Elucidation and inhibition of the biosynthetic pathway to the anthrax stealth siderophore petrobactin

炭疽杆菌是引起炭疽病的细菌，炭疽病是一种经常致命的动物和人类疾病。近年来，它引起了相当大的关注，因为它的孢子有可能被用作生物恐怖毒剂。虽然炭疽病可以用现有的抗生素治疗，如环丙沙星，但炭疽杆菌可以通过基因工程使其对目前所有可用的抗生素产生抗药性。这种基因工程菌株构成的生物恐怖主义威胁将是相当大的，因此需要开发对炭疽杆菌有效的新抗生素。铁是包括炭疽杆菌在内的几乎所有细菌增殖所必需的元素。因此，感染性细菌从宿主获取铁的系统是开发治疗剂的潜在目标。铁载体是大多数细菌分泌的代谢物，与铁离子紧密结合。在哺乳动物细胞内和细胞之间，铁与蛋白质(如转铁蛋白和乳铁蛋白)紧密结合。由感染性细菌产生的铁载体能够从这些蛋白质中去除铁，并将其输送到细菌细胞中。对于几种感染性细菌来说，抑制它们用于铁载体合成的途径被认为会极大地减弱或消除它们引起感染的能力。炭疽杆菌已被证明可以分泌两种铁载体，分别称为杆状杆菌和石油杆菌。虽然杆菌菌素不是炭疽杆菌在小鼠感染模型中生长所必需的，但Petrobactin起着重要的作用。这被归因于Petrobactin而不是bacillibactin避开哺乳动物免疫系统的能力，这表明旨在抑制Petrobactin催化组装的酶的小分子可能是对抗炭疽杆菌的有效抗生素。为了设计这样的抑制剂，需要在分子水平上对这些酶如何催化Petrobactin的组装有一个基本的了解。遗传学研究表明，Petrobactin生物合成途径是两条已知的铁载体生物合成途径的独特杂交，其中一条几乎完全没有在分子水平上被探索。生物化学研究已经开始揭示Petrobactin生物合成的分子细节，并导致了新的和有趣的酶的发现，这些酶在生产有价值的候选药物和其他精细化学品的构建块方面具有潜在的应用价值。本研究的目的是详细研究石油肌动蛋白合成中的一个关键酶的催化性质，并设计、合成和测试该酶家族的第一批抑制剂。它还旨在研究与Petrobactin组装有关的另外两种重要酶的催化特性。这将澄清炭疽杆菌用于合成Petrobactin的途径，目前尚不清楚。

Bacillus anthracis is the bacterium that causes anthrax, a frequently fatal disease of animals and humans. It has attracted considerable attention in recent years because of the potential to use its spores as a biological terror agent. While anthrax can be treated using currently available antibiotics e.g. ciprofloxacin, Bacillus anthracis could be genetically engineered to make it resistant to all currently available antibiotics. The bioterrorism threat posed by such genetically engineered strains would be considerable and there is thus a need to develop new antibiotics that are active against B. anthracis. Iron is an essential element for the proliferation of virtually all bacteria including B. anthracis. As a consequence, the systems used by infectious bacteria to acquire iron from their hosts represent potential targets for the development of therapeutic agents. Siderophores are metabolites excreted by most bacteria that bind tightly to ferric iron. Within and between mammalian cells ferric iron is tightly bound by proteins (e.g. transferrin and lactoferrin). Siderophores produced by infectious bacteria are able to remove the ferric iron from these proteins and transport it into the bacterial cell. For several infectious bacteria, inhibition of the pathways they use for siderophore synthesis is known to strongly attenuate or abrogate their ability to cause infection. B. anthracis has been shown to excrete two siderophores called bacillibactin and petrobactin. While bacillibactin is not required for B. anthracis growth in mouse models of infection, petrobactin plays a significant role. This has been attributed to the ability of petrobactin, but not bacillibactin, to avoid the mammalian immune system, suggesting that small molecules designed to inhibit the enzymes catalysing assembly of petrobactin may be effective antibiotics against B. anthracis. To design such inhibitors a fundamental understanding at the molecular level of how these enzymes catalyse the assembly of petrobactin is required. Genetic studies have shown that the petrobactin biosynthetic pathway is a unique hybrid of two well-known pathways for siderophore biosynthesis, one of which is almost completely unexplored at the molecular level. Biochemical studies have begun to reveal the molecular details of petrobactin biosynthesis and have led to the discovery of novel and interesting enzymes with potential applications in the production of valuable building blocks for the synthesis of drug candidates and other fine chemicals. This proposal aims to investigate the catalytic properties of a key enzyme in petrobactin synthesis in detail, as well as design, synthesise and test the first inhibitors of this enzyme family. It also aims to investigate the catalytic properties of two other important enzymes involved in petrobactin assembly. This will clarify the pathway used by B. anthracis for petrobactin synthesis, which at present is unclear.