Identification of new substrates for the CYP106A1-mediated 11-oxidation and investigation of the reaction mechanism.

CYP106A1 from Bacillus megaterium DSM319 was recently shown to catalyze steroid and terpene hydroxylations. Besides producing hydroxylated steroid metabolites at positions 6β, 7β, 9α and 15β, the enzyme displayed previously unknown 11-oxidase activity towards 11β-hydroxysteroids. Novel examples for 11-oxidation were identified and confirmed by (1)H and (13)C NMR for prednisolone, dexamethasone and 11β-hydroxyandrostenedione. However, only 11β-hydroxyandrostenedione formed a single 11-keto product. The latter reaction was chosen to investigate the kinetic solvent isotope effect on the steady-state turnover of the CYP106A1-mediated 11-oxidation. Our results reveal a large inverse kinetic isotope effect (∼0.44) suggesting the involvement of the ferric peroxoanion as a reactive intermediate.
(Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)