TCB Publications - Abstract

Ilia G. Denisov, Yelena V. Grinkova, Prithviraj Nandigrami, Mrinal S. Shekhar, Emad Tajkhorshid, and Stephen G. Sligar. Allosteric interactions in human cytochrome P450 CYP3A4: The role of phenylalanine 213. Biochemistry, 58:1411-1422, 2019.

DENI2019-ET The role of Phe213 in the allosteric mechanism of human cytochrome P450 CYP3A4 was studied using a combination of progesterone (PGS) and carbamazepine (CBZ) as probe substrates. We expressed, purified and incorporated in POPC Nanodiscs three mutants, F213A, F213S, and F213Y, and compared them with the wild-type CYP3A4 monitoring spectral titration, the rate of NADPH oxidation and steady-state product turnover rates with pure substrates and substrate mixtures. All mutants demonstrated higher activity with CBZ, lower activity with PGS, and reduced activation of CBZ epoxidation by PGS, most pronounced in F213A mutant. Using all-atom molecular dynamics simulations we compared dynamics of WT CYP3A4 and the F213A mutant incorporated in the lipid bilayer and the effect of the presence of PGS molecule at the allosteric peripheral site and evaluated the critical role of Phe213 in mediating the heterotropic allosteric interactions in CYP3A4.


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