Iron-sulfur center of biotin synthase and lipoate synthase.
Biotin synthase and lipoate synthase are homodimers that are required for the C-S bond formation at nonactivated carbon in the biosynthesis of biotin and lipoic acid, respectively. Aerobically isolated monomers were previously shown to contain a (2Fe-2S) cluster, however, after incubation with dithionite 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 (4Fe-4S) cluster. Here we report Mössbauer studies of (57)Fe-reconstituted biotin synthase showing that anaerobically prepared enzyme can accommodate two (4Fe-4S) clusters per dimer. The (4Fe-4S) cluster is quantitatively converted into a (2Fe-2S)(2+) cluster upon exposure to air. Reduction of the air-exposed enzyme with dithionite or photoreduced deazaflavin yields again (4Fe-4S) clusters. The (4Fe-4S) cluster is stable in both the 2+ and 1+ oxidation states. The Mössbauer and EPR parameters were DeltaE(q) = 1.13 mm/s and delta = 0.44 mm/s for the diamagnetic (4Fe-4S)(2+) and DeltaE(q) = 0.51 mm/s, delta = 0.85 mm/s, g(par) = 2.035, and g(perp) = 1.93 for the S = (1)/(2) state of (4Fe-4S)(1+). Considering that we find two (4Fe-4S) clusters per dimer, our studies argue against the early proposal that the enzyme contains one (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 [DeltaE(q) = 1.20 mm/s and delta = 0.44 mm/s for the diamagnetic (4Fe-4S)(2+) and g(par) = 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.
Ollagnier-De Choudens
S
S
Laboratoire de Chimie et Biochimie des Centres Rédox Biologiques, DBMS-CB, CEA/CNRS/Université Joseph Fourier, 17 Avenue des Martyrs, 38054 Grenoble Cedex 09, France.
Sanakis
Y
Y
Hewitson
K S
KS
Roach
P
P
Baldwin
J E
JE
Münck
E
E
Fontecave
M
M
D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D004926:Escherichia coli; D007501:Iron; D013379:Substrate Specificity; D013455:Sulfur; D013466:Sulfurtransferases / Q000737:chemistry*
D001426:Bacterial Proteins; C079999:LipA protein, Bacteria; D013455:Sulfur; D007501:Iron; D013466:Sulfurtransferases; C057985:biotin synthetase