Regulation and function of Pseudomonas drug efflux pumps

假单胞菌药物外排泵的调节和功能

• 批准号: 7057771
• 负责人: HERBERT P. SCHWEIZER
• 金额: $ 24.65万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2007-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7057771
• 关键词: DNA binding protein; Pseudomonas aeruginosa; SDS polyacrylamide gel electrophoresis; active transport; antibiotics; bacteria infection mechanism; bacterial genetics; enteric bacteria; gel mobility shift assay; gene expression; genetic mapping; genetic regulation; liquid chromatography mass spectrometry; membrane transport proteins; multidrug resistance; operon; polymerase chain reaction; protein structure function; site directed mutagenesis

DESCRIPTION (provided by applicant): Pseudomonas aeruginosa is an opportunistic pathogen that causes a multitude of infections, which are difficult to treat because of this bacterium's intrinsic and acquired antibiotic resistance. This antibiotic resistance can be attributed to synergy between a low-permeability outer membrane (OM) and active efflux from the cell. The genome of P. aeruginosa encodes 35 proposed drug efflux systems belonging to five different families, but our working hypothesis is that the clinically relevant efflux pumps contributing to intrinsic and/or acquired drug resistance belong to the resistance nodulation division (RND) family which contains 12 members. Genome sequence and expression analysis of these systems poses several questions that remain unanswered. First, because only a subset of the RND operons contains genes for OM channel proteins, the question is whether these systems must recruit other OM channels from elsewhere on the genome to function as tripartite efflux systems for drug efflux across the entire cell envelope, as current dogma says, or whether they can function as two-component systems for other substrates. Second, with the exception of a single pump, the other RND pumps are tightly regulated but the regulatory networks governing their expression and the inducing substrates remain unknown. Our other working hypotheses therefore are: (i) efflux pumps not encoding their own OM channels may either recruit hitherto unidentified proteins for function with certain substrates but may also function as two-component efflux systems for other substrates; (ii) Efflux pump expression is governed by specific and probably also global transcriptional regulators. We will test these hypotheses using the recently discovered MexJK efflux pump that is normally silent and does not encode its own OM channel. We also propose to amend these studies to probe whether any global transcriptional regulators may be involved in RND efflux pump expression. Specifically we propose to: Aim 1-Establish the regulation of MexJK by its cognate regulator MexL using biochemical and molecular studies; identify an inducer that may be present in certain growth media; identify a putative activating factor present in P. aeruginosa but absent in E. coil. Aim 2: Establish the molecular architecture of MexJK, specifically its OM membrane channel requirement, if any, using genetic and biochemical methods. Aim 3: Probe other, perhaps global regulators of efflux operon expression, specifically MexS and PA4878.

描述（由申请人提供）：铜绿假单胞菌是一种机会性病原体，会引起多种感染，由于这种细菌的固有和获得的抗生素耐药性，难以治疗。这种抗生素耐药性可以归因于低渗透性外膜（OM）和来自细胞的主动排出之间的协同作用。铜绿假单胞菌的基因组编码35个属于五个不同家族的药物外排系统，但我们的工作假设是，临床相关的外排泵有助于固有和/或获得耐药性的耐药性属于耐药性结节司（RND）家族，其中包含12名成员。这些系统的基因组序列和表达分析提出了几个尚未得到答复的问题。首先，因为只有RND操纵子的一个子集包含OM通道蛋白的基因，所以问题是这些系统是否必须从基因组上其他地方募集其他OM通道，以充当整个细胞包膜中药物外排的三方外排系统，正如当前的教条所说的那样，或者它们是否可以充当其他底物的两组成分系统。其次，除了单个泵外，另一个RND泵受到严格调节，但是控制其表达的调节网络和诱导的底物仍然未知。因此，我们的其他工作假设是：（i）未编码自己的OM通道的外排泵可以募集迄今未识别的蛋白质用于某些底物的功能，但也可能充当其他底物的两组分解系统； （ii）外排泵的表达受特定的，可能也可能是全球转录调节剂的控制。我们将使用最近发现的MEXJK外排泵测试这些假设，该泵通常是无声的，不会编码自己的OM通道。我们还建议修改这些研究，以探究是否可能参与RND外排泵表达。具体而言，我们建议：通过生化和分子研究，通过其同源调节剂MexL对Mexjk进行调节；确定某些增长培养基中可能存在的诱导剂；确定铜绿假单胞菌中存在的假定激活因子，但在E.卷轴中不存在。目标2：使用遗传和生化方法建立MexJK的分子结构，特别是其OM膜通道需求（如果有）。 AIM 3：探测外排操作子表达的其他，可能是全球调节剂，尤其是MEX和PA4878。

项目成果

Discovery and characterization of high-affinity, potent SARS-CoV-2 neutralizing antibodies via single B cell screening.

• DOI: 10.1038/s41598-021-99401-x
• 发表时间: 2021-10-20
• 期刊: Scientific reports
• 影响因子: 4.6
• 作者: Schardt JS;Pornnoppadol G;Desai AA;Park KS;Zupancic JM;Makowski EK;Smith MD;Chen H;Garcia de Mattos Barbosa M;Cascalho M;Lanigan TM;Moon JJ;Tessier PM
• 通讯作者: Tessier PM

Evidence of MexT-independent overexpression of MexEF-OprN multidrug efflux pump of Pseudomonas aeruginosa in presence of metabolic stress.

• DOI: 10.1371/journal.pone.0026520
• 发表时间: 2011
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Kumar A;Schweizer HP
• 通讯作者: Schweizer HP

TNFRSF13B Diversification Fueled by B Cell Responses to Environmental Challenges-A Hypothesis.

• DOI: 10.3389/fimmu.2021.634544
• 发表时间: 2021
• 期刊: Frontiers in immunology
• 影响因子: 7.3
• 作者: Cascalho M;Platt JL
• 通讯作者: Platt JL