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Protein Engineering of the Progesterone Hydroxylating P450‐Monooxygenase CYP17A1 Alters Its Regioselectivity
Author(s) -
Morlock Lisa K.,
Grobe Sascha,
Balke Kathleen,
Mauersberger Stephan,
Böttcher Dominique,
Bornscheuer Uwe T.
Publication year - 2018
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201800371
Subject(s) - cyp17a1 , pregnenolone , chemistry , enzyme , residue (chemistry) , active site , regioselectivity , monooxygenase , stereochemistry , protein engineering , substrate (aquarium) , alanine , biochemistry , steroid , biology , cytochrome p450 , amino acid , hormone , ecology , catalysis
The CYP171 enzyme is known to catalyse a key step in the steroidogenesis of mammals. The substrates progesterone and pregnenolone are first hydroxylated at the C17 position, and this is followed by cleavage of the C17−C20 bond to yield important precursors for glucosteroids and androgens. In this study, we focused on the reaction of the bovine CYP17A1 enzyme with progesterone as a substrate. On the basis of a created homology model, active‐site residues were identified and systematically mutated to alanine. In whole‐cell biotransformations, the importance of the N202, R239, G297 and E305 residues for substrate conversion was confirmed. Additionally, mutation of the L206, V366 and V483 residues enhanced the formation of the 16α‐hydroxyprogesterone side product up to 40 % of the total product formation. Furthermore, residue L105 was found not to be involved in this side activity, which contradicts a previous study with the human enzyme.