Purification and characterization of bisulfite reductase (desulfofuscidin) from Desulfovibrio thermophilus and its complexes with exogenous ligands.
A dissimilatory bisulfite reductase has been purified from a thermophilic sulfate-reducing bacterium Desulfovibrio thermophilus (DSM 1276) and studied by EPR and optical spectroscopic techniques. The visible spectrum of the purified bisulfite reductase exhibits absorption maxima at 578.5, 392.5 and 281 nm with a weak band around 700 nm. Photoreduction of the native enzyme causes a decrease in absorption at 578.5 nm and a concomitant increase in absorption at 607 nm. When reduced, the enzyme reacts with cyanide, sulfite, sulfide and carbon monoxide to give stable complexes. The EPR spectrum of the native D. thermophilus bisulfite reductase shows the presence of a high-spin ferric signal with g values at 7.26, 4.78 and 1.92. Upon photoreduction the high-spin ferric heme signal disappeared and a typical 'g = 1.94' signal of [4Fe-4S] type cluster appeared. Chemical analyses show that the enzyme contains four sirohemes and eight [4Fe-4S] centers per mol of protein. The molecular mass determined by gel filtration was found to be 175 kDa. On SDS-gel electrophoresis the enzyme presents a main band of 44 to 48 kDa. These results suggest that the bisulfite reductase contains probably one siroheme and two [4Fe-4S] centers per monomer. The dissimilatory bisulfite reductase from D. thermophilus presents some homologous properties with desulfofuscidin, the bisulfite reductase isolated from Thermodesulfobacterium commune (Hatchikian, E.C. and Zeikus, J.G. (1983) J. Bacteriol. 153, 1211-1220).
Fauque
G
G
Département de Biologie, C.E.N. Cadarache, Saint Paul lez Durance, France.
Lino
A R
AR
Czechowski
M
M
Kang
L
L
DerVartanian
D V
DV
Moura
J J
JJ
LeGall
J
J
Moura
I
I
D000596:Amino Acids / Q000032:analysis; D003901:Desulfovibrio / Q000201:enzymology* / Q000254:growth & development; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D008024:Ligands; D046911:Macromolecular Substances; D008970:Molecular Weight; D010088:Oxidoreductases / Q000302:isolation & purification*; D050862:Oxidoreductases Acting on Sulfur Group Donors / Q000302:isolation & purification* / Q000378:metabolism; D011485:Protein Binding; D013045:Species Specificity; D013053:Spectrophotometry
D000596:Amino Acids; D008024:Ligands; D046911:Macromolecular Substances; D010088:Oxidoreductases; D050862:Oxidoreductases Acting on Sulfur Group Donors