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Spectroscopic Investigations of [FeFe] Hydrogenase Maturated with [57Fe2(adt)(CN)2(CO)4]2–

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journal contribution
posted on 22.07.2015, 00:00 by Ryan Gilbert-Wilson, Judith F. Siebel, Agnieszka Adamska-Venkatesh, Cindy C. Pham, Edward Reijerse, Hongxin Wang, Stephen P. Cramer, Wolfgang Lubitz, Thomas B. Rauchfuss
The preparation and spectroscopic characterization of a CO-inhibited [FeFe] hydrogenase with a selectively 57Fe-labeled binuclear subsite is described. The precursor [57Fe2(adt)­(CN)2(CO)4]2– was synthesized from the 57Fe metal, S8, CO, (NEt4)­CN, NH4Cl, and CH2O. (Et4N)2[57Fe2(adt)­(CN)2(CO)4] was then used for the maturation of the [FeFe] hydrogenase HydA1 from Chlamydomonas reinhardtii, to yield the enzyme selectively labeled at the [2Fe]H subcluster. Complementary 57Fe enrichment of the [4Fe-4S]H cluster was realized by reconstitution with 57FeCl3 and Na2S. The Hox-CO state of [257Fe]H and [457Fe-4S]H HydA1 was characterized by Mössbauer, HYSCORE, ENDOR, and nuclear resonance vibrational spectroscopy.