牦牛金属硫蛋白-Ⅲ(MT-Ⅲ)cDNA分子克隆及序列分析

利用基因特异引物YMT-ⅢSP1和YMT-ⅢSP2,通过RT-PCR从牦牛大脑组织中克隆出了牦牛MT-Ⅲ基因编码区全长．将牦牛MT-ⅢcDNA序列在CBI上进行同源性搜索发现牦牛MT-Ⅲ编码区序列与羊、猪、人、鼠的同源性分别为95％、92％、89％、86％,该序列在不同哺乳动物中相当保守,并通过牦牛MTs(Ⅰ、Ⅱ、Ⅲ)编码区序列之间的分析表明,MT-Ⅰ与MT-Ⅱ编码的蛋白质性质、功能之间差异较小,而MT-Ⅲ与MT-Ⅰ／Ⅱ编码的蛋白质性质、功能之间差异相对较大．牦牛MT-Ⅲ基因编码的MT-Ⅲ蛋白由68个氨基酸组成,其中19个半胱氨酸,具有MTs保守的短肽结构如:C-X-C、C-C-X-C-C、C-X-X-C、KKS以及MT-Ⅲ特异的MDPE、CPCP等结构,在分子进化上很保守．分析表明了牦牛MT-Ⅲ蛋白质是一个低分子量,富含半胱氨酸．不含芳香族氨基酸,也无二硫键的蛋白质．

Molecular cloning and sequence analyzing of cDNA encoding yak metallothionein-Ⅲ

Metallothionein-Ⅲ (MT-Ⅲ),as a new member of the metallothionein(MT) family, has specific physiological effect different from known MT-Ⅰ and MT-Ⅱ,and the difference in functional aspect might origin from its own unique structure. In this study,the yak MT-Ⅲgene coding region was amplified and cloned by RT-PCR from brain tissue of yak using YMT-ⅢSP1 and YMT-ⅢSP2 as specific primers of gene. The cDNA sequence of MT-Ⅲ was subjected to Blastn searching in CBI and the results showed that the MT-Ⅲ coding region sequences of sheep,pig,human,mouse shared 95 %,92 %,89 % and 86 % homology with yak,respectively,and it was comparative conservative in different mammals species. Analysis between three metallothionein coding sequences in Yak suggested that property and function of MT-Ⅰ and MT-Ⅱ had less differences, while those of MT-Ⅲ and MT-Ⅰ/Ⅱ had more differences. MT-Ⅲ protein was composed of 68 amino acid,including 19 cysteine. There were lots of conserved motifs in MTs,such as C-X-C,C-C-X-C-C,C-X-X-C,KKS,and specific structures,MDPE,CPCP, etc. in MT-Ⅲ ,this indicated it's comparatively conservative in MT-Ⅲ molecular evolution. The analysis showed that MT-Ⅲ was a low-molecular-weight,abundant-cysteine,no-aromatic-amino-acid and no-disul-fide-bond protein.