日本血吸虫腺苷脱氨酶的克隆与融合表达
[Cloning and fusion expression of adenosine deaminase of Schistosoma japonicum].
作者信息
Yang Zhong, Xu Bin, Wang Wei, Feng Zheng, Wei Dong-Zhi, Hu Wei
机构信息
State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China Uiniversity of Science and Technology, Shanghai 200237, China.
出版信息
Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi. 2007 Feb 28;25(1):6-11.
OBJECTIVE
To clone and express a new protein adenosine deaminase of Schistosoma japonicum (SjADA).
METHOD
Specific primers were designed according to the EST sequence and used for amplification of the encoding sequence from the cDNA clone containing SjADA. The gene was sub-cloned into pET32 plasmid and expressed in E.coli BL31 (DE) pLys strain and the recombinant proteins were purified by chelating sepharose FF. The structure and phylifunctions of this protein were analyzed by MrBayes and GeneAtlas.
RESULT
The full length sequence of SjADA gene was obtained and the recombinant protein was cloned, expressed and purified successfully. The bioinformatics analysis showed that the identity of SjADA and SmADA gene sequence was only 25% and they belonged to different subfamily. The structure of SjADA had 41% identity with it's PDB template 1A4M.
CONCLUSION
The recombinant protein of SjADA has been obtained. The purine salvage pathway of S. japonicum may be different with that of S. mansoni.
目的
克隆并表达日本血吸虫一种新的蛋白——腺苷脱氨酶（SjADA）。
方法
根据EST序列设计特异性引物，用于从含SjADA的cDNA克隆中扩增编码序列。将该基因亚克隆到pET32质粒中，在大肠杆菌BL31(DE)pLys菌株中表达，重组蛋白通过螯合琼脂糖FF纯化。利用MrBayes和GeneAtlas分析该蛋白的结构和系统功能。
结果
获得了SjADA基因的全长序列，成功克隆、表达并纯化了重组蛋白。生物信息学分析表明，SjADA与曼氏血吸虫ADA基因序列的同一性仅为25%，它们属于不同的亚家族。SjADA的结构与其PDB模板1A4M有41%的同一性。
结论
已获得SjADA重组蛋白。日本血吸虫的嘌呤补救途径可能与曼氏血吸虫不同。
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