Cell envelope of bacteria producing glycopeptide antibiotics

糖肽类抗生素通过阻断转肽和转糖基化来抑制细菌细胞壁的合成。大多数病原体对糖肽的耐药机制涉及替代细胞壁前体的合成。由于抗性机制的复杂性，其中包括至少三个必需的酶，预计糖肽生产者是抗性基因的进化起源。然而，没有发现生产Amycolatopsis物种的抗生素生物合成基因簇中的必需的抗性基因。唯一的功能，这是本地化的集群，并可能与电阻是一个假定的ABC-转运，以及两个组件的调节系统和羧肽酶。后者在万古霉素抗性转座子上具有对应物，但不表达抗性。拟议的研究应回答以下问题：* 生产者如何保护其细胞壁免受抗生素的作用？* 辅助抗性基因是如何参与抗性的？* 不同的细胞壁前体在指数期和稳定期（=生产期）合成，它们的合成是如何调节的？这将通过假定耐药基因的突变和过表达研究进行分析。编码“正常”细胞壁合成的基因将被分离和表征。将研究基因的调节以及蛋白质的合成，关于抗性和抗生素生产的共同调节。此外，我们计划研究不同时间点和突变体中胞壁素前体的化学结构（与化学合作）。

Glycopeptide antibiotics inhibit the synthesis of the bacterial cell wall by blocking the transpeptidation and transglycosylation. The resistance mechanism in most pathogens against glycopeptides involves the synthesis of alternative cell wall precursors. Due to the complexity of the resistance mechanism which comprises of at least three essential enzymes, it was expected that glycopeptide producers are the evolutionary origin of the resistance genes. However, none of the essential resistance genes was found in the antibiotic biosynthesis gene clusters of the producing Amycolatopsis species. The only functions which are localised in the cluster and may be related to resistance are a putative ABC-transporter, as well as a two component regulatory system and a carboxypeptidase. The latter have counterparts on vancomycin resistance transposons, but are dispensable for expressing resistance. The proposed research should answer the following questions: * How does the producer protect its cell wall from the action of its antibiotic? * How are the accessory resistance genes involved in resistance? * Are different cell wall precursors synthesized during exponential and stationary (=production) phase and how is their synthesis regulated? This will be analysed by mutation and overexpression studies of the putative resistance genes. The genes encoding the synthesis of the "normal" cell wall will be isolated and characterised. The regulation of the genes as well as the synthesis of the proteins will be investigated, with respect to co-regulation of resistance and antibiotic production. Furthermore, we plan to study the chemical structure of the murein precursors at different time points and in the mutants (cooperation with chemistry).