Thiazolyl peptide analogs that provide insight into antibiotic targeting

描述(由申请人提供)：噻唑基多肽具有新颖的支架和已被证明的对危险的抗药性病原体的活性，如耐甲氧西林金黄色葡萄球菌(MRSA)和恶性疟原虫，这使它们成为抗生素开发的理想候选者。尽管它们的效力和迫切需要新的抗生素来对抗新出现的耐药性，但由于缺乏多样化的方法，噻唑基多肽一直未能作为治疗药物开发。这一缺陷也阻碍了对结构-活性关系的研究，以确定它们的新型支架如何指导抗生素的作用模式。最近发现，这些分子来自核糖体前体多肽，这些多肽经过广泛的翻译后修饰，打开了通过基因操作实现快速多样化的大门。为此，本研究旨在创建一种新型的噻唑基多肽表达平台，以提高对噻唑基多肽支架如何为细菌核糖体蛋白质合成提供精致靶向的理解。我们的具体假设是，修饰后的噻唑基肽环结构将具有交替或多个靶点，以抑制革兰氏阳性细菌的生长。初步数据表明，该噻唑基多肽生物合成设备应该容易接受环大小的适度变化。我们打算通过以下具体目标来实现这些目标：1.确定链霉菌中产生噻唑基多肽GE37468的基因簇。ATCC 55365，并将其转移到遗传易驯化的变铅青链霉菌中，创建稳定的噻唑基多肽表达平台。这将建立第一个来自公众可访问的有机体的基因簇，该有机体产生带有29个成员环的噻唑基肽，并使人们对独特的GE37468结构的生物合成有深入的了解。2.利用新颖的变铅青链霉菌表达平台构建合理的、高多样性的环大小改变的GE37468类似物文库。3.以高通量的方式评估GE37468类似物的抗生素活性模式，以更好地理解结构如何提供靶向性。本文提出的研究将在创建、理解和筛选环肽文库的能力方面取得前所未有的飞跃。这项工作有可能通过构建高多样性的遗传文库来创造具有新活性光谱的噻唑基多肽。这对于创造能够对抗病原菌新出现的耐药性的噻唑基多肽疗法的长期目标至关重要。 与公共健康相关：该项目与公共健康相关，因为它提议更好地理解噻唑基多肽如何抑制病原菌中的蛋白质合成。这一理解对于开发新的治疗方法以对抗新出现的细菌耐药性至关重要。

DESCRIPTION (provided by applicant): Thiazolyl peptides possess novel scaffolds and proven activity against dangerous drug resistant pathogens such as methicillin-resistant Staphylococcus auerus (MRSA) and Plasmodium falciparum that make them ideal candidates for antibiotic development. Despite their potency and the pressing need for new antibiotics to combat emerging resistance, thiazolyl peptides have eluded development as therapeutics due to insufficient methods for their diversification. This deficiency has also hindered the study of structure-activity relationships to define how their novel scaffolds direct the antibiotic mode of action. The recent discovery that these molecules arise from ribosomal precursor peptides which undergo extensive post-translational modifications opens the door to rapid diversification through genetic manipulation. Towards this end, this research aims to create a novel thiazolyl peptide expression platform to enhance the understanding of how thiazolyl peptide scaffolds provide exquisite targeting of ribosomal protein synthesis in bacteria. Our specific hypothesis is that modified thiazolyl peptide ring structures will have alternate or multiple targets that inhibit the growth of gram positive bacteria. Preliminary data indicates the thiazolyl peptide biosynthetic machinery should readily accept modest changes in ring size. We intend to accomplish these goals through the following specific aims: 1. Identify the gene cluster responsible for the thiazolyl peptide GE37468 production in Streptomyces sp. ATCC 55365 and transfer it to the genetically tractable Streptomyces lividans to create a stable thiazolyl peptide expression platform. This will establish the first gene cluster from a publically accessible organism which produces a thiazolyl peptide with a 29-member ring and give insights into the biosynthesis of the unique GE37468 structure. 2. Create rational and high-diversity libraries of GE37468 analogs with altered ring sizes using the novel S. lividans expression platform. 3. Evaluate the mode of antibiotic activity for GE37468 analogs in a high through-put manner to create a better understanding of how structure provides targeting. The research proposed herein will take an unprecedented leap in the ability to create, understand, and screen libraries of cyclic peptides. This work has the potential to create thiazolyl peptides with novel spectrums of activity through the construction of high diversity genetic libraries. This is vital to the long-term goal of creating thiazolyl peptide therapeutics that can combat emerging resistance in pathogenic bacteria. PUBLIC HEALTH RELEVANCE: This project is relevant to public health because it proposes to develop greater understanding of how thiazolyl peptides inhibit protein synthesis in pathogenic bacteria. This understanding is vital to the development of new therapeutics to combat emerging bacterial resistance.