Formation and properties of [4Fe-4S] clusters on the IscU scaffold protein

Rapid and quantitative reductive coupling of two 2Fe-2S]2+ clusters to form a single 4Fe-4S]2+ cluster on the homodimeric IscU Fe-S cluster scaffold protein has been demonstrated by UV-visible absorption, M?ssbauer, and resonance Raman spectroscopies, using dithionite as the electron donor. Partial reductive coupling was also observed using reduced Isc ferredoxin, which raises the possibility that Isc ferredoxin is the physiological reductant. The results suggest that reductive coupling of adjacent 2Fe-2S]2+ clusters assembled on IscU provides a general mechanism for the final step in the biosynthesis of 4Fe-4S]2+ clusters. The 4Fe-4S]2+ center on IscU can be reduced to a S = 1/24Fe-4S]+ cluster (g parallel = 2.06 and g perpendicular = 1.92), but the low midpoint potential (< -570 mV) and instability of the reduced cluster argue against any physiological relevance for the reduced cluster. On exposure to O2, the 4Fe-4S]2+ cluster on IscU degrades via a semistable 2Fe-2S]2+ cluster with properties analogous to those of the 2Fe-2S]2+ center in 2Fe-2S]2+ IscU. It is suggested that the ability of IscU to accommodate either 2Fe-2S]2+ or 4Fe-4S]2+ clusters in response to cellular redox status and/or oxygen levels may provide an effective way to populate appropriately cluster-loaded forms of IscU for maturation of different types of Fe-S] proteins.