Physical Mechanisms of Bacterial Genome Maintenance

描述(申请人提供)：细菌单链(Ss)DNA结合蛋白(SSB)在基因组生物学中起着重要的保护和组织作用。为了保护单链DNA免受潜在的损伤，多个SSB组装成“核小体”支架，其结构尚不清楚。SSDNA/SSB复合体远不是惰性的，而是活跃的DNA加工中心，至少有12种不同的酶通过利用与SSB直接的蛋白质-蛋白质相互作用来获得基因组底物。在迄今为止研究的所有案例中，SSB的柔性C末端(SSB-Ct)形成了异源蛋白的对接位置。蛋白质如何与SSB-Ct序列结合，以及这些重要的相互作用如何影响基因组维护酶的活性仍然没有明确的定义。鉴于SSB与异源蛋白相互作用的重要性，阻断这些SSB蛋白复合体形成的抑制剂作为新型抗菌剂具有巨大的潜力。这项建议的最终目的是阐明单链DNA/单链断裂底物的结构，揭示酶如何利用与单链断裂的直接结合来处理这些结构，并表征阻断异源蛋白与单链断裂结合的抑制剂的抗菌性能。该提案结合了生化、结构和遗传方法来解决这些问题。与公共健康相关：基因组维护过程确保细胞中遗传信息的准确性，并提供机制，使这些信息能够被忠实地复制并分发给子细胞。这些都是所有细胞必不可少的过程，需要许多不同蛋白质的精确协调。这项提议旨在了解这些蛋白质成分中的几个是如何在细胞中协调的，并研究选择性地阻断细菌中这种协调的抑制剂的抗菌机制。

DESCRIPTION (provided by applicant): Bacterial single-strand (ss) DNA-binding proteins (SSBs) play essential protective and organizational roles in genome biology. To shield ssDNA from potential damage, multiple SSBs assemble into "nucleosome-like" scaffolds, the structures of which are not well understood. Far from being inert, ssDNA/SSB complexes are active DNA processing centers where at least a dozen different enzymes gain access to genomic substrates by exploiting direct protein-protein interactions with SSB. In all cases examined to date, SSB's flexible C-terminus (SSB-Ct) forms a docking site for heterologous proteins. How proteins bind to the SSB-Ct sequence and how these essential interactions affect the activities of genome maintenance enzymes remains poorly defined. Given the importance of SSB's interactions with heterologous proteins, inhibitors that block formation of these SSB protein complexes have great potential as novel anti-bacterial agents. The ultimate goals of this proposal are to elucidate the structures of ssDNA/SSB substrates, to reveal how enzymes take advantage of direct binding to SSB to process these structures, and to characterize the anti-bacterial properties of inhibitors that block heterologous protein association with SSB. The proposal brings together biochemical, structural, and genetic approaches to address these questions. PUBLIC HEALTH RELEVANCE: Genome maintenance processes ensure the accuracy of genetic information in cells and provide mechanisms whereby this information can be faithfully duplicated and distributed to daughter cells. These are essential process for all cells and require precise coordinate of many different proteins. This proposal aims to understand how several of these protein components are coordinated in cells and to investigate the anti-bacterial mechanisms of inhibitors that selectively block this coordination in bacteria.