REGULATION AND STRUCTURE OF THE MER OPERON

我们将研究细菌质粒中金属反应基因的调控- 测定汞(Hg)抗性(Mer)操纵子。MER轨迹是非常 广泛存在于真细菌中，并介导无机物和 有机汞化合物。汞结合基序的惊人保守性 人类铜贮藏病中蛋白质中的聚合体蛋白 (孟克斯和威尔逊病)强调了MER作为一种 了解人类接触有毒金属的模型。此外，急性或 动物宿主长期暴露于汞化合物会导致 携带Mer基因座的细菌在正常菌群中的增殖。 在细菌中，汞解毒基因的表达是 由一种新的汞结合转录调控因子MERR控制。 因此，汞对这个系统中基因表达的响应调节是， 一种过渡金属调控基因的模型，已被发现 有几个非金属基因调控中看不到的不同寻常的方面 代谢物。(L)我们将用分子遗传学来探索相互作用 野生型和突变型MERR蛋白的Mer操纵子-启动子区域 并用RNA聚合酶建立了分子基础 转录抑制和激活。(2)我们已经发现 表达也受转录提前终止的控制 (即，Mer有一个非常有效的中间操纵子转录衰减器)。我们 将利用遗传学来分析这种衰减器的成分并确定 并对所涉及的顺式和反式作用剂进行表征。(3)我们有 最近发现了宿主细胞氢过氧化物酶令人惊讶的作用 (包括过氧化氢酶)在汞抗性的表达中，并将定义 这一作用以及探索其他宿主染色体基因在汞中的重要作用 解毒。(4)我们还将启动一个新的项目，关于 通过分离识别金属的MERR突变体与金属离子结合 除汞外，并通过对其纯化的蛋白质产品进行表征 核磁共振、电子自旋共振和共振拉曼光谱。

We will study metal-responsive gene regulation in the bacterial plasmid- determined mercury (Hg) resistance (mer) operon. The mer locus is very widely found in eubacteria and mediates detoxification of inorganic and organic Hg compounds. The striking conservation of Hg binding motifs of mer proteins in the proteins altered in the human copper storages diseases (Menkes' and Wilson's diseases) underscores the importance of mer as a model for understanding human exposure to toxic metals. Moreover, acute or chronic exposure of an animal host to mercury compounds results in the proliferation in the normal flora of bacteria which carry the mer locus. In the bacterium, expression of the mercurial detoxification genes is controlled by a novel Hg-binding transcriptional regulator, MerR. Mercury-responsive regulation of gene expression in this system is, thus, a model for regulation of genes by transition metals and has been found to have several unusual aspects not seen in gene regulation by non-metallic metabolites. (l) We will use molecular genetics to probe the interactions of wildtype and mutant merR proteins with the mer operator-promoter region and with RNA polymerase in order to establish the molecular basis for transcriptional repression and activation. (2) We have discovered that mer expression is also controlled by premature transcriptional termination (i.e. mer has a very efficient mid-operon transcriptional attenuator). We will use genetics to dissect the elements of this attenuator and identify and characterize the cis- and trans-acting agents involved. (3) We have recently discovered a surprising role for the host cell hydroperoxidases (including catalase) in the expression of Hg resistance and will define this role and explore other host chromosomal genes important in Hg detoxification. (4) We will also begin a new project on the interaction of MerR with metal ions by isolating merR mutants which recognize metals other than Hg and by characterizing their purified protein products using NMR, ESR, and resonance Raman spectroscopy.