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编码与人类Ro/SS-A自身抗原同源的曼氏血吸虫抗原的基因的克隆

Cloning of the gene encoding a Schistosoma mansoni antigen homologous to human Ro/SS-A autoantigen.

作者信息

Khalife J, Trottein F, Schacht A M, Godin C, Pierce R J, Capron A

机构信息

Centre d'Immunologie et de Biologie Parasitaire, Unité Mixte INSERM U167-CNRS URA 624, Institut Pasteur, Lille, France.

出版信息

Mol Biochem Parasitol. 1993 Feb;57(2):193-202. doi: 10.1016/0166-6851(93)90195-4.

Abstract

A cDNA library was constructed from the mRNA of adult worms of Schistomsoma mansoni in the expression vector lambda gt11 and screened with a rabbit antiserum raised against a 60-65-kDa electroeluted adult worm fraction. Two overlapping clones were selected and a partial nucleotide sequence was deduced (1172 bp). The full-length sequence was obtained by the amplification of the 5' end of first strand cDNA using PCR. The overall mRNA size was 1335 nt including a 25 nt 5' non-coding region and a 131 nt untranslated region with the poly(A) tail. The predicted amino acid sequence of 393 aa (45 kDa) has 52% identity with the human Ro/SS-A autoantigen, which is considered to be the human calreticulin. As for the human Ro/SS-A, the protein encoded by the cDNA described here contains a hydrophobic leader sequence and a carboxyl terminal sequence, HDEL consensus signal sequence for retention in the ER. An antiserum raised against the fusion protein of one clone recognized a 58-kDa antigen in homogenates of cercariae and of adult worms. The expression of the protein in the pGEX-2T fusion system allowed us to show the presence of specific antibodies in S. mansoni infected patients' sera and in the sera of patients with systemic lupus erythematosus, reflecting a cross-immunoreactivity between the S. mansoni protein and the human calreticulin autoantigen.

摘要

从曼氏血吸虫成虫的mRNA构建了一个cDNA文库,该文库克隆于表达载体λgt11中,并用针对60 - 65 kDa电洗脱成虫组分产生的兔抗血清进行筛选。挑选出两个重叠克隆,并推导了部分核苷酸序列(1172 bp)。通过使用PCR扩增第一链cDNA的5'端获得了全长序列。总的mRNA大小为1335 nt,包括一个25 nt的5'非编码区和一个带有poly(A)尾的131 nt非翻译区。预测的393个氨基酸(45 kDa)的氨基酸序列与人类Ro/SS - A自身抗原具有52%的同一性,该自身抗原被认为是人类钙网蛋白。与人类Ro/SS - A一样,此处描述的cDNA编码的蛋白质包含一个疏水前导序列和一个羧基末端序列,即用于保留在内质网中的HDEL共有信号序列。针对一个克隆的融合蛋白产生的抗血清识别尾蚴和成虫匀浆中的一种58 kDa抗原。该蛋白在pGEX - 2T融合系统中的表达使我们能够在曼氏血吸虫感染患者的血清和系统性红斑狼疮患者的血清中检测到特异性抗体,这反映了曼氏血吸虫蛋白与人类钙网蛋白自身抗原之间的交叉免疫反应性。

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