BINDING OF PURIFIED MULTIPLE ANTIBIOTIC-RESISTANCE REPRESSOR PROTEIN (MARR) TO MAR OPERATOR SEQUENCES
BINDING OF PURIFIED MULTIPLE ANTIBIOTIC-RESISTANCE REPRESSOR PROTEIN (MARR) TO MAR OPERATOR SEQUENCES
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DOI：
10.1073/pnas.92.12.5456
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发表时间：
1995-06-06
影响因子：
11.1
通讯作者：
ROSNER, JL
中科院分区：
文献类型：
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作者：
MARTIN, RG;ROSNER, JL
marORAB多重耐药操纵子的表达升高可增强大肠杆菌对各种医学上重要的抗生素的耐药性，该操纵子的转录在体内被marR编码的蛋白质抑制。马尔，并通过水杨酸和某些抗生素解阻遏，阻遏结果从马尔与marO操纵子-启动子区域的相互作用的可能性进行了研究，在体外使用纯化的马尔和含有marO的DNA片段。马尔R与marO DNA形成至少两种复合物，其与marO DNA的结合比与鲑鱼精子DNA的结合紧密30倍以上，并保护marO内两个独立的21 bp位点不被DNase I消化。位点I邻接推定的-35转录起始信号的下游侧，并包括4 bp的-10信号，位点II开始于位点I下游13 bp处，终止于马尔R的第一个碱基对之前。位点II与位点I同源性约为80%，不需要进行阻遏，因为位点II缺失的突变体（mar 0133）被野生型马尔R反式阻遏。位点II的缺失并不能阻止马尔R与mar 0133的位点I复合。水杨酸与马尔R结合（Kd约为0.5 mM），减弱了马尔R与位点I和II的相互作用，从而抑制了马尔操纵子的表达，抑制了大肠杆菌的耐药性。大肠杆菌，与MarR-位点I复合物的形成相关。水杨酸盐似乎通过与马尔R结合并抑制复合物形成来诱导马尔操纵子，而四环素和氯霉素既不结合马尔R也不抑制复合物形成，必须通过间接机制诱导。
Elevated expression of the marORAB multiple antibiotic-resistance operon enhances the resistance of Escherichia coli to various medically significant antibiotics, Transcription of the operon is repressed in vivo by the marR-encoded protein. MarR, and derepressed by salicylate and certain antibiotics, The possibility that repression results from MarR interacting with the marO operator-promoter region was studied in vitro using purified MarR and a DNA fragment containing marO. MarR formed at least two complexes with marO DNA bound >30-fold more tightly to it than to salmon sperm DNA, and protected two separate 21-bp sites within marO from digestion by DNase I. Site I abuts the downstream side of the putative -35 transcription-start signal and includes 4 bp of the -10 signal, Site II begins 13 bp downstream of site I, ending immediately before the first base pair of marR. Site Il, approximate to 80% homologous to site I, is not required for repression since a site II-deleted mutant (mar0133) was repressed in trans by wild-type MarR, The absence of site II did not prevent MarR from complexing with the site I of mar0133. Salicylate bound to MarR (K-d approximate to 0.5 mM) and weakened the interaction of MarR with sites I and II, Thus, repression of the mar operon, which curbs the antibiotic resistance of E. coli, correlates with the formation of MarR-site I complexes. Salicylate appears to induce the mar operon by binding to MarR and inhibiting complex formation, whereas tetracycline and chloramphenicol which neither bind MarR nor inhibit complex formation, must induce by an indirect mechanism.