Iron-sulfur center of biotin synthase and lipoate synthase

Biotin synthase and lipoate synthase are homodimers that are required for t he C-S bond formation at nonactivated carbon in the biosynthesis of biotin and lipoic acid, respectively. Aerobically isolated monomers were previousl y shown to contain a (2Fe-2S) cluster, however, after incubation with dithi onite one (4Fe-4S) cluster per dimer was obtained, suggesting that two (2Fe -2S) clusters had combined at the interface of the subunits to form the (4F e-4S) cluster. Here we report Mossbauer studies of Fe-57-reconstituted biot in synthase showing that anaerobically prepared enzyme can accommodate two (4Fe-4S) clusters per dimer. The (4Fe-4S) cluster is quantitatively convert ed into a (2Fe-2S)(2+) cluster upon exposure to air. Reduction of the air-e xposed enzyme with dithionite or photoreduced deazaflavin yields again (4Fe -4S) clusters. The (4Fe-4S) cluster is stable in both the 2+ and If oxidati on states. The Mossbauer and EPR parameters were Delta E-q = 1.13 mm/s and delta = 0.44 mm/s for the diamagnetic (4Fe-4S)(1+) and Delta E-q = 0.51 mm/ s, delta = 0.85 mm/s, delta = 2.035, and g(perp) = 1.93 for the S = 1/2 sta te of (4Fe-4S)(1+). Considering that we find two (4Fe-4S) clusters per dime r, our studies argue against the early proposal that the enzyme contains on e (4Fe-4S) cluster bridging the two subunits. Our study of lipoate synthase gave results similar to those obtained for BS, under strict anaerobiosis, lipoate synthase can accommodate a (4Fe-4S) cluster per subunit [Delta E-q = 1.20 mm/s and delta = 0.44 mm/s for the diamagnetic (4Fe-4S)(2+) and g(pa r) = 2.039 and g(perp) = 1.93 for the S = 1/2 state of (4Fe-4S)(1+)], which reacts with oxygen to generate a (2Fe-2S)(2+) center.