THE COPPER, ZINC-SUPEROXIDE DISMUTASE GENE OF SACCHAROMYCES-CEREVISIAE - CLONING, SEQUENCING, AND BIOLOGICAL-ACTIVITY
THE COPPER, ZINC-SUPEROXIDE DISMUTASE GENE OF SACCHAROMYCES-CEREVISIAE - CLONING, SEQUENCING, AND BIOLOGICAL-ACTIVITY
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DOI：
10.1073/pnas.85.13.4789
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发表时间：
1988-07-01
影响因子：
11.1
通讯作者：
VALENTINE, JS
中科院分区：
文献类型：
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作者：
BERMINGHAMMCDONOGH, O;GRALLA, EB;VALENTINE, JS
从酿酒酵母（Saccharomycescerevisiae）中克隆了铜、锌-超氧化物歧化酶（Cu，Zn-SOD：EC 1.14.1.1）的基因，并对其进行了测序，结果表明该基因具有生理活性。通过使用长的独特的dexoyoligonucleotide探针从cDNA gt 11文库中分离该基因。通过计算机生成的酵母密码子偏好表，从已知的氨基酸序列推导出探针序列。报告了编码区和侧翼区的序列。克隆的基因在体内表达并显示出活性。将含有编码区和上游160个碱基的3.2-SOD酶片段亚克隆到酵母/大肠杆菌穿梭载体中，转化缺乏Cu，Zn-SOD活性的酵母菌株。含Cu，Zn-SOD基因质粒的存在纠正了该菌株的特征性氧敏感性。用酵母Cu，Zn-SOD抗体进行电泳转移印迹，结果表明在转化体和野生型酵母中存在该蛋白，但在突变体中不存在。根据这些发现，Cu，Zn-SOD在防御双氧毒性中的作用进行了讨论。
The gene for copper, zinc-superoxide dismutase (Cu,Zn-SOD: EC 1.14.1.1) from the yeast Saccharomyces cerevisiae has been cloned, sequenced, and shown to have physiological activity. The gene was isolated from a .lambda.gt11 library by using a long, unique dexoyoligonucleotide probe. The probe sequence was deduced from the known amino acid sequence by using a computer-generated yeast codon preference table. The sequence of the coding and flanking regions is reported. The cloned gene was expressed and shown to be active in vivo. A 3.2-kilobase fragment containing the coding region and 160 upstream bases, subcloned in a yeast/Escherichia coli shuttle vector, was used to transform a yeast strain lacking Cu,Zn-SOD activity. The presence of the Cu,Zn-SOD gene-containing plasmid corrected the characteristic dioxygen sensitivty of this strain. Electrophoretic transfer blots with antibody to yeast Cu,Zn-SOD showed the presence of the protein in transformants and wild-type yeast but not in the mutant. The role of Cu,Zn-SOD in defense against dioxygen toxicity is discussed in the light of these findings.