Characterization of the Key Enzymes in Polycholorophenol Biodegradation

多氯苯酚是美国最普遍的有机污染物之一。几种类型的多氯苯酚主要通过在木材工业中用作防腐剂、在农业中用作除草剂以及在消费品中用作一般的杀生物剂而被引入环境中。它们在环境中持久存在，因为卤素取代使它们不易于微生物降解。值得注意的是，我们已经确定并表征了参与降解这些有毒化合物的几种微生物酶。这些酶的PI表征揭示了几种新的反应，促使从生物化学和结构的角度进行进一步的研究。PI的长期目标是对降解机制有一个基本的了解，并确定这些外源性污染物生物降解途径中所有参与酶的底物特异性参数。本论文的主要目的是对多氯酚生物降解过程中的三种单加氧酶（TcpA、TftD和PcpB）和一种特殊的双加氧酶（PcpA）进行比较研究。在这些多氯芳族化合物的分解过程中，脱氯至关重要，因为部分或完全脱氯必须在开环和随后的矿化之前发生。然而，酶促脱氯尚未得到大量研究。这项研究提供了一个全面的了解不寻常的特异性和反应催化这些酶。这有助于确定活性位点结构的修饰以拓宽底物范围或加速动力学的可行性。 这可能会导致更好的清理战略，以补救有毒的环境场所。更广泛的影响这项研究正在提高催化效率和范围的底物特异性的生物降解酶能够生物修复的外源性污染物。它正在通过结合生物化学、遗传学、生物物理学和机械学专门知识，对这一问题采取多学科办法。 包括Emiliamo Sanchez在内的两名研究生和几名本科生正在参与这项研究。 高中生也通过华盛顿州立大学现有的NSF资助的K-12外展计划参与研究，将酶学，结构生物学和环境微生物学带到高中课堂和当地的探索中心。

Polychlorophenols are among the most pervasive organic pollutants in the United States. Several types of polychlorophenols are primarily introduced into the environment through their use as preservatives in the wood industry, as herbicides in agriculture, and as general biocides in consumer products. They persist in the environment because halogen substitution makes them recalcitrant to microbial degradation. Significantly we have identified and characterized several microbial enzymes involved in the degradation of those toxic compounds. The PIs characterization of these enzymes has revealed several novel reactions that prompt further investigation from both biochemical and structural perspectives. The PIs long-term goal is to develop a fundamental understanding of the degradation mechanisms and to define the parameters for the substrate specificity of all the participating enzymes in the biodegradation pathway of those xenobiotic pollutants. The target of the current research is focused on the comparative investigation of three monooxygenases (TcpA, TftD and PcpB) and one special dioxygenase (PcpA), all of which catalyze the dechlorination step in polychlorophenol biodegradations. In the breakdown process of those polychlorinated aromatic compounds, dechlorination is critical because partial or complete dechlorination must occur before ring-cleavage and the subsequent mineralization. However, enzymatic dechlorination has not been significantly investigated. This investigation is providing a comprehensive understanding of the unusual specificities and reactions catalyzed by those enzymes. It is contributing to the determination of the feasibility of modification of the structures of the active sites to broaden the substrate range or accelerate the kinetics. This may lead to better clean-up strategies for remediating toxic environmental sites. Broader impactsThis research is improving the catalytic efficiency and range of substrate specificity for the biodegradative enzymes capable of bioremediation of xenobiotic pollutants. It is applying a multidisciplinary approach to the problem by combining biochemical, genetic, biophysical and mechanistic expertise. Two graduate students, including Emiliamo Sanchez, and several undergraduate students are participating in the research. High school students are also involved in the research through existing NSF-funded K-12 outreach programs at Washington State University, bringing enzymology, structural biology and environmental microbiology to the high school classroom and local Discovery Center.