金属硫蛋白加质子常数及与Cd(II)络合常数测定

用pH计和Cd离子选择电极测定了金属硫蛋白的加质子常数及其与Cd(Ⅱ)的络合常数, 用改进的简化络合模型处理实验结果, 得到了去金属硫蛋白(apo MT)中6类不同的加质子基团的数目及其加质子常数。对Cd(Ⅱ)滴定数据的计算表明, MT中两个结构域——α和β对Cd(Ⅱ)的络合常数相差约1000倍。从热力学定量描述了MT中两个结构域结合金属离子选择优先顺序。

Determination of Apo-Metallothionein's Protonation Constants and Complexing Constants With Cadmium Ion

Metallothionein (MT) is a protein of molecular weight about 6000 Dalton obtained from liver of rat, rabbit and silver carp in our laboratory. Mammal's MT consists of 61 amino acid residues including 20 cysteine residues which has high affinity to metal and it appears to coordinate metals in two distinct configurations. Ions of at least eighteen different metals were associated with the protein. Most metals exhibited saturation binding at 7 mol eq forming M_7-MT. These included Bi(Ⅲ), Cd(Ⅱ), Co(Ⅱ), Hg(Ⅱ), In(III), Ni(II), Pb(II), Sb(III), and Zn(II). Other metals including Os(III), Pd(II), Pt(IV), Re(V), Rh(III) and Tl(III) give a positive indication of binding but their stoichiometries remained unclear. Ag(I) and Cu(I) bound in clusters as M_(12)-MT.In this paper, rabbit liver MT which combined with 5 Cd(II) and 2 Zn(II) ion per protein was obtained by gel filtration and ion exchange after homogenization, centrifugation and deposition with ethanol from rabbits received cadmium chloride by subcutaneously injection. In 0.01 mol·L~(-1) perchloric acid medium, the metal ions released from MT and apo-MT were obtained by gel filtration on a Sephadex G-25 column, the titration with sodium hydroxide and cadmium nitrate solution were carried out by combined pH electrode and cadmium ion selective electrode under nitrogen at 298 K.According to the sequences of amino acid from rabbit liver MT-2, the binding sites were divide into six types. By using a simplified complexing model, six types of protonation group and two types of metal complexing region (α- and β- domain) were obtained. The results were listed as follows....