8VKF

The crystal structure of wild-type CYP199A4 bound to 4-propionylbenzoic acid

Summary for 8VKF

• Entry DOI: 10.2210/pdb8vkf/pdb
• Descriptor: Cytochrome P450, PROTOPORPHYRIN IX CONTAINING FE, 4-propanoylbenzoic acid, ... (6 entities in total)
• Functional Keywords: cytochrome p450, cyp199a4, oxidoreductase
• Biological source: Rhodopseudomonas palustris HaA2
• Total number of polymer chains: 1
• Total formula weight: 45441.86

Authors

Podgorski, M.N.,Bell, S.G. (deposition date: 2024-01-09, release date: 2024-05-08, Last modification date: 2025-02-19)

Primary citation

Lee, J.H.Z.,Coleman, T.,Mclean, M.A.,Podgorski, M.N.,Hayball, E.F.,Stone, I.S.J.,Bruning, J.B.,Whelan, F.,Voss, J.J.,Sligar, S.G.,Bell, S.G.
Selective alpha -Hydroxyketone Formation and Subsequent C-C Bond Cleavage by Cytochrome P450 Monooxygenase Enzymes.
Acs Catalysis, 14:8958-8971, 2024

PubMed Abstract: The heme enzymes of the cytochrome P450 superfamily (CYPs) catalyze oxidation reactions with a high level of selectivity. Here, the CYP199A4 enzyme from the bacterium HaA2 is used to catalyze the hydroxylation of carbonyl-containing compounds to generate -hydroxyketones. Both 4-propionyl- and 4-(2-oxopropyl)-benzoic acids were regioselectively hydroxylated by this enzyme to generate -hydroxyketone metabolites, 4-(2-hydroxypropanoyl)benzoic acid and 4-(1-hydroxy-2-oxopropyl)benzoic acid, respectively, with high stereoselectivity. Co-crystallization of CYP199A4 with each substrate allowed high-resolution X-ray crystal structures of the enzyme bound with both to be determined. These provide a rationale for biochemical observations related to substrate binding and activity. As these versatile enzymes have a demonstrated ability to support carbon-carbon (C-C) bond cleavage (lyase) reactions on -hydroxyketones, we assessed if this activity would be catalyzed by wild-type (WT) CYP199A4. Molecular dynamics (MD) simulations predicted the regioselective hydroxylation of each substrate but indicated that the WT enzyme would not be a good catalyst for lyase activity, in agreement with the experimental observations. The MD simulations also suggested the F182L mutant of CYP199A4 would permit closer approach of the substrate to the ferric-peroxo intermediate, enabling the formation of the lyase transition state. Indeed, this variant was observed to catalyze the cleavage reaction. Furthermore, the F182A variant of CYP199A4 was used to catalyze both the hydroxylation and C-C bond cleavage reactions with both 4-propionyl- and 4-(2-oxopropyl)-benzoic acids using hydrogen peroxide as the oxidant. This dual CYP activity is analogous to that supported by the mammalian CYP17A1 enzyme in steroid biosynthesis.

PubMed: 39911918
DOI: 10.1021/acscatal.4c01766

Experimental method

X-RAY DIFFRACTION (1.7 Å)