Insight into antibiotic resistance of pathogenic bacteria via structural studies of a multidrug transporter

通过多药转运蛋白的结构研究深入了解病原菌的抗生素耐药性

• 批准号: BB/K014226/1
• 负责人: Christopher Law
• 金额: $ 41.82万
• 依托单位: Queen's University Belfast
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2013
• 资助国家: 英国
• 起止时间: 2013 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FK014226%2F1
• 关键词: Insight; into; antibiotic; resistance; pathogenic

Antibiotic resistance is a major global health concern. One of the mechanisms that infectious bacteria have evolved to resist the effects of antibiotics is that of multidrug efflux. This process is mediated by molecular pumps, made of protein, that are embedded in the fatty membranes that surround the bacterial cell. An understanding of the structure and mechanism of these membrane proteins is vital if we are to understand how they recognise and transport their antibiotic substrates and, in turn, use that knowledge to our advantage in the fight against bacterial infections. Although membrane proteins are important biological molecules that represent about one third of all the proteins made by a typical cell, and nearly half of them are targets of drugs on the market today, the structures of relatively few membrane proteins have been solved. One of the reasons for this is the challenge of working with proteins that are embedded in the oil-like environment of biological membranes. This means they first need to be isolated from their membrane environment using detergents before they can be highly purified to free them of contaminants. The purified protein-detergent complex then needs to be crystallised and exposed to X-rays in order to obtain a detailed, three-dimensional atomic structure. We have isolated, purified and grown crystals of a membrane protein from E. coli that plays a role in the transport of a broad range of antibiotics out of the cell, thereby contributing to multidrug resistance. This is a vital first step towards solving the structure of this transporter protein using a technique called X-ray crystallography. The structural information gained will provide insight into how these proteins recognise and bind to a diverse range of antibiotic substrate molecules and how they use energy to drive those substrates out of the cell. This information, in combination with biochemical studies designed to test if the protein performs other physiologically relevant functions, could be of use in the fight against antibiotic resistance in harmful bacteria.

抗生素耐药性是一个主要的全球健康问题。感染性细菌进化出的抵抗抗生素作用的机制之一是多药外排。这个过程是由蛋白质组成的分子泵介导的，分子泵嵌在细菌细胞周围的脂肪膜中。了解这些膜蛋白的结构和机制至关重要，如果我们要了解它们如何识别和运输抗生素底物，并反过来利用这些知识来对抗细菌感染。尽管膜蛋白是重要的生物分子，占典型细胞所产生的所有蛋白质的约三分之一，并且其中近一半是当今市场上药物的靶点，但相对较少的膜蛋白的结构已经被解决。其中一个原因是与嵌入在生物膜的油状环境中的蛋白质一起工作的挑战。这意味着它们首先需要使用洗涤剂从膜环境中分离出来，然后才能高度纯化以去除污染物。然后，纯化的蛋白质-去污剂复合物需要结晶并暴露于X射线，以获得详细的三维原子结构。我们从大肠杆菌中分离、纯化并生长了一种膜蛋白晶体。大肠杆菌中的一种，它在将多种抗生素运输出细胞中发挥作用，从而导致多药耐药性。这是使用称为X射线晶体学的技术解决这种转运蛋白结构的重要第一步。获得的结构信息将提供这些蛋白质如何识别和结合各种抗生素底物分子以及它们如何使用能量将这些底物赶出细胞的见解。这些信息，结合旨在测试蛋白质是否具有其他生理相关功能的生物化学研究，可以用于对抗有害细菌的抗生素耐药性。

项目成果

Prokaryotes: Physiology, Biochemistry and Cell Behavior

原核生物：生理学、生物化学和细胞行为

• 发表时间: 2014
• 作者: Alegre, K.O

Purification of a Multidrug Resistance Transporter for Crystallization Studies.

• DOI: 10.3390/antibiotics4010113
• 发表时间: 2015-03-05
• 期刊: Antibiotics (Basel, Switzerland)
• 作者: Alegre KO;Law CJ
• 通讯作者: Law CJ

Multidrug resistance protein MdtM adds to the repertoire of antiporters involved in alkaline pH homeostasis in Escherichia coli.

• DOI: 10.1186/1471-2180-13-113
• 发表时间: 2013-05-23
• 期刊: BMC microbiology
• 影响因子: 4.2
• 作者: Holdsworth SR;Law CJ
• 通讯作者: Law CJ

A single-component multidrug transporter of the major facilitator superfamily is part of a network that protects Escherichia coli from bile salt stress.

• DOI: 10.1111/mmi.12597
• 发表时间: 2014-05
• 期刊: Molecular microbiology
• 影响因子: 3.6
• 作者: Paul S;Alegre KO;Holdsworth SR;Rice M;Brown JA;McVeigh P;Kelly SM;Law CJ
• 通讯作者: Law CJ

Measurement of Proton-driven Antiport in Escherichia coli

大肠杆菌中质子驱动的反向转运的测量

• DOI: 10.21769/bioprotoc.1278
• 发表时间: 2014
• 期刊: BIO-PROTOCOL
• 影响因子: 0.8
• 作者: Holdsworth S