Suppressed expression of hypoxanthine-guanine phosphoribosyltransferase (HGPRT) in an irradiation-attenuated Plasmodium berghei XAT strain.
Plasmodium berghei XAT (XAT) is a non-reversible, non-lethal type malaria parasite strain derived from the highly virulent lethal P. berghei NK65 (NK65) by X-irradiation. The difference in polypeptide expression between NK65 and XAT was examined in this study. Western blot patterns of the parasite polypeptides showed that a 30-kDa polypeptide was not detected in XAT. In the present paper, we focused the study on the difference in the expression of the 30-kDa polypeptide between XAT and NK65. Although several other significant differences were noted in the spots shown by two-dimensional gel electrophoresis, the 30-kDa polypeptide was isolated by means of preparative 2D-gel electrophoresis followed by HPLC, and N-terminal amino acid sequence of the polypeptide was eventually determined. Complementary DNA clones encoding the 30-kDa polypeptide were isolated and characterized. Full-length cDNA clones from XAT encoded a protein of 231 amino acid residues with a 693-bp open reading frame. The deduced amino acid sequence exhibited 67% identity with that for P. falciparum hypoxanthine-guanine phosphoribosyltransferase (HGPRT; EC 2.4.2.8), suggesting that this protein is P. berghei HGPRT. Northern blot analysis revealed that expression of HGPRT in XAT was only one-eighth of that in NK65. This finding indicates that HGPRT gene expression is markedly suppressed in XAT. The amino acid sequence of HGPRT from NK65 was identical to that from XAT. This finding showed that the amino acid sequence of XAT-HGPRT was not mutated and had not undergone deletion.
Onda
T
T
Department of Parasitology, Gunma University School of Medicine, Maebashi, Japan.
Miyamoto
K
K
Sugioka
Y
Y
Kangawa
K
K
Kano
S
S
Suzuki
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D018076:DNA, Complementary / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D004912:Erythrocytes / Q000469:parasitology; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D008288:Malaria / Q000469:parasitology*; D051379:Mice; D008969:Molecular Sequence Data; D010962:Plasmodium berghei / Q000201:enzymology* / Q000472:pathogenicity / Q000528:radiation effects*; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014774:Virulence
D018076:DNA, Complementary; D007041:Hypoxanthine Phosphoribosyltransferase