Siderophore Synthetases in Three Bacterial Pathogens

超出提供的空间。 缺铁微环境中的细菌合成、输出和回收铁络合剂 因此，螯合物，也被称为铁载体，通过以下方式作为脊椎动物感染的毒力因子 鼠疫中的鼠疫耶尔森氏菌、霍乱流行中的霍乱弧菌和假单胞菌等病原体 铜绿假单胞菌在CF患者肺部感染中的应用各自的铁载体，Yersiniabactin(Ybt)， 弧菌素(Vib)和脓球蛋白(PCH)是由非核糖体多肽合成酶(NRPS)激活产生的 由于铁的枯竭。Yersiniabactin是由非核糖体多肽和聚酮(PK)组成的杂交体。 由NRPS/PKS混合装配线制造。这条装配线的NRPS/PKS和PKS/NRPS开关点 将用纯化的Ybt合成酶多模块亚基进行检测。YBT还具有串联型双噻唑啉 环系统和嵌入环化(Cy)和C-甲基化(MT)结构域的异常C-甲基化 17域Ybt装配线。我们将研究Cy和MT催化域的机制。这个 五氯环己烷装配线涉及四种蛋白质和一系列将被分析的酰基-S酶中间体 用于特定链伸长步骤的时间和位置，包括杂环噻唑啉环还原到 然后通过PchG亚单位和PchF的N-MT结构域进行N-甲基化 分别为亚基。由于Ybt和PCH铁载体蛋白-蛋白质识别的相似性 将检查PCH和YBT装配线内部和之间的域和模块，以开始 建立这种NRP分子的组合生物合成规则。第三种致病铁载体 系统中，Vib合成酶组装一个分支的铁载体，并产生恶唑啉环(从苏氨酸) 而不是PCH和Ybt中的噻唑啉环(来自半胱氨酸)。有两个Cy结构域和三个 可控震源装配线中的缩合(C)结构域和所有五个链延长的功能 催化结构域将通过生化分析和诱变来破译分子逻辑 这些NR.PS组件。理解机制，例如杂环化Cy结构域可能允许 抑制剂的设计，以阻止铁载体的生产这些多模块酶。

EXCEED THE SPACE PROVIDED. Iron chelators are synthesized, exported and retrieved by bacteria in iron-deficient microenvironments aid thus chelatars, also known as siderophores, serve as virulence factors for infections in vertebrates by pathogens such as Yersinia pestis in plague, Vibrio cholerae in cholera epidemics, and Pseudomonas aeruginosa in lung infection in CF patients. The respective siderophores, yersiniabactin (Ybt), vibriobactin(Vib), and pyochelin(Pch),are produced by non-ribosomal peptide synthetases(NRPS),activated by iron depletion. Yersiniabactin is assembled as a hybrid of a nonribosornal peptide and a polyketide (PK) by a mixed NRPS/PKS assembly line. The NRPS/PKS and PKS/NRPS switchpointsof this assembly line will be examined with purified multimodular subunits of Ybt synthetase. Ybt also has a tandem bithiazoline ring system and unusual C-methylations from Cyclization(Cy) and C-methylation(MT) domains embedded in the 17 domain Ybt assembly line. The mechanisms of the Cy and MT catalytic domains will be examined. The Pch assembly line involves four proteins and a cascade of acyl-S-enzyme intermediates that will be analyzed for timing and locus of specific chain elongation steps, including heterocyclic thiazoline ring reduction to thiazolidine and then subsequent N-methylationby the PchG subunit and the N-MT domain of the PchF subunit respectively. Because of the similarity of the Ybt and PCh siderophores protein-protein recognitionof domains and modules within and between the Pch and Ybt assembly lines will be examined to begin to establish rules for combinatorial biosynthesis of such NRP molecules. The third pathogenic siderophore system, the Vib synthetase assembles a branched siderophore and produces an oxazoline ring (from threonine) rather than the thiazoline riongs (from cysteine) in Pch and Ybt. There are two Cy domains and three Condensation (C) domains in the Vib assembly line and the function of all five of thesechain-elongation catalytic domains will be examined by biochemical analysis and mutagenesis to decipher the molecular logic of these NR.PS assemblies. Understanding of mechanism, e.g. of the heterocyclizing Cy domains may allow inhibitor design to block siderophore production by these multimodular enzymes.