Characterization of a sodium-dependent concentrative nucleobase-transport system in guinea-pig kidney cortex brush-border membrane vesicles.
The characteristics of hypoxanthine transport were examined in purified brush-border membrane vesicles isolated from guinea-pig kidney. Hypoxanthine uptake in the vesicles was specifically stimulated by both Na+ and an inside-negative potential, resulting in a transient accumulation of intravesicular hypoxanthine. Na(+)-dependent hypoxanthine influx was saturable (apparent Km 4.4 +/- 2.1 microM, Vmax. 128 +/- 29 pmol/min per mg of protein at 100 mM NaCl and 22 degrees C). Guanine, thymine, 5-fluorouracil and uracil inhibited hypoxanthine uptake (Ki values 1-30 microM), but adenine and the nucleosides inosine and thymidine were without effect. Guanine competitively inhibited Na(+)-dependent hypoxanthine influx, suggesting that it was a substrate for the active nucleobase transporter in guinea-pig renal membrane vesicles. A sigmoidal dependence between hypoxanthine influx and Na+ concentration was obtained (KNa 13 +/- 2 mM; Hill coefficient, h, 2.13 +/- 0.14), suggesting that at least two Na+ ions are transported per hypoxanthine molecule. This system differs from the Na(+)-nucleobase carrier in cultured LLC-PK1 renal cells, which has a stoichiometric coupling ratio of 1:1. These results represent the first demonstration of an active electrogenic nucleobase carrier in renal apical membrane vesicles.
Research School of Biosciences, University of Kent, Canterbury, U.K.
D A
DA
Griffith
S M
SM
Jarvis
D000818:Animals; D001692:Biological Transport; D006168:Guinea Pigs; D019271:Hypoxanthine; D007042:Hypoxanthines / Q000378:metabolism; D007672:Kidney Cortex / Q000378:metabolism* / Q000502:physiology / Q000648:ultrastructure; D007700:Kinetics; D008297:Male; D008564:Membrane Potentials; D008871:Microvilli / Q000378:metabolism / Q000502:physiology; D012964:Sodium / Q000378:metabolism*
D007042:Hypoxanthines; D019271:Hypoxanthine; D012964:Sodium