人生长激素释放因子基因的高效表达

The High Expression of hGRF Gene

将人生长激素释放因子 ( h GRF)基因和编码 L-天门冬酰胺酶 ( L- ans B) C-末端 12 6个氨基酸残基的基因片断进行融合表达 ,融合蛋白在菌体总蛋白中的比率达到 30 %左右 ,比和 L- ans B全基因融合表达时高得多 ,表达条件优化后 ,比率可增加到 4 5%。融合蛋白以不溶性的包含体形式存在。包含体的尿素抽提液经乙醇分级沉淀后得到较纯的融合蛋白 ,60 mmol/ L的盐酸水解融合蛋白 ,通过 SDS- PAGE和电喷雾质谱分析 ,融合蛋白释放出一分子量为 52 35.59Da的物质 ,而 GRF的分子量是 52 35.4 8Da,也就是说融合蛋白酸水解释放出了

The hGRF was fused with C terminal 126 amino acids fragment of \%L\% ansB and the expression level of the fusion protein increased to 30 % comparing with its fusion expression to the whole \%L\% ansB molecule. After improving the expressive condition such as media and inductive condition, the expression level can reach to 45 %. The fusion protein was insoluble in \%E.coli\% cytoplasm. After dissolving the inclusion body in 8 mmol/L urea and precipitating by ethanol, the partially purified fusion protein was obtained. 60 mmol/L hydrogen chloride hydrolyzed the fusion protein. Through the analysis of SDS PAGE and the electrospray mass spectrum, a material with MW 5235.59 Da was released from the fusion protein. It is GRF whose MW is 5235.48 Da.