Identification of the Cl- transport site of human red blood cells by a kinetic analysis of the inhibitory effects of a chemical probe.
H2DIDS, the dihydro analog of DIDS (4,4'-diisothiocyanostilbene-2,2'-disulfonic acid) can interact covalently with membrane sites, resulting in an irreversible inhibition of anion exchange. At low temperatures (0 degrees C) and for relatively short times, however, its interaction is largely reversible, so that a kinetic analysis of the nature of its inhibitory effect on Cl- self exchange can be performed. The effects of variations in the chloride concentration on the inhibitory potency of H2DIDS are consistent with the concept that Cl- and H2DIDS compete for the transport site of the anion exchange system. The value of Ki for H2DIDS is 0.046 micrometer, indicating that H2DIDS has a higher affinity for the transport system than any other inhibitor so far examined. If, as seems probable, the covalent labelling of H2DIDS occurs at the same site as the reversible binding, H2DIDS can be used as a covalent label for the transport site. The specific localization of H2DIDS in the band-3 protein thus indicates that this protein participates directly in anion exchange.
D001665:Binding Sites; D001692:Biological Transport / Q000187:drug effects; D002712:Chlorides / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004910:Erythrocyte Membrane / Q000502:physiology; D004912:Erythrocytes / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D013267:Stilbenes / Q000378:metabolism / Q000494:pharmacology*
D002712:Chlorides; D008565:Membrane Proteins; D013267:Stilbenes