title: Characterization of an electrogenic ATP and chloride-dependent proton translocating pump from rat renal medulla.
abstract: To study acidification mechanisms in the distal nephron, microsomes were prepared from rat renal medulla by differential centrifugation. Microsomes were enriched in the enzyme marker gamma-glutamyl transferase and contained an ATP-dependent proton pump, as evidenced by ATP-dependent, 3,3',4',5-tetrachlorosalicylanilide-reversible quenching of acridine orange fluorescence. Acidification was vanadate-insensitive, but was completely inhibited by micromolar N-ethylmaleimide. Maximal acidification was achieved in the presence of halide (Cl-, Br-) only and was not attainable with potassium-valinomycin diffusion potentials without halide ion. Microsomal ATPase activity was neither chloride- nor N-ethylmaleimide-sensitive. A chloride conductance was observed only with vesicles which had undergone ATP-dependent acidification. An ATP-dependent, N-ethylmaleimide-inhibitable, 3,3',4',5-tetrachlorosalicylanilide-reversible, and chloride-attenuated quench of bis(1,3-dibutylbarbituric acid-(5] pentamethinoxonol fluorescence was seen, consistent with net transfer of positive charge into the vesicles. Nonetheless, positive intravesicular potentials increased the ATP-dependent initial acidification rate, perhaps by increasing availability of chloride ion to the transport site. Our results are consistent with an electrogenic, ATP-dependent proton pump regulated by a voltage-sensitive chloride site.
mesh_terms: D000251:Adenosine Triphosphatases / Q000032:analysis; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D001692:Biological Transport; D002712:Chlorides / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007679:Kidney Medulla / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D009399:Nephrons / Q000378:metabolism; D010539:Permeability; D011522:Protons; D051381:Rats; D011919:Rats, Inbred Strains
chemical_list: D002712:Chlorides; D011522:Protons; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases