Crystallographic snapshots of TsrM, a radical S‐adenosylmethionine enzyme whose reaction is not so radical

Abstract

TsrM methylates C2 of the indole ring of L‐tryptophan (Trp) as the first step in the biosynthesis of the quinaldic acid moiety of thiostrepton, a ribosomally synthesized and post‐translationally modified thiazolyl peptide natural product that exhibits potent in vitro activity against many Gram‐positive bacteria of urgent clinical relevance. TsrM is annotated as a member of the radical S‐adenosylmethionine (SAM) superfamily and exhibits hallmarks of these enzymes, such as a [4Fe–4S] cluster ligated by cysteines residing in a CxxxCxxC motif and a binding site for S‐adenosylmethionine. However, unlike all other characterized members of the radical SAM (RS) superfamily, TsrM does not catalyze a reductive cleavage of SAM to the almost universal 5′‐deoxyadenosyl 5′‐radical intermediate. Instead, recent studies suggest that it catalyzes its reaction via two polar SN2 displacements, an attack on the methyl group of SAM by a cob(I)alamin species and a subsequent attack of C2 of Trp onto methylcobalamin. Herein we report the first crystal structures of a class B RS methylase. The structure contains both the cobalamin and [4Fe‐4S] cluster cofactors bound and show the presence of unique ligands to each. We additionally have solved a structure with substrate analogs bound, which reveals a novel active site, suggesting that TsrM is only masquerading as a radical SAM enzyme.