Reaction Coordinate Leading to H_2 Production in [FeFe]-Hydrogenase Identified by Nuclear Resonance Vibrational Spectroscopy and Density Functional Theory

摘要

[FeFe]-hydrogenases are metalloenzymes that reversibly reduce protons to molecular hydrogen at exceptionally high rates. We have characterized the catalytically competent hydride state (H_(hyd)) in the [FeFe]-hydrogenases from both Chlamydomonas reinhardtii and Desulfovibrio desulfuricans using ~(57)Fe nuclear resonance vibrational spectroscopy (NRVS) and density functional theory (DFT). H/D exchange identified two Fe-H bending modes originating from the binuclear iron cofactor. DFT calculations show that these spectral features result from an iron-bound terminal hydride, and the Fe-H vibrational frequencies being highly dependent on interactions between the amine base of the catalytic cofactor with both hydride and the conserved cysteine terminating the proton transfer chain to the active site. The results indicate that H_(hyd) is the catalytic state one step prior to H_2 formation. The observed vibrational spectrum, therefore, provides mechanistic insight into the reaction coordinate for H_2 bond formation by [FeFe]-hydrogenases.