牦牛MT-Ⅰ/-Ⅱ cDNA分子克隆及其蛋白质结构分析  

Molecular Cloning of cDNA Encoding for MT-I/-II and Analyzing of the Protein Structure in Yak

作　　者：马彬云[1] 任宏伟[2] 吴建平[1] 徐明旭[3] 张利平[1] 向云[4] 贺鹏飞[2] 蔡欣[5] 

机构地区：[1]甘肃农业大学动物科学技术学院,兰州730070 [2]北京大学蛋白质工程国家重点实验室,北京100871 [3]北京大学人类疾病基因研究中心,北京100083 [4]北京师范大学生命科学学院,北京100875 [5]西北农林科技大学生命科学学院,杨陵712100

出　　处：《中国生物工程杂志》2005年第4期62-68,共7页China Biotechnology

基　　金：教育部科学技术研究重点项目(03130)

摘　　要：利用基因特异引物YMTSP1和YMTSP2,通过RT-PCR从牦牛肝脏组织RNA中克隆出了牦牛 MT-Ⅰ(Genbank Accession No:AY513744)和MT-Ⅱ(Genbank Accession No:AY513745)基因编码区全 长。将牦牛MT-Ⅰ和MT-Ⅱ cDNA序列在CBI上进行同源性搜索发现,牦牛MT-Ⅰ/-Ⅱ编码区序列在 不同哺乳动物中相当保守。牦牛MT-Ⅰ和MT-Ⅱ编码的MT-Ⅰ和MT-Ⅱ蛋白分别由61个氨基酸组 成,其具有保守的短肽结构如:C-X-C,C-C-X-C-C,C-X-X-C等,其决定MT蛋白分子的整个三维结 构,在分子进化上十分保守。同时对牦牛MT的疏水性和跨膜区分析表明,牦牛MT蛋白可能不 存在跨膜区,也不存在信号肽,是1种非分泌蛋白。并通过同源比较模建,预测和构建了牦牛 MT-Ⅰ和MT-Ⅱ蛋白的分子空间结构,表明牦牛MT-Ⅰ和MT-Ⅱ由α-和β-两个结构域组成,在α-结构 域含有5个Cys短肽结构,β-结构域有4个Cys短肽结构,且2个结构域由保守的三肽序列KKS 相连。In the present study, the yak (B.Grunniens) MT-I(Genbank Accession No. AY513744) and MT-II (Genbank Accession No. AY513745) cDNA encoding sequence was amplified and cloned by RT-PCR using the gene specific primers YMTSPl and YMTSP2. The cDNA sequences of MT-I and MT-II were subjected to Blastn searching in CBI and the results showed that the nucleotide sequences of yak MT-I and MT-II between different species mammals are comparatively conservative. The yak cDNA sequences encoding for yak MT-I/-II protein composed of 61 amino acids, respectively. There are lots of conserved motifs, such as C-X-C, C-C-X-C-C, C-X-X-C, etc. and these motifs determine the overall three dimensional structure of the protein, which are comparatively conservative in their evolution. The hydrophobicity and transmembrane region analysis showed that the MTs are probably no transmembrane proteins and the signal peptides analysis suggested that they are non-secretory proteins. Then, the molecular spatial structures of yak MT-I/-II have been predicted by homology comparative modeling, which are composed of α- and β-domain. The α-domain (N-terminal domain) comprises residues 1 - 29 and contains five motifs, and the β-domains (C-terminal domain) begins at residue 33 and contains four cycteine motifs. There are little or no interactions between the two domains of a metallothionein molecule but they are linked by a short conserved tripetide KKS with residues 30-32.

关 键 词：cDNA分子克隆 蛋白质结构分析 牦牛 CDNA序列 基因编码区 氨基酸组成 结构域 特异引物 肝脏组织 哺乳动物 三维结构 蛋白分子 分子进化 分泌蛋白 同源比较 空间结构 跨膜区 CYS MT RNA PCR 同源性 CBI 短肽 分析表 疏水性 

分 类 号：Q51[生物学—生物化学] S968.251[农业科学—水产养殖]