Enzymatic Studies on the Initial Steps of Anaerobic Toluene and Xylene Metabolism

本研究将探索催化芳香烃厌氧降解第一步的新型酶。最近的研究表明，甲基苯、甲苯和间二甲苯的降解的第一步是在富马酸盐的双键上加成甲基碳。苄基琥珀酸合成酶，催化甲苯加成富马酸，将作为该反应的原型进行研究。这种酶加成反应代表了一种新的激活芳烃的生化反应。此外，该活性还可以定义一类新的酶，通过迄今未知的反应机制催化碳-碳键的形成。苄基琥珀酸合酶可能在一系列不同细菌种类的厌氧矿化途径中起主要作用。这方面的研究旨在更好地理解这些不寻常的酶促反应。本项目将包括苄基琥珀酸合成酶的纯化和酶的反应机理的研究。除了描述一类新的酶外，这项研究的结果将对我们理解石油烃矿化细菌使用的厌氧途径具有重要意义。由于在燃料污染含水层的典型条件下缺乏分子氧，因此能够降解石油碳氢化合物（如甲苯和二甲苯异构体）的厌氧细菌至关重要。对这些途径的关键酶的研究可能会导致分子工具的发展，这些工具可能有助于监测相关微生物的丰度和活性。本研究将探索催化芳烃厌氧降解第一步的新型酶。这些化合物是重要的环境污染物。直到最近，人们还认为这类化合物的降解需要氧气。最近的工作表明，在降解甲苯和间二甲苯的第一步，新颖。除了描述这类新型酶外，这项研究的结果将对我们理解石油烃矿化细菌使用的厌氧途径具有重要意义。由于在燃料污染含水层的典型条件下缺乏分子氧，因此能够降解石油碳氢化合物（如甲苯和二甲苯异构体）的厌氧细菌至关重要。对这些途径的关键酶的研究可能会导致分子工具的发展，这些工具可能有助于监测相关微生物的丰度和活性，并有助于生物修复。

9723312 Spormann This research will investigate novel enzymes catalyzing the first step in anaerobic degradation of aromatic hydrocarbons. Recent work showed that the inital step in degradation of the methyl-benzenes toluene and m-xylene is the addition of the methyl carbon to the double bond of fumarate. Benzylsuccinate synthase, which catalyzes the addition of toluene to fumarate, will be studied as the prototype for this reaction. This enzymatic addition represents a novel biochemical reaction to activate aromatic hydrocarbons. Further, the activity may also define a novel class of enzymes that catalyze the formation of carbon-carbon bonds by a heretofore unknown reaction mechanism. Benzylsuccinate synthases may be of primary importance in anaerobic mineralization pathways in a range of different bacterial species. Research in this proposals directed towards gaining a better understanding of these unusual enzymatic reactions. This project will include purification of benzylsuccinate synthase and investigation of the enzyme's reaction mechanism. In addition to characterizing a novel class of enzymes, the results of this research will have important implications for our understanding of anaerobic pathways used by petroleum hydrocarbon-mineralizing bacteria. Because of the absence of molecular oxygen under typical conditions in fuel-contaminated aquifers, anaerobic bacteria capable of degrading petroleum hydrocarbons, such as toluene and the xylene isomers, are of key importance. Investigations of the key enzymes of these pathways may lead to the development of molecular tools that may assist in monitoring file abundance and activity of relevant microbes in situ. This research will investigate novel enzymes catalyzing the first step in anaerobic degradation of aromatic hydrocarbons. These compounds are significant environmental pollutants. Until recently it was thought that degradation of such compounds required oxygen. Recent work showed that the inital step in degradation of toluene and m-xylene, novel. In addition to characterizing this novel class of enzymes, the results of this research will have important implications for our understanding of anaerobic pathways used by petroleum hydrocarbon-mineralizing bacteria. Because of the absence of molecular oxygen under typical conditions in fuel-contaminated aquifers, anaerobic bacteria capable of degrading petroleum hydrocarbons, such as toluene and the xylene isomers, are of key importance. Investigations of the key enzymes of these pathways may lead to the development of molecular tools that may assist in monitoring the abundance and activity of relevant microbes in situ, and aid in bioremediation.