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大肠杆菌的supX基因座是topA,即DNA拓扑异构酶I的结构基因。

The Escherichia coli supX locus is topA, the structural gene for DNA topoisomerase I.

作者信息

Margolin P, Zumstein L, Sternglanz R, Wang J C

出版信息

Proc Natl Acad Sci U S A. 1985 Aug;82(16):5437-41. doi: 10.1073/pnas.82.16.5437.

Abstract

Mutations in the supX locus, which result in the absence of DNA topoisomerase I enzyme activity in both Salmonella typhimurium and Escherichia coli, are all selected as suppressors of the leu-500 promoter mutation in S. typhimurium. To determine whether the supX locus is the structural gene topA for the DNA topoisomerase I enzyme or is a positive-acting regulator/activator gene for a nearby topA structural gene, nonsense mutations were selected in the E. coli supX gene carried on an F' episome in S. typhimurium cells. The cysB-topA region of the episomes with nonsense-mutant supX alleles were then cloned onto plasmid pBR322 and transformed into E. coli cells lacking a chromosomal supX gene. Three such E. coli strains, each carrying cloned DNA from episomes with different nonsense-mutant supX alleles, all lacked DNA topoisomerase I activity but expressed antigenic determinants specific to the enzyme; control cells lacked both enzyme activity and antigenic determinants. Maxicell studies of plasmid-coded proteins demonstrated the absence of the DNA topoisomerase I protein (100 kDa) in the three strains but the appearance of a new smaller peptide in each (36, 47, and 64 kDa). These new peptides must represent fragments of the enzyme resulting from translation termination at the supX nonsense codons and confirm the interpretation that the supX gene is topA, the structural gene for DNA topoisomerase I.

摘要

在鼠伤寒沙门氏菌和大肠杆菌中,supX基因座发生突变会导致DNA拓扑异构酶I酶活性缺失,这些突变均被选作鼠伤寒沙门氏菌中leu - 500启动子突变的抑制子。为了确定supX基因座是DNA拓扑异构酶I的结构基因topA,还是附近topA结构基因的正向作用调节/激活基因,在鼠伤寒沙门氏菌细胞中携带F'附加体的大肠杆菌supX基因中选择了无义突变。然后将带有无义突变supX等位基因的附加体的cysB - topA区域克隆到质粒pBR322上,并转化到缺乏染色体supX基因的大肠杆菌细胞中。三个这样的大肠杆菌菌株,每个都携带来自带有不同无义突变supX等位基因的附加体的克隆DNA,均缺乏DNA拓扑异构酶I活性,但表达该酶特有的抗原决定簇;对照细胞既缺乏酶活性也缺乏抗原决定簇。对质粒编码蛋白的最大细胞研究表明,这三个菌株中不存在DNA拓扑异构酶I蛋白(100 kDa),但每个菌株中都出现了一种新的较小肽段(36、47和64 kDa)。这些新的肽段必定代表了在supX无义密码子处翻译终止产生的酶片段,并证实了supX基因是topA这一解释,即DNA拓扑异构酶I的结构基因。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/dddd/390584/468851f5cccb/pnas00356-0211-a.jpg

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