The renal proximal tubule actively secretes many organic anions, including anionic metabolites and drugs. It is generally accepted that the basolateral membrane constitutes the active carrier mediated step in transepithelial anion secretion. The apical membrane step is also thought to be carrier mediated. However, the physico-chemical nature of the carrier is obscure. We have demontrated that the photoaffinity probe, N-(4-azido-2-nitrophenyl)-2-aminoethyl-sulfonate (NAP-taurine), a compound secreted by the kidney in the same manner as PAH, inhibits PAH transport by purified basolateral and apical membrane vesicles isolated from the rabbit kidney. This suggests NAP-taurine is capable of interacting with carrier proteins in both membranes. Utilizing this probe we were able to covalently label four membrane proteins in basolateral membranes. Labeling was competitively inhibited by PAH. There was a high correlation between labeling and inhibition of transport. The labeled proteins were separated by differential detergent solubilization. We propose to reconstitute these detergent solubilized proteins in proteoliposomes as the first step in purificatin of the system. In addition, we propose to utilize the same methodology (successfully applied to basolateral membranes) to identify putative transport elements in rabbit brush border membranes and Necturus basolateral and brush border membranes. The latter system is interesting since NAP-taurine competitively inhibits PAH transport but is not itself transported. This is probably due to the presence of a sulfonate moiety. Sulfonate containing anions are not transported in this species. These studies should yield information about the molecular characteristic of these membrane carriers which is essential to the understanding of normal kidney function as well as for prediction of drug interaction with the kidney.
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