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首页> 外文期刊>BMC Biotechnology >Molecular cloning and functional characterization of an ATP-binding cassette transporter OtrC from Streptomyces rimosus
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Molecular cloning and functional characterization of an ATP-binding cassette transporter OtrC from Streptomyces rimosus

机译：核糖链霉菌ATP结合盒转运蛋白OtrC的分子克隆和功能表征

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Background The otrC gene of Streptomyces rimosus was previously annotated as an oxytetracycline (OTC) resistance protein. However, the amino acid sequence analysis of OtrC shows that it is a putative ATP-binding cassette (ABC) transporter with multidrug resistance function. To our knowledge, none of the ABC transporters in S. rimosus have yet been characterized. In this study, we aimed to characterize the multidrug exporter function of OtrC and evaluate its relevancy to OTC production. Results In order to investigate OtrC’s function, otrC is cloned and expressed in E. coli The exporter function of OtrC was identified by ATPase activity determination and ethidium bromide efflux assays. Also, the susceptibilities of OtrC-overexpressing cells to several structurally unrelated drugs were compared with those of OtrC-non-expressing cells by minimal inhibitory concentration (MIC) assays, indicating that OtrC functions as a drug exporter with a broad range of drug specificities. The OTC production was enhanced by 1.6-fold in M4018 ( P =?0.000877) and 1.4-fold in SR16 ( P =?0.00973) duplication mutants, while it decreased to 80% in disruption mutants ( P =?0.0182 and 0.0124 in M4018 and SR16, respectively). Conclusions The results suggest that OtrC is an ABC transporter with multidrug resistance function, and plays an important role in self-protection by drug efflux mechanisms. This is the first report of such a protein in S. rimosus , and otrC could be a valuable target for genetic manipulation to improve the production of industrial antibiotics.
机译：背景技术先前已将线状链霉菌的otrC基因注释为土霉素（OTC）抗性蛋白。但是，OtrC的氨基酸序列分析表明它是一种具有多重耐药功能的推定ATP结合盒（ABC）转运蛋白。据我们所知，还没有特征性地描述过金黄色葡萄球菌的ABC转运蛋白。在这项研究中，我们旨在表征OtrC的多药出口商功能，并评估其与OTC生产的相关性。结果为了研究OtrC的功能，将otrC克隆并在大肠杆菌中表达。通过ATPase活性测定和溴化乙锭外排试验确定了OtrC的输出功能。同样，通过最小抑制浓度（MIC）分析比较了OtrC过表达细胞对几种与结构无关的药物的敏感性与OtrC非表达细胞的敏感性，表明OtrC作为具有广泛药物特异性的药物输出者起作用。在M4018中，OTC产量增加了1.6倍（P =？0.000877），而在SR16中OTC产量增加了1.4倍（P =？0.00973），而在突变型突变体中，OTC产量下降了80％（P =？0.0182和0.0124）。和SR16）。结论结果表明，OtrC是一种具有多药耐药功能的ABC转运蛋白，在药物外排机制的自我保护中起着重要作用。这是首次在核糖链球菌中发现这种蛋白，otrC可能成为遗传操作以改善工业抗生素生产的重要靶标。

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