重组谷氨酰胺转胺酶的表达、复性及纯化 
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湖北省自然科学基金项目(2010CDZ046)


Expression,renaturation and purification of recombinant transglutaminase
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    摘要:

    以吸水链霉菌基因组DNA为模板,通过PCR扩增得到谷氨酰胺转胺酶基因片段,构建pGEX-TGase重组质粒,并转化到大肠杆菌中得到重组菌BL21/pGEX-TGase。重组菌经乳糖诱导产生融合蛋白,并对诱导条件进行了优化。表达产物主要以包涵体形式存在,采取稀释及尿素梯度液相色谱结合的方法,成功地实现了包涵体蛋白的复性。通过GST亲和柱纯化目的蛋白,得到酶活为29 U/mg的电泳纯目的蛋白。

    Abstract:

    Using the genomic DNA of Streptomyces hygroscopicus as a template,transglutaminase gene segments were obtained by PCR amplification.The recombinant plasmid pGEX-TGase was transformed into E.coli to get the recombinant strain BL21/pGEX-TGase.The recombinant strain produced fusion protein induced by lactose and the conditions of induction were optimized.Expression products existed mainly in the form of inclusion bodies.Dilution and urea gradient liquid chromatography was used to refold the inclusion bodies successfully.The target proteins with electrophoretical purity and activity of 29 U/mg were purified by GST affinity column.

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邵坤彦,彭宝玉,叶程,刘威,何冬兰.重组谷氨酰胺转胺酶的表达、复性及纯化 [J].华中农业大学学报,2013,32(1):14-18

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