HydE Catalytic Mechanism Is Powered by a Radical Relay with Redox-Active Fe(I)-Containing Intermediates.

Abstract

[FeFe]-hydrogenases are enzymes that catalyze the redox interconversion of H⁺ and H₂ using a six-iron active site, known as the H-cluster, which consists of a structurally unique [2Fe]_H subcluster linked to a [4Fe-4S]_H subcluster. A set of enzymes, HydG, HydE, and HydF, are responsible for the biosynthesis of the [2Fe]_H subcluster. Among them, it is well established that HydG cleaves tyrosine into CO and CN^- and forms a mononuclear [Fe(II)(Cys)(CO)₂(CN)] complex. Recent work using EPR spectroscopy and X-ray crystallography show that HydE uses this organometallic Fe complex as its native substrate. The low spin Fe(II) center is reduced into an adenosylated Fe(I) species, which is proposed to form an Fe(I)Fe(I) dimer within HydE. The highly unusual transformation catalyzed by HydE draws interest in both biochemistry and organometallic chemistry. Due to the instability of the substrate, the intermediates, and the proposed product, experimental characterization of the detailed HydE mechanism and its final product is challenging. Herein, the catalytic mechanism of HydE is studied using hybrid quantum mechanics/molecular mechanics (QM/MM) molecular dynamics simulations. A radical relay mechanism was found for the cleavage of the cysteine S-Cβ bond that is energetically favored with respect to a closed-shell mechanism involving unconventional proton transfer. In addition, we propose a pathway for the dimerization of two Fe(I) complexes within the HydE hydrophobic cavity, which is consistent with the recent experimental result that HydF can perform [FeFe]-hydrogenase maturation with a synthetic dimer complex as the substrate. These simulation results take us further down the path to a more complete understanding of these enzymes that synthesize one of Nature's most efficient energy conversion catalysts.