Spectroscopic
Investigations of [FeFe] Hydrogenase
Maturated with [<sup>57</sup>Fe<sub>2</sub>(adt)(CN)<sub>2</sub>(CO)<sub>4</sub>]<sup>2–</sup>
- Publication date
- Publisher
Abstract
The preparation and spectroscopic
characterization of a CO-inhibited
[FeFe] hydrogenase with a selectively <sup>57</sup>Fe-labeled binuclear
subsite is described. The precursor [<sup>57</sup>Fe<sub>2</sub>(adt)(CN)<sub>2</sub>(CO)<sub>4</sub>]<sup>2–</sup> was synthesized from
the <sup>57</sup>Fe metal, S<sub>8,</sub> CO, (NEt<sub>4</sub>)CN,
NH<sub>4</sub>Cl, and CH<sub>2</sub>O. (Et<sub>4</sub>N)<sub>2</sub>[<sup>57</sup>Fe<sub>2</sub>(adt)(CN)<sub>2</sub>(CO)<sub>4</sub>] was then used for the maturation of the [FeFe] hydrogenase HydA1
from <i>Chlamydomonas reinhardtii</i>, to yield the enzyme selectively labeled at the [2Fe]<sub>H</sub> subcluster. Complementary <sup>57</sup>Fe enrichment of the
[4Fe-4S]<sub>H</sub> cluster was realized by reconstitution with <sup>57</sup>FeCl<sub>3</sub> and Na<sub>2</sub>S. The H<sub>ox</sub>-CO
state of [2<sup>57</sup>Fe]<sub>H</sub> and [4<sup>57</sup>Fe-4S]<sub>H</sub> HydA1 was characterized by Mössbauer, HYSCORE, ENDOR,
and nuclear resonance vibrational spectroscopy
