Characterization and immunolocalization of an NTP diphosphohydrolase of Trypanosoma cruzi

被引:52
作者
Fietto, JLR
DeMarco, R
Nascimento, IP
Castro, IM
Carvalho, TMU
de Souza, W
Bahia, MT
Alves, MJM
Verjovski-Almeida, S
机构
[1] Univ Sao Paulo, Inst Quim, Dept Bioquim, BR-05599970 Sao Paulo, SP, Brazil
[2] Univ Fed Ouro Preto, Nucl Pesquisas Ciencias Biol, BR-35400000 Ouro Preto, MG, Brazil
[3] Univ Fed Rio de Janeiro, IBCCF, BR-21246900 Rio De Janeiro, RJ, Brazil
基金
巴西圣保罗研究基金会;
关键词
Trypanosoma cruzi; ecto-NTPDase; apyrase; immuno-localization; plasma-membrane; purine salvage pathway;
D O I
10.1016/j.bbrc.2004.02.071
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
An ecto-NTP diphosphohydrolase (NTPDase) activity, insensitive to inhibitors of ATPases and phosphatases, was characterized on the surface of live Trypanosoma cruzi intact parasites. The enzyme exhibits broad substrate specificity, typical of NTPDases, and a high hydrolysis rate for GTP. A 2282 bp message encoding a full-length NTPDase was cloned by RT-PCR using epimastigote mRNA. A single protein was immunoprecipitated from [S-35]methionine-labeled parasites using antibodies against Toxoplasma gondii NTPase I. This antibody localized an NTPDase on the external surface of all forms of T cruzi, as seen by confocal immunofluorescence microscopy. The NTPDase could be part of the parasite's purine salvage pathway. Additionally, trypomastigotes (infective form) presented a 2:1 ATP/ADP hydrolysis ratio, while epimastigotes (non-infective form) presented a 1:1 ratio, suggesting a possible role for the NTPDase in the parasite's virulence mechanisms. (C) 2004 Elsevier Inc. All rights reserved.
引用
收藏
页码:454 / 460
页数:7
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