Chloride-bicarbonate exchange through the human red cell ghost membrane monitored by the fluorescent probe 6-methoxy-N-(3-sulfopropyl)quinolinium

Analytical Biochemistry
T M Calafut, J A Dix

Abstract

The exchange of chloride and bicarbonate across the human red cell membrane has been characterized by quenching of an intracellular fluorophore, 6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ). In 20 mM Hepes, pH 7.4, and varying concentrations of chloride and bicarbonate with total anion concentration of 150 mM, SPQ is quenched dynamically by chloride with an apparent Stern-Volmer quenching constant 0.071 +/- 0.016 mM-1 at 25 degrees C. Hepes alone quenched SPQ fluorescence with a quenching constant of 0.030 +/- 0.003 mM-1. Stopped-flow kinetic experiments give fluorescence time courses that, when corrected for Stern-Volmer quenching, are first order. Disulfonic stilbenes (inhibitors of anion exchange) decrease the rate of fluorescence equilibration. Transport of bicarbonate via hydration-dehydration of CO2 does not contribute to the observed kinetics. The chloride-bicarbonate exchange rate is independent of the anion concentration gradient, but increases at 25 degrees C from 1 to 4 s-1 as equilibrium chloride concentration increases from 20 to 130 mM (with concomitant decrease in bicarbonate concentration from 130 to 20 mM). The data indicate that the translocation rate of the chloride-loaded transport protein is greater than th...Continue Reading

Citations

Dec 5, 1998·The American Journal of Physiology·A E Milton, I D Weiner
Mar 10, 2000·Biochemical and Biophysical Research Communications·L SoldatiG Vezzoli

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