Synthesis of New Ligands for the Characterization of Siderophore-ExFABP Interacti

摘要 癌症和癌症治疗通常伴随着免疫系统的严重削弱。 细菌是导致大多数贫血癌症患者的外源性感染的原因， 严重的健康风险。初步研究表明，人类铁运载蛋白，脂质运载蛋白的成员， 结合蛋白家族，与微生物铁载体结合，通过 铁的螯合作用。考虑到哺乳动物的铁水平会随着病原体和肿瘤的生长而下降， 通过含铁微生物铁载体复合物靶向结合铁可能是治疗 癌症和细菌感染。我们已经发现，前FABP，一个相关的脂质运载蛋白表达过程中， 鸡胚发育，也结合不同结构类别的铁载体复合物。我们的目标是 化学合成天然存在和设计的细菌铁载体类似物，以实现 对铁载体特异性先天免疫系统相互作用的完整理解。为了解析 铁黄素和Ex-FABP的识别模式，我们将制备绿脓菌螯铁蛋白的氧代和亚甲基变体， 与Ex-FABP结合但不与铁螯铁蛋白结合的假单胞菌铁载体。具体的假设是， 绿脓菌螯铁蛋白的硫唑啉环中的硫原子由于与一个紧密的 花萼中的束缚水分子。用恶唑啉环替代噻唑啉将潜在地模拟 其他高亲和力铁运载蛋白如羧基分枝杆菌素的结合模式。因为自然发生的 绿脓菌螯铁蛋白也以非对映异构体的不稳定混合物存在，我们建议按构型合成 稳定的脯氨酸基类似物，以确定Ex-FABP的离散立体化学要求 约束力我们还将进行（I组）绿脓菌荧光素的第一次化学合成，这是一种结合 特别是前FABP全化学合成的优点包括能够追求合成 类似物和难以从细菌来源获得的同质配体的分离。一旦 这些侧载体的合成纯样品是可用的，我们将进行定量结合研究， 体外生长停滞测定和X射线晶体学。这项研究的结果将有助于 鉴定其他蛋白质特异性微生物铁载体，并为蛋白质突变奠定基础， 微生物选择性抗生素治疗的方法。

ABSTRACT Cancer and cancer therapy are commonly accompanied by a severe weakening of the immune system. Bacteria are responsible for the majority of adventitious infections in neutropenic cancer patients and thus pose a significant health risk. Preliminary studies have shown that human siderocalin, a member of the lipocalin family of binding proteins, binds to microbial ferric siderophores and inhibits bacetrial growth through the sequestration of iron. Given that mamalian iron levels drop in response to pathogens and tumor growth, targeted binding of iron via ferric microbial siderophore complexes may be a useful strategy in the treatment of cancer and bacterial infections. We have found that Ex-FABP, a related lipocalin protein expessed during chicken embryo development, also binds ferric siderophore complexes of distinct structural classes. Our aim is to chemically synthesize naturally occurring and designed analogs of bacterial siderophores in order to achieve a complete understanding of siderophore-specific innate immune system interactions. In an effort to parse the recognition modes of siderocalin and Ex-FABP, we will prepare oxo and methylene variants of pyochelin, a Pseudomonas siderophore that binds to Ex-FABP, but not to siderochelin. The specific hypothesis is that the sulfur atom in the thiozoline ring of pyochelin preculdes binding to siderocalin due to a steric clash with a tightly bound water molecule in the calyx. Replacement of a thiazoline with an oxazoline ring will potentially mimic the binding mode of other high-affinity siderocalins such as the carboxymycobactins. Because naturally occuring pyochelin also exists as an unstable mixture of diastereomers, we propose to synthesize configurationally stable proline-based analogs in order to determine the discrete stereochemical requirements of Ex-FABP binding. We will also undertake the first chemical synthesis of (group I) pyoverdin, a complex peptide that binds specifically to Ex-FABP. The advantages of total chemical synthesis include the ability to pursue synthetic analogs and the isolation of homogenous ligand that is difficult to obtain from bacterial sources. Once synthetically pure samples of these sideophores are available, we will proceed with quanitative binding studies, in vitro growth arrest assays, and x-ray crystallography. Results emanating from this research will aid in the identification of other protein-specific microbial siderophores and lay the foundation for protein mutation as a means toward microbe-selective antibiotic therapy.