Characterization of the [4Fe-4S]+ cluster at the active site of aconitase by 57Fe, 33S, and 14N electron nuclear double resonance spectroscopy.

57 Fe, 33 S, and 14 N electron nuclear double resonance (ENDOR) studies have been performed to characterize the [4Fe-4S] + cluster at the active site of aconitase. Q-band 57 Fe ENDOR of isotopically enriched enzyme, both substrate free and in the enzyme-substrate complex, reveals four inequivalent iron sites. In agreement with Mossbauer studies, one of the iron ions, Fe a , which is easily removed by oxidation to yield the [3Fe-4S] + cluster of inactive aconitase, shows a dramatic change in the presence of substrate. The remaining iron sites, Fe b1,2,3 , show minor changes when substrate is bound.