.004

Experimental method

X-RAY DIFFRACTION (1.85 Å)