Affinity identification of organic anion transporters in brush-border membrane vesicles from rat kidney.
The inhibitory properties of bromoacetyl-p-aminohippuric acid as the affinity probe of the organic anion transport system were studied. Bromoacetylated p-aminohippurate was shown to be able to inhibit irreversibly the p-aminohippurate (PAH) uptake in brush-border membrane vesicles. The inhibition depends on both the time of treatment and the affinity probe concentration. The treatment of brush-border membrane with 1 mM bromoacetyl-p-aminohippurate for 1.5 h results in 100% irreversible inhibition of PAH transport but no changes were observed in the activity of alkaline phosphatase, gamma-glutamyltranspeptidase or maltase. The affinity labelling of the organic anion transporters was performed with bromoacetyl-p-amino[3H]hippuric acid. It was shown, by means of SDS-polyacrylamide gel electrophoresis, that the probe bound covalently to the brush-border membrane proteins with molecular masses of 28 kDa, 63 kDa, 98 kDa, and > 150 kDa. The data obtained with SITS and probenecid as the organic anion transport inhibitors indicate that brush-border membrane proteins of 28 kDa, 63 kDa, 98 kDa may correspond to the organic anion transport system.
Orlov YuN
St. Petersburg Institute of Nuclear Physics of the Academy of Sciences, Gatchina, Russia.
Zherebtsova
M A
MA
Kazbekov
E N
EN
D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D000345:Affinity Labels; D000618:Aminohippuric Acids; D000818:Animals; D007672:Kidney Cortex / Q000378:metabolism* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000032:analysis*; D008871:Microvilli / Q000201:enzymology / Q000378:metabolism*; D051381:Rats; D010130:p-Aminohippuric Acid / Q000378:metabolism*
D000345:Affinity Labels; D000618:Aminohippuric Acids; D008565:Membrane Proteins; C087730:bromoacetyl-4-aminohippuric acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D010130:p-Aminohippuric Acid