Structural analysis of cofactor binding for a prolyl 4-hydroxylase from the pathogenic bacterium Bacillus anthracis.

Abstract

The prolyl 4-hydroxylases (P4Hs) are mononuclear nonheme iron enzymes that catalyze the formation of 4R-hydroxyproline from many different substrates, with various biological implications. P4H is a key player in collagen accumulation, which has implications in fibrotic disorders. The stabilization of collagen triple-helical structure via prolyl hydroxylation is the rate-limiting step in collagen biosynthesis, and therefore P4H has been extensively investigated as a potential therapeutic target of fibrotic disease. Understanding how these enzymes recognize cofactors and substrates is important and will aid in the future design of inhibitors of P4H. In this article, X-ray crystal structures of a metallocofactor- and α-ketoglutarate (αKG)-bound form of P4H from Bacillus anthracis (BaP4H) are reported. Structures of BaP4H were solved at 1.63 and 2.35 Å resolution and contained a cadmium ion and αKG bound in the active site. The αKG-Cd-BaP4H ternary complex reveals conformational changes of conserved residues upon the binding of metal ion and αKG, resulting in a closed active-site configuration required for dioxygen, substrate binding and catalysis.