摘要
参考E .coli偏爱密码子 ,设计并合成人MT 3α结构域 (α MT 3)的DNA片段 ,克隆到表达载体pGEX 4T 1中。经IPTG诱导 ,在E .coli中高效表达出谷胱甘肽 S 转移酶 (GST) α融合蛋白。凝血酶酶切融合蛋白 (用CdCl2 或ZnSO4辅助 )后 ,得到纯化的结合镉或锌的α MT 3蛋白。质谱分子量检测及氨基酸组成分析证明得到了目的蛋白。紫外吸收光谱及圆二色性光谱研究重组α MT 3的结构。MT 3的与其余MT的α结构域不同 ,α MT 3的圆二色性光谱在 2 2 0nm左右为负峰 ,说明α MT 3可能包含一个α 螺旋的二级结构。
According to the E.coli preference codon,six DNA fragments of α domain of human metallothionein\|3(α\|MT\|3) were synthesized and cloned into the vector pGEX\|4T\|1.Upon IPTG induction,Glutathione\|S\|transferase (GST)\|α fusion protein was efficiently expressed in E.coli .After digestion by thrombin (combined with CdCl\-2 or ZnSO\-4),the desired protein binding Cd\+\{2+\} or Zn\+\{2+\} was released,which was confirmed by molecular mass and amino acid composition analysis.Ultraviolet (UV) and Circular Dichroism (CD) spectra were used to test the secondary structure of α\|MT\|3.Distinct from other α\|MT,α\|MT\|3 showed a negative trough at about 220?nm in the CD spectrum,suggesting that an α\|helix might exist in its secondary structure.
出处
《北京大学学报（自然科学版）》
CAS
CSCD
北大核心
2001年第4期470-476,共7页
Acta Scientiarum Naturalium Universitatis Pekinensis
基金
国家九五攻关资助项目 ( 96 C0 2 0 1 0 9)
