Assignment of haem ligands and detection of electronic absorption bands of molybdenum in the di-haem periplasmic nitrate reductase of Paracoccus pantotrophus.
The periplasmic nitrate reductase (NAP) from Paracoccus pantotrophus is a soluble two-subunit enzyme (NapAB) that binds two c-type haems, a [4Fe-4S] cluster and a bis-molybdopterin guanine dinucleotide cofactor that catalyses the reduction of nitrate to nitrite. In the present work the NapAB complex has been studied by magneto-optical spectroscopy to probe co-ordination of both the NapB haems and the NapA active site Mo. The absorption spectrum of the NapAB complex is dominated by features from the NapB c-type cytochromes. Using a combination of electron paramagnetic resonance spectroscopy and magnetic circular dichroism it was demonstrated that both haems are low-spin with bis-histidine axial ligation. In addition, a window between 600 and 800 nm was identified in which weak absorption features that may arise from Mo could be detected. The low-temperature MCD spectrum shows oppositely signed bands in this region (peak 648 nm, trough 714 nm) which have been assigned to S-to-Mo(V) charge transfer transitions.
Butler
C S
CS
Centre for Metalloprotein Spectroscopy and Biology, School of Biological and Chemical Sciences, University of East Anglia, Norwich NR4 7TJ, UK.
Ferguson
S J
SJ
Berks
B C
BC
Thomson
A J
AJ
Cheesman
M R
MR
Richardson
D J
DJ
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D004578:Electron Spin Resonance Spectroscopy; D006418:Heme / Q000737:chemistry; D008969:Molecular Sequence Data; D008982:Molybdenum / Q000737:chemistry*; D050901:Nitrate Reductase; D009565:Nitrate Reductases / Q000737:chemistry*; D010230:Paracoccus / Q000737:chemistry / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid
D006418:Heme; D008982:Molybdenum; D009565:Nitrate Reductases; D050901:Nitrate Reductase