Inactivation of the renal microvillus membrane Na+-H+ exchanger by histidine-specific reagents.
We examined the effect of histidine-specific reagents on the transport activity of the Na+-H+ exchanger in microvillus (brush-border) membrane vesicles isolated from the rabbit renal cortex. Rose bengal-catalyzed photo-oxidation caused irreversible inhibition of the rate of Na+-H+ exchange but also caused significant loss of vesicle integrity. Treatment of the membrane vesicles with diethylpyrocarbonate caused inactivation of Na+-H+ exchange that could not be attributed to vesicle disruption or collapse of transmembrane H+ gradients. Inactivation of Na+-H+ exchange by diethylpyrocarbonate followed pseudo-first order kinetics to below 10% residual activity, could be reversed by hydroxylamine, was reflected by a decreased Vmax with no change in the Km for Na+, was dependent on external pH but not internal pH, was blocked by amiloride, and was enhanced by Na+. These data are consistent with the hypothesis that a diethylpyrocarbonate-sensitive imidazolium residue is the titratable group found in kinetic studies to bind H+ at the external transport site of the Na+-H+ exchanger.
D000818:Animals; D002352:Carrier Proteins / Q000037:antagonists & inhibitors*; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D004285:Dogs; D006639:Histidine / Q000378:metabolism*; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000494:pharmacology; D007672:Kidney Cortex / Q000187:drug effects / Q000648:ultrastructure*; D007700:Kinetics; D008871:Microvilli / Q000187:drug effects / Q000378:metabolism; D010777:Photochemistry; D011817:Rabbits; D012395:Rose Bengal / Q000494:pharmacology; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers; D013997:Time Factors
D002352:Carrier Proteins; D006898:Hydroxylamines; D017923:Sodium-Hydrogen Exchangers; D012395:Rose Bengal; D019811:Hydroxylamine; D006639:Histidine; D012964:Sodium; D004047:Diethyl Pyrocarbonate