Identification of a 14-kDa subunit associated with the catalytic sector of clathrin-coated vesicle H+-ATPase

Sheng Bin Peng, Bill P. Crider, Sue Jean Tsai, Xiao Song Xie, Dennis K. Stone

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

The clathrin-coated vesicle H+-ATPase is composed of a peripheral catalytic sector (V(C)) and an integral membrane proton channel (V(B)), both of which are multiple subunit complexes. This study was conducted to determine if subunit F, previously identified in vacuolar proton pumps of tobacco hornworm and yeast, was present in mammalian pumps. Using a polymerase chain reaction-based strategy, we have isolated and sequenced cDNA clones from bovine and rat brain cDNA libraries. A full-length clone from rat brain encodes a 119-amino acid polypeptide with a predicted molecular mass of 13,370 Da and with approximately 72 and 49% identity to subunit F of tobacco hornworm and yeast, respectively. Southern and Northern blot analyses indicate that the protein is encoded by a single gene. An anti-peptide antibody, directed against deduced protein sequence, was affinity-purified and shown to react with a 14-kDa polypeptide that is present in a highly purified pump prepared from clathrin-coated vesicles and also isolated V(C). When stripped clathrin-coated vacuolars and purified chromaffin granule membranes were treated with KI in the presence of ATP, the 14-kDa subunit was released from both membranes, further indicating that it is part of the peripheral catalytic sector. In addition, direct sequencing of this 14-kDa component of the coated vacuolar proton pump confirmed its identity as a subunit F homologue.

Original languageEnglish (US)
Pages (from-to)3324-3327
Number of pages4
JournalJournal of Biological Chemistry
Volume271
Issue number6
DOIs
StatePublished - Feb 9 1996

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Clathrin-Coated Vesicles
Clathrin
Proton-Translocating ATPases
Manduca
Proton Pumps
Tobacco
Membranes
Yeast
Peptides
Rats
Brain
Clone Cells
Yeasts
Pumps
Chromaffin Granules
Polymerase chain reaction
Molecular mass
Southern Blotting
Gene Library
Ion Channels

ASJC Scopus subject areas

  • Biochemistry

Cite this

Identification of a 14-kDa subunit associated with the catalytic sector of clathrin-coated vesicle H+-ATPase. / Peng, Sheng Bin; Crider, Bill P.; Tsai, Sue Jean; Xie, Xiao Song; Stone, Dennis K.

In: Journal of Biological Chemistry, Vol. 271, No. 6, 09.02.1996, p. 3324-3327.

Research output: Contribution to journalArticle

Peng, Sheng Bin ; Crider, Bill P. ; Tsai, Sue Jean ; Xie, Xiao Song ; Stone, Dennis K. / Identification of a 14-kDa subunit associated with the catalytic sector of clathrin-coated vesicle H+-ATPase. In: Journal of Biological Chemistry. 1996 ; Vol. 271, No. 6. pp. 3324-3327.
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