BIODEGRADATION OF MERCURIAL ANTISEPTICS AND PESTICIDES

有机汞的抗性和生物降解相关基因(OMR) 包括防腐剂硫代硫酸盐、消毒剂苯汞 醋酸盐和神经毒性杀菌剂甲基汞，经常在 细菌质粒以及对无机化合物的抗性，如 氯化汞。尽管无机汞的耐受性(HGR)已被研究 广泛地说，人们对有机汞抗性的了解要少得多。 两年前，我们开始研究有机汞的抗性，由 接合的IncM质粒为R831b。虽然R831b的大小为90kb，但它是 携带OMR基因座的最小的自然产生的质粒。vbl.使用 转座子诱变及限制性内切酶分析 R831b只携带两个可选择标记：HGR和OMR。我们发现 这些基因座之间有13.5kb的距离，尽管它们的酶 对编码进行协调管理。我们已经将OMR亚克隆到pBR322中 它与我们之前拥有的HGR mini-IncM复制子兼容 克隆并已开始构建精细结构的限制和缺失 两个地点的地图。在这笔赠款的最后一年，我们将 完成这些基因座的限制性内切酶图谱，并产生两个lac 和Galk融合在亲本质粒和亚克隆衍生物中。 从长远来看，我们建议：(1)使用微细胞多肽分析 鉴定R831b的OmR和HGR基因座的基因产物；(2) 利用插入和缺失检测OMR/HGR的表达调控 突变体和Lac和Galk的融合 R831b基因座)以及异源系统(即包括R831b 和NR1操纵子的相关基因座)，并分离顺式基因。 和R831b中这些基因座的反式调控元件，并比较 (3)采用OMR和HGR DNA作为 探索这些基因座在我们的大量收集中的出现情况 繁殖耐金属细菌；以及(4)对OMR和HGR基因座进行测序 R831b。目前还没有对OMR基因座进行测序，尽管来自 质粒NR1和相关的转座子Tn501已经测序，我们的 多肽数据、DNA杂交数据(我们的未发表和T.Barkay， 个人通信)和免疫扩散数据(S.Silver，个人 通信)表明R831b的HGR基因座没有紧密的亲缘关系 与NR1/Tn501的同源性比较。

Genes for resistance to and biodegradation of organomercurials (Omr) including the antiseptic merthiolate, the disinfectant phenylmercuric acetate, and the neurotoxic fungicide methyl mercury, are often found on bacterial plasmids along with resistance to inorganic compounds such as HgCl2. Though inorganic mercury resistance (Hgr) has been studied extensively, much less is known about organomercury resistance. Two years ago we began to study the organomercury resistance determined by the conjugative IncM plasmid, R831b. Though 90 kb in size, R831b is the smallest naturally occurring plasmid which carries the Omr locus. Using transposon mutagenesis and restriction enzyme analysis we have mapped the only two selectable markers carried by R831b, Hgr and Omr. We found that these loci are separated by 13.5 kb despite the fact that the enzymes they encode are regulated co-ordinately. We have sub-cloned Omr into pBR322 which is compatible with the Hgr mini-IncM replicon we had previously cloned and have begun to construct fine structure restriction and deletion maps of both loci. During the last year of this present grant we will complete the restriction enzyme mapping of these loci and generate both lac and galK fusions in the parental plasmid and in sub-cloned derivatives. We propose for the long term: (1) to use minicell polypeptide analysis to characterize the gene products of the Omr and Hgr loci of R831b; (2) to examine regulation of Omr/Hgr expression using insertion and deletion mutants and lac and galK fusions of both the homologous system (i.e. the R831b loci) as well as of a heterologous system (i.e. including both R831b and the correlate loci of the mer operon of NR1) and to isolate the cis- and trans-acting regulatory elements of these loci in R831b and compare them with those of the NR1 mer operon; (3) to employ the Omr and Hgr DNA as probes to explore the occurrence of such loci in our large collection of multiply metal resistant bacteria; and (4) to sequence the Omr and Hgr loci of R831b. No Omr locus has yet been sequenced and though Hgr loci from plasmid NR1 and the related transposon Tn501 have been sequenced, our polypeptide data, DNA hybridization data (ours unpublished and T. Barkay, personal communication) and immunodiffusion data (S. Silver, personal communication) indicate that the Hgr locus of R831b is not closely related to that of NR1/Tn501.