Cloning, sequencing and expression of a novel cDNA encoding human vacuolar ATPase (14-kDa subunit).
A cDNA encoding the 14-kDa subunit of vacuolar ATPase was cloned from human fetal brain. The sequence was composed of 680 nucleotides containing an open reading frame of 357 nucleotides. The deduced peptide sequence consisted of 119 amino acid residues with a calculated molecular weight of 13,369 Da and a pI of 5.19. Overall, this amino-acid sequence was respectively 69% and 70% identical to those of Manduca sexta and Drosophila melanogaster 14-kDa subunits, although the two representatives of Class Insecta were remarkably similar to one another (91% identity). Three regions in particular (the N-terminal, amino acids 5-36; the middle, residues 58-68; and the C-terminal, residues 88-118) were highly conserved. Hence, we think that the 14-kDa subunits have evolved from the same ancestral gene, and that the three conserved regions are important for the structure and function of vacuolar ATPase. A single 0.8-kb band was detected in various human tissues by Northern blot analysis. Since the human 14-kDa subunit is expressed ubiquitously, it might be a housekeeping protein. A separate transcript found in the cDNA library lacked a 6-bp segment in the 5' non-coding region (nucleotides -40 to -35) and also carried a 23C to T (8Thr to Ile) point mutation in the coding region; these minor differences likely reflected normal polymorphism.
Fujiwara
T
T
Otsuka GEN Research Institute, Otsuka Pharmaceutical Co., Ltd., Tokushima, Japan.
Kawai
A
A
Shimizu
F
F
Hirano
H
H
Okuno
S
S
Takeda
S
S
Ozaki
K
K
Shimada
Y
Y
Nagata
M
M
Watanabe
T
T
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D001921:Brain / Q000201:enzymology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005333:Fetus / Q000201:enzymology; D006801:Humans; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D006180:Proton-Translocating ATPases / Q000096:biosynthesis / Q000235:genetics*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014617:Vacuoles / Q000201:enzymology*
D018076:DNA, Complementary; D006180:Proton-Translocating ATPases