人工合成表皮生长因子基因在甲基营养型酵母中的高效表达  

High Level Expression of Chemically Synthesized Human Epidermal Growth Factor Gene in the Yeast Pichia pastoris

作　　者：黄秉仁[1] 蔡良琬[1] 廖洪涛 唐申秀 周海涛[1] 

机构地区：[1]中国协和医科大学中国医学科学院基础医学研究所医学分子生物学国家重点实验室

出　　处：《中国生物化学与分子生物学报》1998年第5期512-517,共6页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家八五科技攻关课题

摘　　要：选用酵母菌偏爱密码子人工合成了编码５１个氨基酸的人表皮生长因子（ｈＥＧＦ）基因．将合成基因与编码酵母α因子前导肽８５个氨基酸的ＤＮＡ片段融合后克隆到醇氧化酶基因启动子下游，并构建出多拷贝表达载体．此载体转化甲基营养型酵母株ＧＳ１１５后筛选出整合型ＭｕｔＳＨｉｓ＋基因型菌株．高密度培养及诱导表达后该株可分泌具完好生物活性和正确物理化学性质的人表皮生长因子，产量达１００ｍｇ／Ｌ，经３次柱层析纯度达９５％以上。Human epidermal growth factor(hEGF) gene coding 51 amino acids was chemically synthesized by using favored code usage in yeast.The gene was under the control of the alcohol oxidation enzyme promoter and the α factor lead sequence including 85 amino acids coding sequence,and a multicopy insert was constructed as a part of the expression plasmid.The yeast Pichia pastoris was transformed by the expression plasmid and a Mut s His + cell line was screened.High cell density culture of the cell line was carried out and the cells were induced.The human epidermal growth factor with well biologically activity was secreted into the medium.The production was 100 mg/L medium.Its purity was 95% through chromatography three times.All of above mentioned lay the foundation for investigating the biological effects of EGF.

关 键 词：人表皮生长因子 甲基营养型酵母 高效表达 

分 类 号：Q78[生物学—分子生物学]