Neuroglobin: enzymatic reduction and oxygen affinity.
Neuroglobin (Ngb) is a hexacoordinate globin expressed in the nervous system of vertebrates, involved in neuroprotection. O(2) equilibrium measurements on mouse Ngb yielded significantly different P(50) values, ranging from approximately 2 torr to approximately 10 torr. By a kinetic approach minimizing the effects of protein autoxidation, we measured P(50)=2.2 torr at 20 degrees C. As predicted from the structure, O(2) binds to the Y44D Ngb mutant more quickly (k=2.2s(-1) vs 0.15s(-1)) and with slightly higher affinity (P(50)=1.3 torr) than wild-type. In addition, we introduced a novel reduction protocol for metNgb based on NADH:flavorubredoxin oxidoreductase (FlRd-red) from Escherichia coli, a candidate for the Ngb reducing activity recently identified in E. coli extracts. Interestingly, E. coli FlRd-red shares sequence similarity with the FAD-binding domain of the human apoptosis-inducing factor, a finding which may have unexpected significance with reference to the mechanism of neuroprotection by Ngb.
Giuffrè
Alessandro
A
Department of Biochemical Sciences and CNR Institute of Molecular Biology and Pathology, Sapienza University of Rome, I-00185 Rome, Italy.
Moschetti
Tommaso
T
Vallone
Beatrice
B
Brunori
Maurizio
M
D000818:Animals; D001665:Binding Sites; D004798:Enzymes / Q000737:chemistry*; D005914:Globins / Q000737:chemistry*; D051379:Mice; D009419:Nerve Tissue Proteins / Q000737:chemistry*; D000078787:Neuroglobin; D010084:Oxidation-Reduction; D010100:Oxygen / Q000737:chemistry*; D011485:Protein Binding
D004798:Enzymes; D009419:Nerve Tissue Proteins; D000078787:Neuroglobin; D005914:Globins; D010100:Oxygen