cDNA sequence encoding the 16-kDa proteolipid of chromaffin granules implies gene duplication in the evolution of H+-ATPases.
Vacuolar H+-ATPases function in generating protonmotive force across the membranes of organelles connected with the vacuolar system of eukaryotic cells. This family of H+-ATPases is distinct from the two other families of H+-ATPases, the plasma membrane-type and the eubacterial-type. One of the subunits of the vacuolar H+-ATPase binds N,N'-dicyclohexylcarbodiimide (DCCD) and has been implicated in the proton-conducting activity of these enzymes. We have cloned and sequenced the gene encoding the DCCD-binding protein (proteolipid) of the H+-ATPase of bovine chromaffin granules. The gene encodes a highly hydrophobic protein of 15,849 Da. Hydropathy plots revealed four transmembrane segments, one of which contains a glutamic residue that is the likely candidate for the DCCD binding site. Sequence homology with the vacuolar proteolipid and with the proteolipids of eubacterial-type H+-ATPases was detected. The proteolipids from Escherichia coli, spinach chloroplasts, and yeast mitochondria matched better to the NH2-terminal part of the vacuolar protein. The proteolipids of bovine mitochondria and Neurospora mitochondria matched better to the COOH-terminal end of the vacuolar proteolipid. These findings suggest that the proteolipids of the vacuolar H+-ATPases were evolved in parallel with the eubacterial proteolipid, from a common ancestral gene that underwent gene duplication.
Mandel
M
M
Roche Institute of Molecular Biology, Roche Research Center, Nutley, NJ 07110.
Moriyama
Y
Y
Hulmes
J D
JD
Pan
Y C
YC
Nelson
H
H
Nelson
N
N
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005075:Biological Evolution; D002352:Carrier Proteins / Q000235:genetics*; D002736:Chloroplasts / Q000201:enzymology; D002837:Chromaffin Granules / Q000201:enzymology* / Q000648:ultrastructure; D002838:Chromaffin System / Q000201:enzymology*; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology; D007425:Intracellular Membranes / Q000201:enzymology; D007473:Ion Channels / Q000378:metabolism; D008928:Mitochondria / Q000201:enzymology; D008969:Molecular Sequence Data; D005810:Multigene Family; D011510:Proteolipids / Q000235:genetics*; D006180:Proton-Translocating ATPases / Q000235:genetics*; D011522:Protons; D012689:Sequence Homology, Nucleic Acid; D014617:Vacuoles / Q000201:enzymology
D002352:Carrier Proteins; D007473:Ion Channels; D011510:Proteolipids; D011522:Protons; D004247:DNA; D006180:Proton-Translocating ATPases