Plasma membrane proton ATPase from human kidney

Abstract

Distal urinary acidification is thought to be mediated by a proton ATPase (H+‐ATPase). We isolated a plasma membrane fraction from human kidney cortex and medulla which contained H+‐ATPase activity. In both the cortex and medulla the plasma membrane fraction was enriched in alkaline phosphatase, maltase, Na+,K+‐ATPase and devoid of mitochondrial and lysosomal contamination. In the presence of oligomycin (to inhibit mitochondrial ATPase) in the presence of ouabain (to inhibit Na+,K+‐ATPase) and in the absence of Ca (to inhibit Ca2+‐ATPase) this plasma membrane fraction showed ATPase activity which was sensitive to dicyclohexylcarbodiimide and N‐ethylmaleimide. This ATPase activity was also inhibited by vanadate, 4,4′‐diisothiocyano‐2,2′‐disulfonic stilbene and ZnSO4. In the presence of ATP, but not GTP or UTP, the plasma membrane fraction of both cortex and medulla was capable of quenching of acridine orange fluorescence, which could be dissipated by nigericin indicating acidification of the interior of the vesicles. The acidification was not affected by presence of oligomycin or ouabain indicating that it was not due to mitochondrial ATPase or Na+,K+‐ATPase, respectively. Dicyclohexylcarbodiimide and N‐ethylmaleimide completely abolished the acidification by this plasma membrane fraction. In the presence of valinomycin and an outward‐directed K gradient, there was increased quenching of acridine orange, indicating that the H+‐ATPase is electrogenic. Acidification was not altered by replacement of Na by K, but was critically dependent on the presence of chloride. In summary, the plasma membrane fraction of the human kidney cortex and medulla contains a H+‐ATPase, which is similar to the H+‐ATPase described in other species, and we postulate that this H+‐ATPase may be involved in urinary acidification.

Chlordecone is a potent in vitro inhibitor of oligomycin‐insensitive mg2+ ‐ATPase of rat bile canaliculi–enriched fraction

“…Since DCCD was also a potent inhibitor of mitochondrial ATPase (21), NEM was considered more specific. The higher concentration of NEM required for OIMATPase inhibition was consistent with reports for H+-ATPases in other tissues (21)(22)(23) Colchicine, ethynylestradiol, manganese, phalloidin, and taurocholate failed to inhibit BCEF OIMATPase at concentrations up to 132 pM (Figure 1). Concentration-dependent inhibition of BCEF OIMATPase by ANIT, CD, filipin, and taurolithocholate in a low micromolar range demonstrated their direct actions on the canalicular plasma membrane (Figure 1).…”

“…At 1 mM, NEM inhibited 17.4 2 4.5% (n = 4 individual preparations in duplicate) of BCEF OIMATPase. Both DCCD and NEM inhibited H+-ATPase of turtle bladder epithelium (21) and renal distal tubule of humans (22) and rats (23) associated with urinary acidification. Since DCCD was also a potent inhibitor of mitochondrial ATPase (21), NEM was considered more specific.…”

Vacuolar-type ATPase in the accessory boring organ of Nucella lamellosa (Gmelin) (Mollusca : Gastropoda): role in shell penetration

“…The disparity in the ATPase activities among the three ABO homogenates can be attributed primarily to variation in the amount of V-ATPase actually present in each homogenate (total protein was determined, so inclusion of excess foot tissue, for example, would lower the V-ATPase content) and to variation in the amount of Pi released through dissociation of endogenous ATP or other phosphorylated substrates during tissue processing and assay procedures. The activity of the ABO V-ATPase is comparable to that reported in other animals: Onken and Putzenlechner ( 1995) reported V-ATPase activities of 20 and 10 pmol Pi/ljug protein min) in filtrates from the posterior and anterior gills of Eriocheir sinensis, respectively, whereas Al-Fifi et al (1998) found a higher activity of 192.3 pmol PiAjug protein min) in a 600 X g supernatant obtained from the homogenates of Malpighian tubules in Locusta, and Sallman et al (1986) reported V-ATPase activities of 30.8 pmol PiAjug protein min) and 42.4 pmol Pi/(;ug protein min) in homogenates respectively obtained from the cortex and medulla of human kidneys.…”