Regio- and stereoselective steroid hydroxylation by CYP109A2 from Bacillus megaterium explored by X-ray crystallography and computational modeling.
Cytochrome P450
X-ray crystallography
hydroxylation
near-attack conformations
steroids
Journal
The FEBS journal
ISSN: 1742-4658
Titre abrégé: FEBS J
Pays: England
ID NLM: 101229646
Informations de publication
Date de publication:
10 2023
10 2023
Historique:
revised:
31
05
2023
received:
12
12
2022
accepted:
16
06
2023
medline:
23
10
2023
pubmed:
15
7
2023
entrez:
15
7
2023
Statut:
ppublish
Résumé
The P450 monooxygenase CYP109A2 from Bacillus megaterium DSM319 was previously found to convert vitamin D3 (VD3) to 25-hydroxyvitamin D3. Here, we show that this enzyme is also able to convert testosterone in a highly regio- and stereoselective manner to 16β-hydroxytestosterone. To reveal the structural determinants governing the regio- and stereoselective steroid hydroxylation reactions catalyzed by CYP109A2, two crystal structures of CYP109A2 were solved in similar closed conformations, one revealing a bound testosterone in the active site pocket, albeit at a nonproductive site away from the heme-iron. To examine whether the closed crystal structures nevertheless correspond to a reactive conformation of CYP109A2, docking and molecular dynamics (MD) simulations were performed with testosterone and vitamin D3 (VD3) present in the active site. These MD simulations were analyzed for catalytically productive conformations, the relative occurrences of which were in agreement with the experimentally determined stereoselectivities if the predicted stability of each carbon-hydrogen bond was taken into account. Overall, the first-time determination and analysis of the catalytically relevant 3D conformation of CYP109A2 will allow for future small molecule ligand screening in silico, as well as enabling site-directed mutagenesis toward improved enzymatic properties of this enzyme.
Substances chimiques
Cytochrome P-450 Enzyme System
9035-51-2
Steroids
0
Cholecalciferol
1C6V77QF41
Testosterone
3XMK78S47O
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
5016-5035Informations de copyright
© 2023 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.
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