ELECTRON-SPIN ECHO SPECTROSCOPY OF THE [FE]-HYDROGENASE FROM DESULFOVIBRIO-DESULFURICANS, STRAIN ATCC-7757

Hydrogenase from the sulfate-reducing bacterium Desulfovibrio desulfuricans, strain ATCC 7757, contains three iron-sulfur clusters: two [4Fe-4S] 'F' clusters and one 'H' cluster, believed to represent the active site for reaction with hydrogen. Electron spin-echo envelope modulation (ESEEM) in pulsed electron paramagnetic resonance (EPR) was applied to these clusters, in their paramagnetic oxidation states. Three-pulse, stimulated-echo spectra of the oxidized H-cluster showed modulations indicative of cancellation of the nuclear hyperfine and Zeeman interactions, permitting estimation of the quadrupole coupling. The value for the quadrupole coupling constant is unusually high. These data are compared with previous results from the hydrogenase of Clostridium pasteurianum, and show that the nitrogen coordination is conserved between these distantly related species. Spectra of samples exchanged into (H2O)-H-2 showed deep nuclear modulations for the reduced F clusters, and weaker modulations for the oxidized H clusters. This is evidence for exchangeable protons in close proximity to Fe-S clusters.