Crystal Structure of the [FeFe]-Hydrogenase Maturase HydE Bound to Complex-B

[FeFe]-hydrogenases use a unique organometallic complex, termed the H cluster, to reversibly convert H₂ into protons and low-potential electrons. It can be best described as a [Fe₄S₄] cluster coupled to a unique [2Fe]_H center where the reaction actually takes place. The latter corresponds to two iron atoms, each of which is bound by one CN^- ligand and one CO ligand. The two iron atoms are connected by a unique azadithiolate molecule (^-S-CH₂-NH-CH₂-S^-) and an additional bridging CO. This [2Fe]_H center is built stepwise thanks to the well-orchestrated action of maturating enzymes that belong to the Hyd machinery. Among them, HydG converts l-tyrosine into CO and CN^- to produce a unique l-cysteine-Fe(CO)₂CN species termed complex-B. Very recently, HydE was shown to perform radical-based chemistry using synthetic complex-B as a substrate. Here we report the high-resolution crystal structure that establishes the identity of the complex-B-bound HydE. By triggering the reaction prior to crystallization, we trapped a new five-coordinate Fe species, supporting the proposal that HydE performs complex modifications of complex-B to produce a monomeric "SFe(CO)₂CN" precursor to the [2Fe]_H center. Substrate access, product release, and intermediate transfer are also discussed.