Lipid requirements for reconstitution of the proton-translocating complex of clathrin-coated vesicles.

X. S. Xie, S. J. Tsai, D. K. Stone

Research output: Contribution to journalArticle

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Abstract

We have recently reported the reconstitution of the clathrin-coated vesicle proton-translocating complex with liposomes prepared from ethanol-extracted crude bovine brain lipids. Reconstitution of proton pumping by the isolated proton ATPase has now been achieved with liposomes prepared from pure lipids. Optimal proton pumping, as assessed by ATP-generated acridine orange quenching and 32Pi-ATP exchange, is achieved with liposomes prepared from phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and cholesterol at a weight ratio of 40:26.5:7.5:26, and a lipid-to-protein weight ratio of 200:1. Under such conditions, the extent of decrease of ATP-generated acridine orange quenching is 150-fold greater than that of native clathrin-coated vesicles. Cholesterol is required to stabilize the proteoliposome. Phosphatidylserine, which is most effective in activating ATP-hydrolytic activity of the solubilized enzyme, is not obligatory for reconstitution of proton pumping.

Original languageEnglish (US)
Pages (from-to)8913-8917
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume83
Issue number23
StatePublished - Dec 1986

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Clathrin-Coated Vesicles
Protons
Lipids
Adenosine Triphosphate
Liposomes
Acridine Orange
Phosphatidylserines
Cholesterol
Weights and Measures
Phosphatidylcholines
Adenosine Triphosphatases
Ethanol
Brain
Enzymes
Proteins

ASJC Scopus subject areas

  • General
  • Genetics

Cite this

Lipid requirements for reconstitution of the proton-translocating complex of clathrin-coated vesicles. / Xie, X. S.; Tsai, S. J.; Stone, D. K.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 83, No. 23, 12.1986, p. 8913-8917.

Research output: Contribution to journalArticle

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