David Braley Center for Antibiotic Discovery, M.G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, ON, Canada.
David Braley Center for Antibiotic Discovery, M.G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, ON, Canada.
Methods Enzymol. 2022;665:325-346. doi: 10.1016/bs.mie.2021.11.009. Epub 2021 Dec 7.
Glycopeptide antibiotics are essential drugs used to treat infections caused by multi-drug resistant Gram-positive pathogens. There is a continuous need for new antibiotics, including GPAs, to address emerging resistance and offer desirable pharmacological profiles for improved efficacy. Microbial natural products are proven sources of antibiotics, and this source has dominated drug discovery over the past century. Bacteria from the phylum Actinobacteria are particularly renowned for producing a diverse range of bioactive natural products including glycopeptide antibiotics. The traditional approach to mining this resource is through the culture and extraction of natural products followed by assay for cell-killing activity. Unfortunately, this method no longer efficiently yields new antibiotic leads, delivering instead known compounds. Whole-genome sequencing programs on the other hand are revealing thousands of unexplored natural product biosynthetic gene clusters in the chromosomes of Actinobacteria. These gene clusters encode the necessary enzymes, transport and resistance mechanisms, along with regulatory elements for the biosynthesis of a variety of antibiotics. Identification of uncharacterized or cryptic biosynthetic gene clusters to unlock the chemical "dark matter" represents a new direction for the discovery of new drug candidates. This chapter discusses the identification of glycopeptide antibiotic biosynthetic gene clusters in microbial genomes, the improved production of these antibiotics using the GPAHex synthetic biology platform, and methods for their purification.
糖肽类抗生素是治疗多重耐药革兰阳性病原体感染的重要药物。不断需要新的抗生素，包括糖肽类抗生素，以应对新出现的耐药性，并提供理想的药理学特性以提高疗效。微生物天然产物是抗生素的可靠来源，这一来源在过去一个世纪主导了药物发现。放线菌门的细菌特别以产生多种生物活性天然产物而闻名，包括糖肽类抗生素。挖掘这种资源的传统方法是通过培养和提取天然产物，然后进行细胞杀伤活性测定。不幸的是，这种方法不再有效地产生新的抗生素先导化合物，而是提供已知的化合物。另一方面，全基因组测序计划揭示了放线菌染色体中数以千计尚未开发的天然产物生物合成基因簇。这些基因簇编码合成各种抗生素所需的酶、运输和抗性机制以及调控元件。鉴定未表征或隐匿的生物合成基因簇，以释放化学“暗物质”，代表了发现新药物候选物的一个新方向。本章讨论了在微生物基因组中鉴定糖肽类抗生素生物合成基因簇的方法，使用 GPAHex 合成生物学平台提高这些抗生素的产量，以及它们的纯化方法。
