STRUCTURE/FUNCTION OF BIPHENYL DIOXYGENASES

描述（改编自申请人的摘要）：本项目涉及 多环芳烃降解酶的结构和功能 芳族和卤代芳族化合物。 其中一些化合物，如 多氯联苯和二恶英是众所周知的健康持久性污染物， 环境意义。 X射线晶体学将用于确定 PCB中酶的各种状态的高分辨率结构 降解途径 外二醇双加氧酶2，3-二羟基联苯1，2-双加氧酶催化 开环的关键步骤。 为了测试机械的建议，结构 对来自假单胞菌LB 400的酶的研究将扩展到几个 二元和三元络合物类似于所提出的中间体。 被认为具有催化意义的残基的定点变体 也将被审查。 其他研究将评估结构 与底物选择性和代谢物有关的催化僵局的起源 抑制作用 第二外二醇双加氧酶的晶体结构， 表达了底物偏好的重要差异， 将测定和分析对代谢物抑制的抗性。 这 第二种酶属于不同的结构类别，并且是通用的。 对蛋白质的结构进化感兴趣。 将开始对第二种关键酶的组分进行结构研究 从这个途径，环羟基化双加氧酶联苯 2，3-双加氧酶。 在这种情况下，一个目标是分析 Rieske分量相对于影响其 独特的氧化还原特性。 拟议的研究是国际努力的一部分， 结构研究与其他人的补充研究。 的 这项工作的总体目标是：了解 参与联苯-PCB途径的酶;了解和改变 在途径的关键步骤基板偏好;了解和删除 催化障碍，如双加氧酶的不可逆抑制， 氯化代谢物;并且，在一般意义上，利用 基础研究沿着微生物发展的各个前沿 具有增强的降解广泛的芳香族化合物和 卤代芳香族污染物。

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.