Chemical and Biological Studies of the Guadinomines

简介(申请人提供)：该奖学金有三个目标。第一个目标是通过合成和生物合成相结合的方法，使用生成愈创木酚胺的类似物。将合成假设驱动的类似物，每个类似物都解决一个问题，即哪些结构特征是III型分泌系统(TTSS)抑制所必需的。还将制造类似物，作为前体导向的生物合成的底物，这将允许快速形成愈创木酚胺类似物。第二个目标是设计一种改进的愈创木酚胺分离方案。天然愈创木酚胺A和B的含量约为1毫克/L，需要繁琐的纯化程序。由于愈创木酚胺A和B都含有邻二胺，这是一种不常见的结构亚基，我们将利用其反应性来加强其分离。我们已经证明，二胺与含氟的醛在室温下可以干净地反应生成氨基，而不需要添加催化剂(May，Khosla-未发表的结果)。这将允许从培养介质中的剩余材料中固相萃取氟。第三个目标是确定TTSS对愈创木酚胺类化合物的抑制范围，并确定它们如何影响其靶标。我们已经证明植物病原菌黄单胞菌被愈创木酚胺A抑制(5月，Mudgett-未发表的结果)，并将继续用天然愈创木酚胺及其类似物来研究植物病原菌。我们将使用小肠结肠炎耶尔森菌试验来确定哪些效应蛋白、结构蛋白和伴侣受到愈创木酚胺的影响。这一知识将使第二代设计新的、更强大的类似物成为可能。 公共卫生相关性：致病性革兰氏阴性菌使用III型分泌系统(TTSS)感染宿主。该提案的目标是研究愈创木酚胺类化合物如何抑制TTS，并制造新的更有效的类似物。

DESCRIPTION (provided by applicant): This fellowship has three Aims. The first Aim is to use the create analogs of the guadinomines through a combined synthetic and biosynthetic approach. Hypothesis-driven analogs will be synthesized, each addressing a question about which structural features are necessary for Type III Secretion System (TTSS) inhibition. Analogs will also be made that should serve as substrates for precursor-directed biosynthesis, which should allow the rapid formation of guadinomine analogs. The second Aim is to devise an improved isolation protocol for guadinomines. Natural guadinomines A and B are present at levels of ~1 mg/L of culture media, and require tedious purification procedures. Since guadinomines A and B both contain a vicinal diamine, an uncommon structural subunit, we will exploit its reactivity to enhance its isolation. We have shown that diamines react cleanly with fluorine-containing aldehydes to make aminals at room temperature without added catalysts (May, Khosla - unpublished results). This will allow for solid phase fluorous extraction from the remaining material in the culture media. The third Aim is to determine the scope of TTSS inhibition of the guadinomines, and determine how they affect their targets. We have shown that the plant pathogen Xanthomonas is inhibited by guadinomine A (May, Mudgett - unpublished results) and will continue to study plant pathogens with natural guadinomines and their analogs. We will use a Yersinia enterolitica assay to determine which effector proteins, structural proteins, and chaperones are affected by guadinomines. This knowledge will enable a second-generation design of new and more potent analogs. PUBLIC HEALTH RELEVANCE: Pathogenic Gram-negative bacteria use a Type III Secretion System (TTSS) to infect hosts. The goals of the proposal are to study how the guadinomines inhibit TTSS, and to make new and more potent analogs.