Structural Biology of Bacterial Copper Resistance

Rosenzweig98744081。该项目的长期目标是在原子细节上了解细菌中铜稳态的分子机制。许多细菌对各种有毒金属离子的抗性系统已被遗传学鉴定出来，但尚不清楚特定基因产物如何在分子水平上结合以提供抗性。拟研究大肠杆菌质粒携带的铜抗性系统pco，该系统编码7种不同的蛋白，包括质周蛋白PcoA、PcoC和PcoE。PcoA、PcoC和PcoE都含有不寻常的富含蛋氨酸和/或组氨酸的序列基序，这些基序与任何已知的铜结合基序都不相似。该研究计划涉及x射线晶体学、生物化学和生物物理学方法的结合。具体目标是：1)求解PcoC中Cu（II）、Cu(I)和apo形态的高分辨率x射线结构；2)求解PcoE中铜和apo形态的高分辨率x射线结构；3)求解PcoA的结晶结构；4)研究PcoA与PcoC之间的相互作用。由此得到的结构信息有望揭示新的铜结合位点，并扩展生物学中铜中心的现有知识。铜是微生物的基本元素，但也有潜在的危害。由于大量的铜是有毒的，许多由细菌引起的植物疾病都是用含铜的杀菌剂来治疗的。然而，一些细菌已经进化出了抵抗铜的系统。为了治疗由这些抗性细菌引起的作物疾病，需要了解铜抗性的机制。已经确定了一些参与抗性系统的蛋白质，但这些蛋白质如何与铜相互作用尚不清楚。本提案中描述的项目侧重于发现一个这样的蛋白质家族如何结合和运输铜。所获得的信息有望对农业有重要的实际应用。此外，所提出的工作将扩展铜化学的基础知识。

Rosenzweig98744081. TechnicalThe long term goal of this project is to understand in atomic detail the molecular mechanisms of copper homeostasis in bacteria. A number of bacterial resistance systems for various toxic metal ions have been identified genetically, but it is not known how the specific gene products combine on the molecular level to provide resistance. The proposed research focuses on the Escherichia coli plasmid-borne copper resistance system pco, which encodes 7 different proteins, including the periplasmic proteins PcoA, PcoC, and PcoE. PcoA, PcoC, and PcoE all contain unusual methionine and/or histidine rich sequence motifs which do not resemble any known copper binding motifs. The research plan involves a combination of X-ray crystallographic, biochemical, and biophysical approaches. The specific aims are 1) to solve the high resolution X-ray structures of the Cu(II), Cu(I), and apo forms of PcoC, 2) to solve the high resolution X-ray structures of the copper and apo forms of PcoE, 3) to crystallize and solve the structure of PcoA, and 4) to study interactions between PcoA and PcoC. The resultant structural information is expected to reveal novel copper binding sites and to expand existing knowledge of copper centers in biology.2. Non-technicalCopper is an essential element in microorganisms, but is also potentially harmful. Because large quantities of copper are toxic, a number of plant diseases caused by bacteria are treated with copper containing bacteriocides. Some bacteria have evolved systems to resist copper, however. In order to treat crop afflictions caused by these resistant bacteria, an understanding of the mechanism of copper resistance is required. A number of proteins involved in resistance systems have been identified, but exactly how these proteins interact with copper is not understood. The projects described in this proposal focus on discovering how one such family of proteins binds and transports copper. The information gained is expected to have important practical applications for agriculture. Furthermore, the proposed work will expand fundamental knowledge of copper chemistry.