title: Kinetic solvent isotope effect in steady-state turnover by CYP19A1 suggests involvement of Compound 1 for both hydroxylation and aromatization steps.
abstract: CYP19A1, or human aromatase catalyzes the conversion of androgens to estrogens in a three-step reaction through the formation of 19-hydroxy and 19-aldehyde intermediates. While the first two steps of hydroxylation are thought to proceed through a high-valent iron-oxo species, controversy exists surrounding the identity of the reaction intermediate that catalyzes the lyase and aromatization reaction. We investigated the kinetic isotope effect on the steady-state turnover of Nanodisc-incorporated human CYP19A1 to explore the mechanisms of this reaction. Our experiments reveal a significant (∼ 2.5) kinetic solvent isotope effect for the C10-C19 lyase reaction, similar to that of the first two hydroxylation steps (2.7 and 1.2). These data implicate the involvement of Compound 1 as a reactive intermediate in the final aromatization step of CYP19A1.
authors.affiliation: Department of Biochemistry, University of Illinois Urbana-Champaign, 505 S. Goodwin Avenue, Urbana, IL 61801, United States.
authors.forename: Yogan
authors.initials: Y
authors.lastname: Khatri
authors.affiliation: Department of Biochemistry, University of Illinois Urbana-Champaign, 505 S. Goodwin Avenue, Urbana, IL 61801, United States.
authors.forename: Abhinav
authors.initials: A
authors.lastname: Luthra
authors.affiliation: Department of Biochemistry, University of Illinois Urbana-Champaign, 505 S. Goodwin Avenue, Urbana, IL 61801, United States.
authors.forename: Ruchia
authors.initials: R
authors.lastname: Duggal
authors.affiliation: Department of Biochemistry, University of Illinois Urbana-Champaign, 505 S. Goodwin Avenue, Urbana, IL 61801, United States. Electronic address: s-sligar@illinois.edu.
authors.forename: Stephen G
authors.initials: SG
authors.lastname: Sligar
mesh_terms: D001141:Aromatase / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D006900:Hydroxylation; D007700:Kinetics; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D010100:Oxygen / Q000737:chemistry; D012997:Solvents / Q000737:chemistry*; D014867:Water / Q000737:chemistry
chemical_list: D012997:Solvents; D014867:Water; D009249:NADP; D001141:Aromatase; C561410:CYP19A1 protein, human; D010100:Oxygen
keywords: CYP19A1; C–C lyase; Human aromatase; KSIE; Steady-state kinetics