Hemoglobin conformation couples erythrocyte S-nitrosothiol content to O2 gradients.

Journal Article (Journal Article)

It is proposed that the bond between nitric oxide (NO) and the Hb thiol Cys-beta(93) (SNOHb) is favored when hemoglobin (Hb) is in the relaxed (R, oxygenated) conformation, and that deoxygenation to tense (T) state destabilizes the SNOHb bond, allowing transfer of NO from Hb to form other (vasoactive) S-nitrosothiols (SNOs). However, it has not previously been possible to measure SNOHb without extensive Hb preparation, altering its allostery and SNO distribution. Here, we have validated an assay for SNOHb that uses carbon monoxide (CO) and cuprous chloride (CuCl)-saturated Cys. This assay is specific for SNOs and sensitive to 2-5 pmol. Uniquely, it measures the total SNO content of unmodified erythrocytes (RBCs) (SNO(RBC)), preserving Hb allostery. In room air, the ratio of SNO(RBC) to Hb in intact RBCs is stable over time, but there is a logarithmic loss of SNO(RBC) with oxyHb desaturation (slope, 0.043). This decay is accelerated by extraerythrocytic thiol (slope, 0.089; P < 0.001). SNO(RBC) stability is uncoupled from O(2) tension when Hb is locked in the R state by CO pretreatment. Also, SNO(RBC) is increased approximately 20-fold in human septic shock (P = 0.002) and the O(2)-dependent vasoactivity of RBCs is affected profoundly by SNO content in a murine lung bioassay. These data demonstrate that SNO content and O(2) saturation are tightly coupled in intact RBCs and that this coupling is likely to be of pathophysiological significance.

Full Text

Duke Authors

Cited Authors

  • Doctor, A; Platt, R; Sheram, ML; Eischeid, A; McMahon, T; Maxey, T; Doherty, J; Axelrod, M; Kline, J; Gurka, M; Gow, A; Gaston, B

Published Date

  • April 19, 2005

Published In

Volume / Issue

  • 102 / 16

Start / End Page

  • 5709 - 5714

PubMed ID

  • 15824313

Pubmed Central ID

  • PMC556285

International Standard Serial Number (ISSN)

  • 0027-8424

Digital Object Identifier (DOI)

  • 10.1073/pnas.0407490102


  • eng

Conference Location

  • United States