Evidence from Mössbauer spectroscopy for distinct [2Fe-2S](2+) and [4Fe-4S](2+) cluster binding sites in biotin synthase from Escherichia coli.
Biotin synthase is an AdoMet-dependent radical enzyme that catalyzes the insertion of an FeS cluster-derived sulfur atom into dethiobiotin. The dimeric enzyme is purified containing one [2Fe-2S]2+ cluster per monomer, but it is most active when reconstituted with an additional [4Fe-4S]2+ cluster per monomer. Using Mössbauer spectroscopy coupled with differential reconstitution of each cluster with 57Fe, we show that the reconstituted enzyme has approximately 1:1 [2Fe-2S]2+ and [4Fe-4S]2+ clusters and that the [4Fe-4S]2+ cluster is assembled at an alternate site not previously occupied by the [2Fe-2S]2+ cluster. These data suggest that biotin synthase is evolved to simultaneously accommodate two different clusters with unique roles in catalysis.
Ugulava
Natalia B
NB
Johnson Research Foundation and Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Surerus
Kristene K
KK
Jarrett
Joseph T
JT
D001426:Bacterial Proteins / Q000737:chemistry; D001665:Binding Sites; D003470:Culture Media; D004926:Escherichia coli / Q000201:enzymology*; D007501:Iron / Q000737:chemistry*; D015204:Spectroscopy, Mossbauer; D013440:Sulfides / Q000737:chemistry*; D013466:Sulfurtransferases / Q000737:chemistry*
D001426:Bacterial Proteins; D003470:Culture Media; D013440:Sulfides; C011342:pyrite; D007501:Iron; D013466:Sulfurtransferases; C057985:biotin synthetase