Structural investigation of the molybdenum site of the periplasmic nitrate reductase from Thiosphaera pantotropha by X-ray absorption spectroscopy.
The molybdenum centre of the periplasmic respiratory nitrate reductase from the denitrifying bacterium Thiosphaera pantotropha has been probed using molybdenum K-edge X-ray absorption spectroscopy. The optimum fit of the Mo(VI) EXAFS suggests two ==O, three -S- and either a fourth -S- or an -O-/-N- as molybdenum ligands in the ferricyanide-oxidized enzyme. Three of the -S- ligands are proposed to be the two sulphur atoms of the molybdopterin dithiolene group and Cys-181. Comparison of the EXAFS of the ferricyanide-oxidized enzyme with that of a nitrate-treated sample containing 30% Mo(V) suggests that the Mo(VI)-->Mo(V) reduction is accompanied by conversion of one ==O to -O-. The best fit to the Mo(IV) EXAFS of dithionite-reduced enzyme was obtained using one ==O, one -O- and four -S-/-Cl ligands. The periplasmic nitrate reductase molybdenum co-ordination environment in both the Mo(VI) and Mo(IV) oxidation states is distinct from that found in the membrane-bound respiratory nitrate reductase.
Bennett
B
B
Centre for Metalloprotein Spectroscopy and Biology, School of Chemical Sciences, University of East Anglia, University Plain, Norwich, UK.
Charnock
J M
JM
Sears
H J
HJ
Berks
B C
BC
Thomson
A J
AJ
Ferguson
S J
SJ
Garner
C D
CD
Richardson
D J
DJ
D000595:Amino Acid Sequence; D005292:Ferricyanides / Q000494:pharmacology; D005583:Fourier Analysis; D006091:Gram-Negative Chemolithotrophic Bacteria / Q000201:enzymology*; D008024:Ligands; D008667:Metalloproteins / Q000737:chemistry* / Q000187:drug effects; D008956:Models, Chemical; D008969:Molecular Sequence Data; D008982:Molybdenum / Q000737:chemistry*; D050901:Nitrate Reductase; D009565:Nitrate Reductases / Q000737:chemistry* / Q000187:drug effects; D009566:Nitrates / Q000494:pharmacology; D010084:Oxidation-Reduction; D017386:Sequence Homology, Amino Acid; D013057:Spectrum Analysis; D014965:X-Rays
D005292:Ferricyanides; D008024:Ligands; D008667:Metalloproteins; D009566:Nitrates; C007931:hexacyanoferrate III; D008982:Molybdenum; D009565:Nitrate Reductases; D050901:Nitrate Reductase