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，可引起多种感染，由于这种细菌固有的和获得性的抗生素耐药性，很难治疗。这种抗生素耐药性可归因于低渗透性外膜 (OM) 和细胞主动流出之间的协同作用。铜绿假单胞菌的基因组编码了属于五个不同家族的 35 个药物外排系统，但我们的工作假设是，导致内在和/或获得性耐药性的临床相关外排泵属于耐药结节分裂 (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解答的问题。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的其他地方招募其他 OM 通道，以作为药物在整个细胞包膜上流出的三方流出系统，正如目前的那样教条说，或者它们是否可以作为其他基材的双组分系统。其次，除了单个泵外，其他 RND 泵都受到严格调控，但控制其表达和诱导底物的调控网络仍然未知。因此，我们的其他工作假设是：（i）不编码自己的 OM 通道的外排泵可能会招募迄今为止未识别的蛋白质来与某些底物一起发挥作用，但也可能充当其他底物的双组分外排系统； (ii) 外排泵表达受特定的、也可能是全局转录调节因子的控制。我们将使用最近发现的 MexJK 外排泵来测试这些假设，该泵通常是安静的，并且不会对其自己的 OM 通道进行编码。我们还建议修改这些研究，以探讨是否有任何全局转录调节因子可能参与 RND 外排泵表达。具体来说，我们建议： 目标 1-利用生化和分子研究确定 MexJK 的同源调节因子 MexL 对 MexJK 的调节；识别某些生长培养基中可能存在的诱导剂；鉴定出存在于铜绿假单胞菌中但在大肠杆菌中不存在的推定激活因子。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 膜通道要求（如果有）。目标 3：探索外排操纵子表达的其他（也许是全局）调节因子，特别是 MexS 和 PA4878。

项目成果

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

B 细胞对环境挑战的反应推动了 TNFRSF13B 多样化 - 一个假设。

• 发表时间: 2021
• 作者: Cascalho, Marilia;Platt, Jeffrey L
• 通讯作者: Platt, Jeffrey L

Molecular basis of azithromycin-resistant Pseudomonas aeruginosa biofilms.

耐阿奇霉素铜绿假单胞菌生物膜的分子基础。

• 发表时间: 2005-09
• 影响因子: 4.9
• 作者: Gillis, Richard J;White, Kimberly G;Choi, Kyoung;Wagner, Victoria E;Schweizer, Herbert P;Iglewski, Barbara H
• 通讯作者: Iglewski, Barbara H

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

通过单 B 细胞筛选发现和表征高亲和力、强效 SARS-CoV-2 中和抗体。

• 发表时间: 2021-10-20
• 影响因子: 4.6
• 作者: Schardt, John S;Pornnoppadol, Ghasidit;Desai, Alec A;Park, Kyung Soo;Zupancic, Jennifer M;Makowski, Emily K;Smith, Matthew D;Chen, Hongwei;Garcia de Mattos Barbosa, Mayara;Cascalho, Marilia;Lanigan, Thomas M;Moon, James J;Tessier, Peter M
• 通讯作者: Tessier, Peter M

Molecular characterization of MexL, the transcriptional repressor of the mexJK multidrug efflux operon in Pseudomonas aeruginosa.

MexL 的分子特征，MexL 是铜绿假单胞菌中 mexJK 多药外排操纵子的转录阻遏蛋白。

• 发表时间: 2005-05
• 作者: Chuanchuen, Rungtip;Gaynor, Jared B;Karkhoff;Schweizer, Herbert P
• 通讯作者: Schweizer, Herbert P

Substrate-dependent utilization of OprM or OpmH by the Pseudomonas aeruginosa MexJK efflux pump.

铜绿假单胞菌 MexJK 外排泵对 OprM 或 OpmH 的底物依赖性利用。

• 发表时间: 2005-05
• 作者: Chuanchuen, Rungtip;Murata, Takeshi;Gotoh, Naomasa;Schweizer, Herbert P
• 通讯作者: Schweizer, Herbert P