Abstract: 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 CdCl₂ or ZnSO₄), the desired protein binding Cd²⁺ or Zn²⁺ 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 220nm in the CD spectrum, suggesting that an α-helix might exist in its secondary structure.

Key words: metallothionein-3, α-MT-3, expression, structure

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220nm左右为负峰，说明α-MT-3可能包含一个α-螺旋的二级结构。

关键词: α-MT-3, 表达, 结构, 金属硫蛋白-3