TY - JOUR
T1 - Proteolipid of vacuolar H+-ATPase of Plasmodium falciparum
T2 - cDNA cloning, gene organization and complementation of a yeast null mutant
AU - Yatsushiro, Shouki
AU - Taniguchi, Shinya
AU - Mitamura, Toshihide
AU - Omote, Hiroshi
AU - Moriyama, Yoshinori
N1 - Funding Information:
We are grateful to Drs. Nathan Nelson and Hannah Nelson (Tel Aviv University, Israel) for sending us the yeast cells and cDNA probes. This study was supported in part by a Grant-in-aid for Scientific Research on Priority Areas, Infectious Disease, 16-17264, to Y. M. from the Japanese Ministry of Education, Science, Culture and Sports.
PY - 2005/11/30
Y1 - 2005/11/30
N2 - Vacuolar H+-ATPase (V-ATPase), an electrogenic proton pump, is highly expressed in Plasmodium falciparum, the human malaria parasite. Although V-ATPase-driven proton transport is involved in various physiological processes in the parasite, the overall features of the V-ATPase of P. falciparum, including the gene organization and biogenesis, are far less known. Here, we report cDNA cloning of proteolipid subunit c of P. falciparum, the smallest and most highly hydrophobic subunit of V-ATPase. RT-PCR analysis as well as Northern blotting indicated expression of the proteolipid gene in the parasite cells. cDNA, which encodes a complete reading frame comprising 165 amino acids, was obtained, and its deduced amino acid sequence exhibits 52 and 57% similarity to the yeast and human counterparts, respectively. Southern blot analysis suggested the presence of a single copy of the proteolipid gene, with 5 exons and 4 introns. Upon transfection of the cDNA into a yeast null mutant, the cells became able to grow at neutral pH, accompanied by vesicular accumulation of quinacrine. In contrast, a mutated proteolipid with replacement of glutamate residue 138 with glutamine did not lead to recovery of the growth ability or vesicular accumulation of quinacrine. These results indicated that the cDNA actually encodes the proteolipid of P. falciparum and that the proteolipid is functional in yeast.
AB - Vacuolar H+-ATPase (V-ATPase), an electrogenic proton pump, is highly expressed in Plasmodium falciparum, the human malaria parasite. Although V-ATPase-driven proton transport is involved in various physiological processes in the parasite, the overall features of the V-ATPase of P. falciparum, including the gene organization and biogenesis, are far less known. Here, we report cDNA cloning of proteolipid subunit c of P. falciparum, the smallest and most highly hydrophobic subunit of V-ATPase. RT-PCR analysis as well as Northern blotting indicated expression of the proteolipid gene in the parasite cells. cDNA, which encodes a complete reading frame comprising 165 amino acids, was obtained, and its deduced amino acid sequence exhibits 52 and 57% similarity to the yeast and human counterparts, respectively. Southern blot analysis suggested the presence of a single copy of the proteolipid gene, with 5 exons and 4 introns. Upon transfection of the cDNA into a yeast null mutant, the cells became able to grow at neutral pH, accompanied by vesicular accumulation of quinacrine. In contrast, a mutated proteolipid with replacement of glutamate residue 138 with glutamine did not lead to recovery of the growth ability or vesicular accumulation of quinacrine. These results indicated that the cDNA actually encodes the proteolipid of P. falciparum and that the proteolipid is functional in yeast.
KW - Complementation
KW - Malaria
KW - Plasmodium falciparum
KW - Proteolipid
KW - Subunit c
KW - V-ATPase
KW - Yeast null mutant
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U2 - 10.1016/j.bbamem.2005.08.011
DO - 10.1016/j.bbamem.2005.08.011
M3 - Article
C2 - 16293223
AN - SCOPUS:27944507680
VL - 1717
SP - 89
EP - 96
JO - Biochimica et Biophysica Acta - Biomembranes
JF - Biochimica et Biophysica Acta - Biomembranes
SN - 0005-2736
IS - 2
ER -