Relative role of heme nitrosylation and β-cysteine 93 nitrosation in the transport and metabolism of nitric oxide by hemoglobin in the human circulation

^*Critical Care Medicine Department of the Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892; and ^§Laboratory of Chemical Biology and ^‡Structural Mass Spectrometry Group, National Institute of Diabetes, Digestive, and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892

Edited by Max F. Perutz, Medical Research Council, Cambridge, United Kingdom, and approved June 22, 2000 (received for review April 6, 2000)

Abstract

To quantify the reactions of nitric oxide (NO) with hemoglobin under physiological conditions and to test models of NO transport on hemoglobin, we have developed an assay to measure NO–hemoglobin reaction products in normal volunteers, under basal conditions and during NO inhalation. NO inhalation markedly raised total nitrosylated hemoglobin levels, with a significant arterial–venous gradient, supporting a role for hemoglobin in the transport and delivery of NO. The predominant species accounting for this arterial–venous gradient is nitrosyl(heme)hemoglobin. NO breathing increases S-nitrosation of hemoglobin β-chain cysteine 93, however only to a fraction of the level of nitrosyl(heme)hemoglobin and without a detectable arterial–venous gradient. A strong correlation between methemoglobin and plasma nitrate formation was observed, suggesting that NO metabolism is a primary physiological cause of hemoglobin oxidation. Our results demonstrate that NO–heme reaction pathways predominate in vivo, NO binding to heme groups is a rapidly reversible process, and S-nitrosohemoglobin formation is probably not a primary transport mechanism for NO but may facilitate NO release from heme.

Footnotes

↵ † To whom reprint requests should be addressed at: Warren G. Magnuson Clinical Center, Critical Care Medicine Department, Building 10, Room 7D43, 10 Center Drive, MSC 1662, Bethesda, Maryland 20892-1662. E-mail: mgladwin{at}nih.gov.
This paper was submitted directly (Track II) to the PNAS office.
Article published online before print: Proc. Natl. Acad. Sci. USA, 10.1073/pnas.180155397.
Article and publication date are at www.pnas.org/cgi/doi/10.1073/pnas.180155397
Abbreviations:

NO,

nitric oxide;

SNO-Hb,

S-nitrosohemoglobin;

Hb(FeII)NO,

nitrosyl(heme)hemoglobin;

FeIII,

methemoglobin