CLONING AND CHARACTERIZATION OF THE PSEUDOMONAS-AERUGINOSA LASR GENE, A TRANSCRIPTIONAL ACTIVATOR OF ELASTASE EXPRESSION
CLONING AND CHARACTERIZATION OF THE PSEUDOMONAS-AERUGINOSA LASR GENE, A TRANSCRIPTIONAL ACTIVATOR OF ELASTASE EXPRESSION
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DOI：
10.1128/jb.173.9.3000-3009.1991
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发表时间：
1991-05-01
影响因子：
3.2
通讯作者：
IGLEWSKI, BH
中科院分区：
文献类型：
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作者：
GAMBELLO, MJ;IGLEWSKI, BH
我们报告了 lasR 基因的发现，该基因可正向调节铜绿假单胞菌 PAO1 中的弹性蛋白酶表达。 lasR 基因因其在菌株 PA103 中恢复阳性弹性蛋白酶表型的能力而被克隆，该菌株拥有弹性蛋白酶结构基因 (lasB)，但无法合成该酶。核苷酸序列分析揭示了一个由 716 个核苷酸组成的开放阅读框，编码大约 27 kDa 的蛋白质。使用 T7 RNA 聚合酶表达系统在大肠杆菌中检测到 27 kDa 的标记 LasR 蛋白。通过基因置换在PAO1中构建了lasR基因的染色体缺失突变体。该突变体 (PAO-R1) 缺乏弹性蛋白酶活性和弹性蛋白酶抗原。推导的LasR氨基酸序列与费氏弧菌阳性激活子LuxR和大肠杆菌疑似激活子28K-UvrC有27%同源性。对多拷贝质粒 (pMG1.7) 上含有 lasR 基因的 PAO1、PAO-R1 和 PAO-R1 的总细胞 RNA 进行 Northern (RNA) 分析，结果表明功能性 lasR 基因是弹性蛋白酶结构基因 (lasB) 转录所必需的。
We report the discovery of the lasR gene, which positively regulates elastase expression in Pseudomonas aeruginosa PAO1. The lasR gene was cloned by its ability to restore a positive elastase phenotype in strain PA103, a strain which possesses the elastase structural gene (lasB) but fails to synthesize the enzyme. Nucleotide sequence analysis revealed an open reading frame of 716 nucleotides encoding a protein of approximately 27 kDa. A labeled LasR protein of 27 kDa was detected in Escherichia coli by using a T7 RNA polymerase expression system. A chromosomal deletion mutant of the lasR gene was constructed in PAO1 by gene replacement. This mutant (PAO-R1) is devoid of elastolytic activity and elastase antigen. The deduced amino acid sequence of LasR is 27% homologous to the positive activator LuxR of Vibrio fischeri and the suspected activator 28K-UvrC of E. coli. Northern (RNA) analysis of total cellular RNA from PAO1, PAO-R1, and PAO-R1 containing the lasR gene on a multicopy plasmid (pMG1.7) revealed that a functional lasR gene is required for transcription of the elastase structural gene (lasB).