Mechanism of anion exchange across the red cell membrane by band 3: interactions between stilbenedisulfonate and NAP-taurine binding sites.
Anion exchange across the erythrocyte membrane can be inhibited competitively by stilbenedisulfonates, which bind to the external transport of the band 3 protein, and noncompetitively by external NAP-taurine [2-[N-(4-azido-2-nitrophenyl)amino]ethanesulfonate], which it has been suggested binds to a "modifier site" [Knauf, P. A., Ship. S., Breur, W., MCCulloch, L., & Rothstein, A. (1978) J. Gen. Physiol. 72, 604-630]. The binding of the two types of inhibitor of erythrocyte membranes is shown in the present study to be competitive, indicating that binding to the same subunit of band 3 is mutually exclusive. Covalent labeling of red cells with a stilbenedisulfonate [4-benzamido-4'-isothiocyano-stilbene-2,2'-disulfonate (BIDS)] to 80% saturation had no detectable effect upon the Ki for inhibition of [32P]phosphate influx by NAP-taurine, indicating that when bound to adjacent subunits in the band 3 dimer, the two types of inhibitor do not interact. In addition to the external NAP-taurine site, a second high-affinity NAP-taurine site (Kd = 15 microM) was detected on the cytoplasmic side of red cell membranes. This site is less than 51 A from the disulfonic stilbene binding site, as judged by fluorescence resonance energy transfer from BIDS to NAP-taurine. Binding at this site is not affected by covalent attachment of BIDS, and no clear role for this site in transport could be determined. On the basis of these studies we present a model indicating that disulfonic stilbenes bind to a site which overlaps both the anion transport site and the modifier site on a band 3 monomer and suggests that the modifier site may be part of a transporting gate.
D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000378:metabolism*; D001457:Anion Exchange Protein 1, Erythrocyte; D001665:Binding Sites; D001667:Binding, Competitive; D001693:Biological Transport, Active / Q000187:drug effects; D001798:Blood Proteins / Q000378:metabolism*; D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D008956:Models, Chemical; D013050:Spectrometry, Fluorescence; D013267:Stilbenes / Q000378:metabolism*; D013654:Taurine / Q000031:analogs & derivatives* / Q000378:metabolism
D001457:Anion Exchange Protein 1, Erythrocyte; D001798:Blood Proteins; D008565:Membrane Proteins; D013267:Stilbenes; D013654:Taurine; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; C010861:NAP taurine; C032526:4-benzamido-4'-isothiocyanostilbene-2,2'-disulfonate