V-Type H+-ATPase/synthase from a thermophilic eubacterium, Thermus thermophilus. Subunit structure and operon.
V-type ATPase (V(o)V(1)) capable of ATP-driven H(+) pumping and of H(+) gradient driven ATP synthesis was isolated from a thermophilic eubacterium, Thermus thermophilus. When the enzyme was analyzed by gel electrophoresis in the presence of sodium dodecyl sulfate, it showed eight polypeptide bands of which four were subunits of V(1). We also isolated the V(o)V(1) operon, containing nine genes in the order of atpG-I-L-E-X-F-A-B-D, which encoded proteins with molecular sizes of 13, 43, 10, 20, 35, 11, 64, 53, and 25 kDa, respectively. The last four genes were identified as those for V(1) subunits; atpA, B, D, and F encoded the A, B, gamma, and delta subunits, respectively. The first five genes, atpG-atpX, were identified as genes for the V(o) subunits. The product of atpL, the proteolipid subunit, lacked a 19-amino acid presequence and, unlike V-type ATPases, contained two membrane-spanning domains rather than four. The hydrophobic 43-kDa product of atpI is the smallest member so far found of the eukaryotic 100-kDa subunit family. Its electrophoretic band overlapped with the band of the A subunit. Therefore, all the gene products were found in our purified V(o)V(1). We isolated the A(3)B(3) subcomplex reconstituted from the isolated subunits and the A(3)B(3)gamma subcomplex from subunit-expressing Escherichia coli. Electron microscopic observation of these subcomplexes revealed that the gamma subunit of V(1) filled the central cavity of A(3)B(3) and might be central subunit, similar to the gamma subunit of F(1)-ATPase.
Yokoyama
K
K
Laboratory of Biochemistry, Department of Molecular and Cellular Biology, Faculty of Pharmaceutical Sciences, Kanazawa University, Takara-machi 13-1, Kanazawa, Ishikawa 920-0934, Japan. yokoken
Ohkuma
S
S
Taguchi
H
H
Yasunaga
T
T
Wakabayashi
T
T
Yoshida
M
M
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009876:Operon / Q000235:genetics; D006180:Proton-Translocating ATPases / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D016963:Thermus thermophilus / Q000201:enzymology* / Q000235:genetics
D001426:Bacterial Proteins; D006180:Proton-Translocating ATPases