Structure, Mutagenesis, and QM:MM Modeling of 3-Ketosteroid Δ¹-Dehydrogenase from Sterolibacterium denitrificans─The Role of a New Putative Membrane-Associated Domain and Proton-Relay System in Catalysis.

ABSTRACT: 3-Ketosteroid Δ¹-dehydrogenases (KstD) are important microbial flavin enzymes that initiate the metabolism of steroid ring A and find application in the synthesis of steroid drugs. We present a structure of the KstD from Sterolibacterium denitrificans (AcmB), which contains a previously uncharacterized putative membrane-associated domain and extended proton-relay system. The experimental and theoretical studies show that the steroid Δ¹-dehydrogenation proceeds according to the Ping-Pong bi-bi kinetics and a two-step base-assisted elimination (E2cB) mechanism. The mechanism is validated by evaluating the experimental and theoretical kinetic isotope effect for deuterium-substituted substrates. The role of the active-site residues is quantitatively assessed by poi

SUBMITTER: Wojcik P 

PROVIDER: S-EPMC9960185 | biostudies-literature | 2023 Feb

REPOSITORIES: biostudies-literature