乙型肝炎病毒S基因密码子最佳化及在毕赤酵母中的高效表达  

Codons optimization and high-level expression of hepatitis B virus S gene in Pichia pastoris

作　　者：周才[1] 邝炎波[1] 曾丽[1] 赖有行[1] 

机构地区：[1]广东省妇幼保健院检验科,广州510010

出　　处：《中华生物医学工程杂志》2007年第6期387-392,共6页Chinese Journal of Biomedical Engineering

摘　　要：目的制备适合酵母表达的乙型肝炎病毒S基因，在毕赤酵母中高效表达重组乙型肝炎病毒S蛋白。方法选用酵母偏爱的同义密码子替换野生型S基因的酵母稀有密码子，采取基因搭桥法及递归式聚合酶链反应，制备合成基因，捕入酵母表达载体pPICZB。携带有合成S基因的重组质粒转化毕赤酵母菌株KM71H，经甲醇诱导表达乙型肝炎病毒S蛋白。结果酶切电泳及DNA测序证实合成的S基因正确克隆到表达载体中；聚丙烯酰胺凝胶电泳及免疫印迹显示优化后的乙型肝炎病毒S基因在毕赤酵母中的重组蛋白表达量远高于野生型基因。结论密码子优化的S基因能明显提高重组S蛋白存毕赤酵母表达系统中的表达量。Objective To synthesize hepatitis B virus S gene which is suitable for yeast protein expression and express hepatitis B virus S protein in Pichia pastoris. Methods Synonymous eodons preferred by yeast usage on protein expression were used to replace some native codons of wild-type HBV S gene. Synthetic S gene （synS-gene） was achieved by a recursive PCR （rPCR）. The synS-gene was inserted into the yeast expression vector pPICZB. Recombinant plasmid was transformed into KM71H yeast by electroporatinn. The yeast transformant induced by methanol expressed the S protein. Results The restriction analysis and DNA sequencing confirmed that the synS-gene was inserted into yeast pPICZB in correct orientation. SDS-PAGE and ELISA indicated that the S protein was expressed by the yeast transformant. The expression level of S protein in the strain containing synS-gene was more higher than the strain containing wild-type S-gene. Conclusion Codons optimization can promote the yield of recombinant S protein in Pichia pastoris.

关 键 词：肝炎病毒 乙型 S蛋白 偏爱密码子 毕赤酵母 

分 类 号：R3[医药卫生—基础医学]