1H NMR Spectroscopy of [FeFe] Hydrogenase: Insight into the Electronic Structure of the Active Site.

Abstract

The [FeFe] hydrogenase HydA1 from Chlamydomonas reinhardtii has been studied using 1H NMR spectroscopy identifying the paramagnetically shifted 1H resonances associated with both the [4Fe-4S]H and the [2Fe]H subclusters of the active site “H-cluster”. The signal pattern of the unmaturated HydA1 containing only [4Fe-4S]H is reminiscent of bacterial-type ferredoxins. The spectra of maturated HydA1, with a complete H-cluster in the active Hox and the CO-inhibited Hox–CO state, reveal additional upfield and downfield shifted 1H resonances originating from the four methylene protons of the azadithiolate ligand in the [2Fe]H subsite. The two axial protons are affected by positive spin density, while the two equatorial protons experience negative spin density. These protons can be used as important probes sensing the effects of ligand-binding to the catalytic site of the H-cluster.