STRUCTURE/FUNCTION OF BIPHENYL DIOXYGENASES

描述(改编自申请者摘要)：本项目涉及 多环化合物降解相关酶的结构与功能 芳香族和卤代族化合物。许多这样的化合物，例如 多氯联苯和二恶英是众所周知的持久性健康污染物， 对环境具有重要意义。X射线结晶学将被用来确定 印刷电路板中各种酶的高分辨结构 降解途径。 外源性双加氧酶2，3-二羟基联苯1，2-双加氧酶催化 环裂解的关键步骤。测试机械方案、结构方案 假单胞菌LB400酶的研究将扩展到几个 类似于建议的中间体的二元和三元络合物。 残基的定点变异被认为具有催化意义 还将接受检查。更多的研究将评估结构 与底物偏好和代谢物有关的催化僵局的起源 抑制力。第二种外源性双加氧酶的晶体结构 表达底物偏好的重要差异以及 将测定和分析对代谢物抑制的抵抗力。这 第二种酶属于不同的结构类别，具有通用性 对蛋白质结构进化的兴趣。 将开始对第二种关键酶的成分进行结构研究 从这个途径，环羟化双加氧酶联苯 2，3-双加氧酶。在这种情况下，一个目标是分析 Rieske组件与影响其 独特的氧化还原特性。 拟议的研究是国际努力的一部分，该努力加入了我们的 结构调查与其他研究的补充。这个 这项工作的总体目标是：了解 参与联苯-多氯联苯途径的酶；理解和改变 在途径的关键步骤中的底物偏好；理解和去除 催化僵局，如对双加氧酶的不可逆转抑制 氯化代谢物；以及，在一般意义上，利用 微生物发展中多个前沿领域的基础研究 具有更强的降解多种芳香族和有机污染物能力的系统 卤代芳香污染物。

DESCRIPTION (Adapted from applicant's abstract): This project address the structure and function of enzymes involved in the degradation of polycyclic aromatic and haloaromatic compounds. A number of these compounds, such as the PCBs and dioxins, are well-known as persistent pollutants of health and environmental significance. X-ray crystallography will be used to determine high resolution structures of various states of enzymes in the PCB degradative pathway. The extradiol dioxygenase 2,3-dihydroxybiphenyl 1,2-dioxygenase catalyzes the key step of ring cleavage. To test mechanistic proposals, structural studies of the enzyme from Pseudomonas LB400 will be extended to several binary and ternary complexes analogous to proposed intermediates. Site-directed variants of residues believed to be of catalytic significance will also be examined. Additional studies will evaluate the structural origins of catalytic impasses related to substrate preference and metabolite inhibition. The crystal structure of a second extradiol dioxygenase that expresses important differences in substrate preference as well as resistance to metabolite inhibition will be determined and analyzed. This second enzyme belongs to a different structural class and is of general interest with respect to the structural evolution of proteins. Structural studies will be initiated for components of a second key enzyme from this pathway, the ring-hydroxylating dioxygenase biphenyl 2,3-dioxygenase. In this case, one goal is to analyze the structure of the Rieske component with respect to structural features that influence its distinct redox properties. The proposed studies are part of an international effort that joins our structural investigations with the complementary research of others. The general goals of this effort are: to understand the mechanisms of the enzymes involved in the biphenyl-PCB pathways; to understand and alter substrate preferences at key steps in the pathways; to understand and remove catalytic impasses such as the irreversible inhibition of dioxygenases by chlorinated metabolites; and, in a general sense, to exploit the results of basic research along a variety of fronts in the development of microbial systems with an enhanced ability to degrade a broad range of aromatic and haloaromatic pollutants.