Identification and characterization of a novel Neospora caninum immune mapped protein 1
Identification and characterization of a novel Neospora caninum immune mapped protein 1
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新型犬新孢子虫免疫图谱蛋白 1 的鉴定和表征
DOI：
10.1017/s0031182012000285
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发表时间：
2012-07-01
期刊：
影响因子：
2.4
通讯作者：
Liu, Q.
中科院分区：
文献类型：
--
作者：
Cui, X.;Lei, T.;Liu, Q.
免疫映射蛋白1（IMP1）是在巨型艾美耳球虫中发现的一种新蛋白。它被认为是一种潜在的抗E. maxima和一个高度保守的顶复门寄生虫蛋白。虽然犬新孢子虫IMP 1（NcIMP 1）是E.最大IMP1在N.犬基因组，它仍然没有被确定和表征。在这项研究中，编码NcIMP 1的cDNA序列通过RT-PCR从Nc1速殖子中分离的RNA克隆。NcIMP 1基因编码一个1182 bp的开放阅读框，编码393个氨基酸，预测分子量为42·9 kDa。序列分析表明，NcIMP 1的氨基酸序列中既没有信号肽，也没有跨膜区。然而，几种功能蛋白质基序，包括N-肉豆蔻酰化位点和棕榈酰化位点的预测。在大肠杆菌中表达重组NcIMP 1（rNcIMP 1），纯化后制备小鼠抗血清。针对rNcIMP 1产生的小鼠多克隆抗体识别约43 kDa的天然IMP 1蛋白。免疫荧光分析显示NcIMP 1定位于N.犬速殖子发现N-肉豆蔻酰化位点和棕榈酰化位点有助于NcIMP 1的定位。rNcIMP 1特异性抗体可抑制N.犬速殖子体外培养。这些结果表明NcIMP 1可能是N.犬，并可能参与寄生虫入侵。
SUMMARY Immune mapped protein 1 (IMP1) is a newly discovered protein in Eimeria maxima. It is recognized as a potential vaccine candidate against E. maxima and a highly conserved protein in apicomplexan parasites. Although the Neospora caninum IMP1 (NcIMP1) orthologue of E. maxima IMP1 was predicted in the N. caninum genome, it was still not identified and characterized. In this study, cDNA sequence encoding NcIMP1 was cloned by RT-PCR from RNA isolated from Nc1 tachyzoites. NcIMP1 was encoded by an open reading frame of 1182 bp, which encoded a protein of 393 amino acids with a predicted molecular weight of 42·9 kDa. Sequence analysis showed that there was neither a signal peptide nor a transmembrane region present in the NcIMP1 amino acid sequence. However, several kinds of functional protein motifs, including an N-myristoylation site and a palmitoylation site were predicted. Recombinant NcIMP1 (rNcIMP1) was expressed in Escherichia coli and then purified rNcIMP1 was used to prepare specific antisera in mice. Mouse polyclonal antibodies raised against the rNcIMP1 recognized an approximate 43 kDa native IMP1 protein. Immunofluorescence analysis showed that NcIMP1 was localized on the membrane of N. caninum tachyzoites. The N-myristoylation site and the palmitoylation site were found to contribute to the localization of NcIMP1. Furthermore, the rNcIMP1-specific antibodies could inhibit cell invasion by N. caninum tachyzoites in vitro. All the results indicate that NcIMP1 is likely to be a membrane protein of N. caninum and may be involved in parasite invasion.