Nitric oxide binding to human ferrihemoglobins cross-linked between either alpha or beta subunits.
We have examined the interactions between nitric oxide (NO) and oxidized human hemoglobin, comparing the behavior of unmodified HbA0 with that of two chemically modified hemoglobins. The latter are promising red cell substitute candidates due to their lower oxygen affinity and greater stability as tetramers. The modified forms examined were HbA-DBBF, cross-linked between the alpha chains with bis(3,5-dibromosalicyl) fumarate, and HbA-FMDA, modified between the beta chains with fumaryl monodibromoaspirin. NO binding to the oxidized forms of these hemoglobins is biphasic, due to the differing reactivities of alpha and beta chains. The structural modifications result in altered rate constants for NO binding to both alpha and beta chains. The affinity of the ferric hemes for NO is not correlated with their oxygen affinities in the ferrous state. In a much slower first-order process, the ferric hemes of HbA become reduced. Faster and more heterogeneous kinetics are observed for reduction of the modified hemoglobins. These results may have physiological relevance, since endogenously produced NO is now recognized to play an important role in the relaxation of vascular smooth muscles. If present in vivo, cell-free hemoglobins exposed to NO become rapidly oxidized. Our results show that subsequent interactions of NO with ferrihemoglobin can result in redox cycling. This has the potential of depleting NO and further altering vascular tone with rates dependent on structural parameters of the ferrihemoglobin that are not determined by oxygen affinity.
Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland 20892.
A I
AI
Alayash
J C
JC
Fratantoni
C
C
Bonaventura
J
J
Bonaventura
R E
RE
Cashon
D001241:Aspirin / Q000031:analogs & derivatives; D003432:Cross-Linking Reagents; D004355:Drug Stability; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008706:Methemoglobin / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D013053:Spectrophotometry
D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; C021630:bis(3,5-dibromosalicyl)fumarate; D009569:Nitric Oxide; D008706:Methemoglobin; C059161:mono(3,5-dibromosalicyl)fumarate; D001241:Aspirin; D010100:Oxygen