Biosynthesis of nucleoside antibiotics targeting bacterial translocase

描述(申请人提供)：多重耐药病原体的出现正成为全球范围内的问题，新抗生素的发现继续下降。这项提议的广泛的、长期的目标是发现、表征和开发一种新的结构类抗生素-核苷抗生素-靶向参与肽聚糖细胞壁生物合成的细菌转位酶I。本建议的具体目的是(I)表征脂肽-核苷类抗生素家族中发现的氨基-核糖基部分的组装和并入，以及(Ii)从功能和机械上表征催化新颖或不寻常的生化反应的酶，其中以一类新的丝氨酸羟甲基转移酶为代表，这些酶被假设催化羟醛缩合形成不寻常的非蛋白生成氨基酸。通过使用为脂肽核苷A-90289产生菌开发的强大的遗传系统进行体内研究，采用基因失活和交叉互补，将实现特定目的I和II，并将开发重组蛋白用于体外功能和机制研究。这一结果将建立一种将核糖基单元整合到天然产品支架中的新机制，并将为进入高碳核苷抗生素领域建立一个范例。这一结果将对我们利用遗传信息寻找新的核苷抗生素的长期目标(基因到化学型方法)至关重要，将允许使用组合生物合成和全合成来实现母体支架的结构多样化，并将为设计具有更好的生物兼容性和药理学特性的第二代抗生素提供基础。

DESCRIPTION (provided by applicant): The emergence of multiple drug resistant pathogens is becoming problematic worldwide, and the discovery of new antibiotics continues to decline. The broad, long-term objective of this proposal is to discover, characterize, and develop a new structural class of antibiotics-nucleoside antibiotics-that target bacterial translocase I involved in peptidoglycan cell wall biosynthesis. The specific aims of this proposal are (I) to characterize the assembly and incorporation of the aminoribosyl moiety found within the family of lipopeptidyl-nucleoside antibiotics and (II) to functionally and mechanistically characterize enzymes catalyzing novel or unusual biochemical reactions represented herein by a new family of serine hydroxymethyltransferase-like enzymes that are hypothesized to catalyze an aldol-type condensation to form unusual nonproteinogenic amino acids. Specific Aim I and II will be achieved by using the robust genetic system developed for the lipopeptidyl nucleoside A-90289-producing strain for in vivo studies employing gene inactivation and cross-complementation, and recombinant proteins will be exploited for functional and mechanistic studies in vitro. The results will establish a new mechanism for incorporating ribosyl units into natural product scaffolds and will establish a paradigm for the entry into high-carbon nucleoside antibiotics. The results will be essential for our long-term goals of searching for new nucleoside antibiotics using genetic information (a genotype-to-chemotype approach), will allow for the structural diversification of the parent scaffolds using combinatorial biosynthesis and total synthesis, and will provide the basis for the design of second generation antibiotics with improved biocompatibility and pharmacological properties.