MICROBIAL DEGRADATION OF AGENT ORANGE

描述：(改编自调查人员摘要)环境释放 大量合成的氯化化合物的形式 除草剂/杀虫剂、溶剂、制冷剂等，产生了主要 对其对人类健康的影响表示关切，因为 许多这样的化合物的持久性。这些化合物的持久性是 反映了自然微生物无法将它们用作 是碳和能源的唯一来源。许多微生物可以利用简单的 氯化化合物，如3-氯苯甲酸酯(3Cba)或 2，4-二氯苯氧乙酸酯(2，4-D)作为其唯一碳源，但不能 利用氯化程度较高的形式，如2，4，5-三氯苯氧乙酸 (2，4，5-T)等。在含有2，4，5-T的恒化器中进行强选择 作为碳的唯一主要来源(定向进化)，它一直 可能分离到一株可利用的洋葱假单胞菌AC1100 2，4，5-T作为其唯一的碳和能源来源。大黄鱼的分子克隆 3Cba/2，4-D以及2，4，5，-T降解的基因都表明 而3Cba和2，4-D异化的氯代酚(Clc)基因是 高度同源性，2，4，5-T降解性(TFT)基因没有同源性 与大量假单胞菌和其他 细菌。申请人提议进行实验，以了解 TFT基因及其产物以确定2，4，5-T是如何 被AC1100及其监管模式降级。测序和 将尝试大量TFT基因的高表达以获得 有关允许有效渗透的酶的作用的信息 2，4，5-T及其随后的脱氯和氧化。类似地，一个 调节蛋白参与儿茶酚(CAT)的正向调节 基因将被纯化，其与启动子的结合方式 结构基因将被描绘出来，以定义这一性质 蛋白质-DNA相互作用对猫基因功能表达的影响这些 数据将提供有关生物进化模式的宝贵信息 自然界中的结构和调节基因以及化学药物中的 对氯化化合物的释放和可获得性的反应。

DESCRIPTION: (Adapted from investigators abstract) Environmental release of a large number of synthetic chlorinated compounds in the form of herbicides/pesticides, solvents, refrigerants etc., has created major concerns with regard to their effect on human health because of the persistence of many such compounds. The persistence of these compounds is a reflection of the inability of natural microorganisms to utilize them as a sole source of carbon and energy. Many microorganisms can utilize simple chlorinated compounds such as 3-chlorobenzoate (3Cba) or 2,4-dichlorophenoxyacetate (2,4-D) as their sole carbon source but cannot utilize higher chlorinated forms such as 2,4,5-trichlorophenoxy acetic acid (2,4,5-T) and others. Under strong selection in a chemostat with 2,4,5-T as the only major source of carbon (directed evolution), it has been possible to isolate a strain of Pseudomonas cepacia AC1100 that can utilize 2,4,5-T as its sole source of carbon and energy. Molecular cloning of the genes both for 3Cba/2,4-D as well as 2,4,5,-T degradation has shown that while the chlorocatechol (clc) genes for 3Cba and 2,4-D dissimilation are highly homologous, the 2,4,5-T degradative (tft) genes show no homology with any of the genomic DNA of a large number of pseudomonads and other bacteria. The applicant proposes experiments to understand the nature of the tft genes and their products to determine exactly how 2,4,5-T is degraded by AC1100 and its mode of regulation. Sequencing and hyperexpression of a number of tft genes will be attempted to obtain information about the role of enzymes that allow efficient permeation of 2,4,5-T and its subsequent dechlorination and oxidation. Similarly, a regulatory protein involved in the positive regulation of catechol (cat) genes will be purified and its mode of binding with the promoter of the structural genes will be delineated to define the nature of this protein-DNA interaction for functional expression of the cat genes. These data would provide valuable information about the mode of evolution of structural and regulatory genes in nature as well as in chemostats in response to the release and availability of chlorinated compounds.