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. 1988 Dec;85(24):9590–9594. doi: 10.1073/pnas.85.24.9590

Purification and reconstitution of the proton-translocating ATPase of Golgi-enriched membranes.

G P Young 1, J Z Qiao 1, Q Al-Awqati 1
PMCID: PMC282806  PMID: 2904677

Abstract

Kidney cortex microsomes enriched in Golgi markers and probably also containing endosomes were isolated by cell fractionation and found to contain a proton-translocating ATPase that was inhibited by N-ethylmaleimide (NEM). This NEM-sensitive ATPase was solubilized with n-octyl glucoside and purified using anion-exchange sievorptive chromatography on sequential DEAE-Sephadex and QAE-Sephadex columns followed by a final hydroxyapatite HPLC column. The purified enzyme, with a specific activity of 4.4 mumol.mg-1.min-1 was completely inhibited by NEM. Addition of asolectin and removal of the detergent by dialysis resulted in reconstitution of NEM-sensitive electrogenic proton transport. This vacuolar ATPase is composed of five polypeptides with apparent molecular masses of 68, 58, 40, 37, and 16 kDa.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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