Transcriptomic and biochemical analyses identify a family of chlorhexidine efflux proteins
Transcriptomic and biochemical analyses identify a family of chlorhexidine efflux proteins
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DOI：
10.1073/pnas.1317052110
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发表时间：
2013-12-10
影响因子：
11.1
通讯作者：
Paulsen, Ian. T.
中科院分区：
文献类型：
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作者：
Hassan, Karl A.;Jackson, Scott M.;Paulsen, Ian. T.
洗必泰被广泛用作医院和社区环境中的防腐剂或消毒剂。许多细菌物种显示出对这种膜活性杀生物剂的抗性。我们研究了一种代表性的医院内人类病原体鲍曼不动杆菌对洗必泰的转录组反应，以确定主要的洗必泰耐药元件。最高度上调的基因编码的主要多药外排系统，AdeAB的组件。在洗必泰胁迫下的下一个最高度过表达的基因被注释为编码一种假设的蛋白质，在此命名为AceI。aceI基因的直系同源物在广泛的变形菌物种的基因组内是保守的。在大肠杆菌中表达的aceI或其直向同源物从其他几个γ-或β-变形菌的物种导致显着增加氯己定的耐药性。此外，破坏贝氏不动杆菌中的aceI直系同源物使其对洗必泰更敏感。AceI蛋白在E.杆菌这种蛋白质被纯化，结合试验证明直接和特定的相互作用AceI和洗必泰。使用[C-14]-氯己定的转运试验确定AceI能够介导氯己定的能量依赖性外排。在一个保守的酸性残基中具有突变的E15 Q AceI突变体，虽然不能介导氯己定抗性和转运，但仍然能够结合氯己定。总之，这些数据与AceI是一种活性洗必泰外排蛋白和细菌药物外排转运蛋白家族的创始成员一致。
Chlorhexidine is widely used as an antiseptic or disinfectant in both hospital and community settings. A number of bacterial species display resistance to this membrane-active biocide. We examined the transcriptomic response of a representative nosocomial human pathogen, Acinetobacter baumannii, to chlorhexidine to identify the primary chlorhexidine resistance elements. The most highly up-regulated genes encoded components of a major multidrug efflux system, AdeAB. The next most highly overexpressed gene under chlorhexidine stress was annotated as encoding a hypothetical protein, named here as AceI. Orthologs of the aceI gene are conserved within the genomes of a broad range of proteobacterial species. Expression of aceI or its orthologs from several other gamma- or beta-proteobacterial species in Escherichia coli resulted in significant increases in resistance to chlorhexidine. Additionally, disruption of the aceI ortholog in Acinetobacter baylyi rendered it more susceptible to chlorhexidine. The AceI protein was localized to the membrane after overexpression in E. coli. This protein was purified, and binding assays demonstrated direct and specific interactions between AceI and chlorhexidine. Transport assays using [C-14]-chlorhexidine determined that AceI was able to mediate the energy-dependent efflux of chlorhexidine. An E15Q AceI mutant with a mutation in a conserved acidic residue, although unable to mediate chlorhexidine resistance and transport, was still able to bind chlorhexidine. Taken together, these data are consistent with AceI being an active chlorhexidine efflux protein and the founding member of a family of bacterial drug efflux transporters.