BIO-ORGANIC MECHANISMS OF PEPTIDE ANTIBIOTICS

细菌对抗生素的抗药性严重限制了我们战胜传染病的能力。几乎在所有医院获得性病原体-抗菌药组合中都出现了耐药病例。很快，我们最严重的传染病威胁将无法治愈，因为我们的有效抗菌素武器库正在减少。我们的长期研究目标是开发具有生物学意义的多肽抗生素的合成途径，深入了解它们的作用机制/作用模式，并将所获得的知识应用于开发对耐药表型具有更高活性的替代抗生素。本提案描述了雷莫拉宁的全合成和机制表征，雷莫拉宁是一种新型的β-片状脂多肽抗生素，已被证明对三种重要的革兰氏阳性条件致病菌--甲氧西林耐药金黄色葡萄球菌、万古霉素耐药屎肠球菌和头孢菌素耐药肺炎链球菌具有活性。通过一种不清楚的机制，雷莫拉宁似乎在MURG水平上阻止细菌细胞壁的发育，MURG是一种糖基转移酶，参与肽聚糖生物合成的中间阶段。由于Murg活性对细菌细胞壁的正常发育是必不可少的，因此它是抗菌设计的一个有吸引力的目标。利用雷莫拉宁的临床抗生素潜力，关键在于获得其结构的合成途径，并解开其作用机制的最密切细节。为了实现这一目标，我们将协同融合全合成、机械酶学和蛋白质生物物理学，以完全将结构与抗生素功能相关联。我们计划用固相法合成雷莫拉宁及其类似物，从而提供一条有利于调节其物理化学性质的通用合成路线。我们计划确定雷莫拉宁的分子靶点，并确定抑制复合体的相互作用能量、特异性和结构。我们将评估雷莫拉宁对Murg反应和发生在细菌细胞膜外表面的机械相关的肽聚糖糖基化交联反应的抑制作用。总而言之，这些研究将为拉莫拉宁抑制肽聚糖生物合成的机制提供清晰的图景，并促进能够对抗细菌耐药性的新一代抗生素的设计、合成和生物学评价。

Bacterial resistance to antibiotics has seriously limited our capacity to overcome infectious disease. Cases of resistance have emerged in virtually all hospital-acquired pathogen-antimicrobial combinations. Soon our most serious infectious threats will be untreatable given our dwindling arsenal of effective antibiotics. Our long-term research goals are to develop synthetic access to biologically interesting peptide antibiotics, to gain insight into their mechanism/mode of action, and to apply the knowledge gained to the development of alternative antibiotics with improved activity against resistant phenotypes. This proposal describes the total synthesis and mechanistic characterization of Ramoplanin, a novel beta-sheet lipodepsipeptide antibiotic with proven activity against methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus faecium, and cephalosporin-resistant Streptococcus pneumonia, three important Gram positive opportunistic human pathogens. By an unclear mechanism, Ramoplanin appears to arrest bacterial cell wall development at the level of MurG, a glycosyltransferase involved in an intermediate stage of peptidoglycan biosynthesis. Since MurG activity is essential for proper bacterial cell wall development, it is an attractive target for antibacterial design. Harnessing the clinical antibiotic potential of Ramoplanin critically hinges on gaining synthetic access to its structure and deconvoluting the most intimate details of its mechanism of action. To accomplish this we will synergistically merge total synthesis, mechanistic enzymology and protein biophysics to completely correlate structure to antibiotic function. We plan to synthesize Ramoplanin and related analogues using solid-phase methods, thus providing a general synthetic route to favorably modulate its physiochemical properties. We plan to identify the molecular target of Ramoplanin and determine the interaction energies, specificities, and structure of the inhibitory complex. We will assess the inhibitory effect of Ramoplanin on the MurG reaction and on the mechanistically related peptidoglycan transglycosylation cross-linking reaction that takes place on the outer surface of the bacterial cell membrane. Collectively these studies will provide a clear picture of the mechanism of Ramoplanin inhibition of peptidoglycan biosynthesis and promote the design, synthesis, and biological evaluation of a new generation of antibiotics capable of combating bacterial resistance to antibiotics.