Solvent accessibility in the distal heme pocket of the nitrosyl d(1)-heme complex of Pseudomonas stutzeri cd(1) nitrite reductase.
In nitrite reductase (cd(1) NIR), the c-heme mediates electron transfer to the catalytic d(1)-heme where nitrite (NO(2)(-)) is reduced to nitric oxide (NO). An interesting feature of this enzyme is the relative lability of the reaction product NO bound to the d(1)-heme. Marked differences in the c- to d(1)-heme electron-transfer rates were reported for cd(1) NIRs from different sources, such as Pseudomonas stutzeri (P. stutzeri) and Pseudomonas aeruginosa (P. aeruginosa). The three-dimensional structure of the P. aeruginosa enzyme has been determined, but that of the P. stutzeri enzyme is still unknown. The difference in electron transfer rates prompted a comparison of the structural properties of the d(1)-heme pocket of P. stutzeri cd(1) NIR with those of the P. aeruginosa wild type enzyme (WT) and its Y10F using their nitrosyl d(1)-heme complexes. We applied high field pulse electron paramagnetic resonance (EPR) techniques that detect nuclear spins in the close environment of the spin bearing Fe(II)-NO entity. We observed similarities in the rhombic g-tensor and detected a proximal histidine ligand with (14)N hyperfine and quadrupole interactions also similar to those of P. aeruginosa WT and Y10F mutant complexes. In contrast, we also observed significant differences in the H-bond network involving the NO ligand and a larger solvent accessibility for P. stutzeri attributed to the absence of this tyrosine residue. For P. aeruginosa, cd(1) NIR domain swapping allows Tyr(10) to become H-bonded to the bound NO substrate. These findings support a previous suggestion that the large difference in the c- to d(1)-heme electron transfer rates between the two enzymes is related to solvent accessibility of their d(1)-heme pockets.
Department of Chemical Physics, Weizmann Institute of Science, Israel.
Marina
M
Radoul
Yoav
Y
Barak
Serena
S
Rinaldo
Francesca
F
Cutruzzolà
Israel
I
Pecht
Daniella
D
Goldfarb
D003580:Cytochromes / Q000737:chemistry*; D041961:Deuterium Exchange Measurement; D004578:Electron Spin Resonance Spectroscopy; D006418:Heme / Q000737:chemistry*; D009572:Nitrite Reductases / Q000737:chemistry*; D011550:Pseudomonas aeruginosa / Q000201:enzymology; D044223:Pseudomonas stutzeri / Q000201:enzymology; D013056:Spectrophotometry, Ultraviolet
D003580:Cytochromes; D006418:Heme; D009572:Nitrite Reductases; C028123:nitrite reductase (NO-forming)