INHIBITORS OF BACTERIAL RNA POLYMERASE: "SWITCH REGION"

描述（由申请人提供）：初步研究证实三种天然产物——聚酮衍生的a-pyrone myxpyronin、聚酮衍生的a-pyrone corallopyronin和聚酮衍生的大环内酯ripostatin——通过与RNAP“开关区”的相互作用抑制细菌RNA聚合酶（RNAP）， RNAP是一种结构元件，介导RNAP在转录中结合和保留DNA模板所需的构象变化。这些化合物不抑制真核RNAP I、RNAP II或RNAP III。这些化合物能有效抑制革兰氏阳性和革兰氏阴性细菌的生长，与目前用于治疗细菌感染的细菌RNAP抑制剂（利福霉素抗菌剂、利福平、利福喷丁和利福布汀）无交叉耐药，并且与正在评估用于治疗细菌感染的其他细菌RNAP抑制剂无交叉耐药。这项工作将利用x射线晶体学、集合和单分子荧光共振能量转移、单分子纳米操作、分子克隆、代宿主表达、基于结构的筛选和从头筛选来解决四个具体目标：

DESCRIPTION (provided by applicant): Preliminary work establishes that three natural products-the polyketide-derived a-pyrone myxopyronin, the Dolyketide-derived a-pyrone corallopyronin, and the polyketide-derived macrocylic lactone ripostatin-inhibit bacterial RNA polymerase (RNAP) through interactions with the RNAP "switch region," a structural element that mediates conformational changes required for RNAP to bind and retain the DNA template in transcription. The compounds do not inhibit eukaryotic RNAP I, RNAP II, or RNAP III. The compounds potently inhibit Gram-positive and Gram-negative bacterial growth, exhibit no cross-resistance with the inhibitors of bacterial RNAP in current clinical use in therapy of bacterial infection (the rifamycin antibacterial agents, rifampicin, rifapentine, and rifabutin), and exhibit no cross-resistance with other inhibitors of bacterial RNAP under evaluation for future clinical use in therapy of bacterial infection. The proposed work will use x-ray crystallography, ensemble and single-molecule fluorescence resonance energy transfer, single-molecule nanomanipulation, molecular cloning, surrogate-host expression, structure-based screening, and de novo screening, to address four specific aims: Specific Aim 1: Determination of structures of complexes of RNAP with switch-region-target inhibitors Specific Aim 2: Determination of mechanisms of inhibition of RNAP by switch-region-target inhibitors Specific Aim 3: Cloning, characterization, and surrogate-host expression of biosynthetic gene clusters for switch-region-target inhibitors Specific Aim 4: Identification and characterization of novel switch-region-target inhibitors. The results will enable development of new broad-spectrum antibacterial agents that will be effective against bacterial strains resistant to currently used antibacterial agents. As such, the results will have direct relevance to public health and to development of countermeasures against bacterial strains that could be used in biowarfare or bioterrorism. In addition, the results will contribute to understanding RNAP structure and function and will provide tools for analysis of RNAP structure and function.