Rapid electrogenic sulfate-chloride exchange mediated by chemically modified band 3 in human erythrocytes.
One of the modes of action of the red blood cell anion transport protein is the electrically silent net exchange of 1 Cl- for 1 SO4= and 1 H+. Net SO4(=)-Cl- exchange is accelerated by low pH or by conversion of the side chain of glutamate 681 into an alcohol by treatment of intact cells with Woodward's reagent K (WRK) and BH4-. The studies described here were performed to characterize the electrical properties of net SO4(=)-Cl- exchange in cells modified with WRK/BH4-. The SO4= conductance measured in 100 mM SO4= medium is smaller in modified cells than in control cells. However, the efflux of [35S] SO4= into a 150-mM KCl medium is 80-fold larger in modified cells than in control cells and is inhibited 99% by 10 microM H2DIDS. No detectable H+ flux is associated with SO4(=)-Cl- exchange in modified cells. In the presence of gramicidin to increase the cation permeability, the stoichiometry of SO4(=)-Cl- exchange is not distinguishable from 1:1. In modified cells loaded with SO4=, the valinomycin-mediated efflux of 86Rb+ into an Na-gluconate medium is immediately stimulated by the addition of 5 mM extracellular Cl-. Therefore, SO4(=)-Cl- exchange in modified cells causes an outward movement of negative charge, as expected for an obligatory 1:1 SO4(=)-Cl- exchange. This is the first example of an obligatory, electrogenic exchange process in band 3 and demonstrates that the coupling between influx and efflux does not require that the overall exchange be electrically neutral. The effects of membrane potential on SO4(=)-SO4= exchange and SO4(=)-Cl- exchange in modified cells are consistent with a model in which nearly a full net positive charge moves inward through the transmembrane field during the inward Cl- translocation event, and a small net negative charge moves with SO4= during the SO4= translocation event. This result suggests that, in normal cells, the negative charge on Glu 681 traverses most of the transmembrane electric field, accompanied by Cl- and the equivalent of two protein-bound positive charges.
Jennings
M L
ML
Department of Physiology and Biophysics, University of Texas Medical Branch, Galveston 77555, USA.
D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry / Q000378:metabolism*; D001894:Borohydrides / Q000737:chemistry; D002712:Chlorides / Q000097:blood*; D004912:Erythrocytes / Q000187:drug effects / Q000378:metabolism*; D005110:Extracellular Space / Q000378:metabolism; D006096:Gramicidin / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007202:Indicators and Reagents / Q000737:chemistry; D007474:Ion Exchange; D007555:Isoxazoles / Q000737:chemistry; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D012414:Rubidium Radioisotopes; D013431:Sulfates / Q000097:blood*; D014634:Valinomycin / Q000494:pharmacology
D001457:Anion Exchange Protein 1, Erythrocyte; D001894:Borohydrides; D002712:Chlorides; D007202:Indicators and Reagents; D007555:Isoxazoles; D012414:Rubidium Radioisotopes; D013431:Sulfates; D006096:Gramicidin; D014634:Valinomycin; C009686:N-ethyl-5-phenylisoxazolium-3'-sulfonate