Cloning and expression in Escherichia coli of a Fasciola hepatica gene encoding a calcium-binding protein
Cloning and expression in Escherichia coli of a Fasciola hepatica gene encoding a calcium-binding protein
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DOI：
10.1016/s0166-6851(99)00012-2
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发表时间：
1999-06-25
影响因子：
1.5
通讯作者：
Parra, F
中科院分区：
文献类型：
--
作者：
de Eguino, ADR;Machín, A;Parra, F
用兔抗肝片吸虫排泄分泌抗原的血清从成虫cDNA表达文库中克隆了一个994 bp的肝片吸虫cDNA克隆。cDNA克隆的核苷酸序列显示存在572 bp的开放阅读框，其编码显示推定的EF-手结构域的22 kDa多肽（Fh 22）。该基因在大肠杆菌中表达，重组蛋白用于生产特异性抗体。使用抗Fh 22血清的免疫印迹研究表明，在成虫寄生虫的排泄分泌提取物中存在类似分子量的多肽。重组Fh 22多肽显示钙依赖性电泳迁移率（随着Ca 2+离子降低，随着EGTA增加）。凝胶过滤实验中观察到的重组Fh 22的行为也表明钙诱导的构象变化。(C)1999年由Elsevier Science B. V.出版，版权所有。
A Fasciola hepatica cDNA clone of 994 bp was isolated from an adult worm cDNA expression library using a rabbit serum against the excretory-secretory antigens. The nucleotide sequence of the cDNA clone revealed the presence of an open reading frame of 572 bp which encoded a 22 kDa polypeptide (Fh22) showing putative EF-hand domains. This gene was expressed in Escherichia coli and the recombinant protein used for the production of specific antibodies. Immunoblotting studies using the anti-Fh22 serum showed the presence of a polypeptide of similar molecular mass in the excretory-secretory extract of the adult parasite. The recombinant Fh22 polypeptide showed calcium-dependent electrophoretic mobility (decreased with Ca2+-ions and increased with EGTA). The observed behaviour of recombinant Fh22 in gel filtration experiments also suggested calcium-induced conformational changes. (C) 1999 Published by Elsevier Science B.V. All rights reserved.