The Evolutionary Origin and Potential of Newly Recruited Enzymes

描述（由申请人提供）：酶的进化是地球上生命出现的关键因素，它随后的分化占据了广泛的生态位。大多数酶都具有令人印象深刻的催化能力，经过38亿年的进化。我们对新酶进化的早期阶段很感兴趣。该应用程序解决了最近在Sphingobium chlorophenolicum（一种从PCP严重污染的土壤中分离出来的细菌）中降解五氯酚（PCP）途径中发挥新功能的酶。S. chlorophenolicum似乎利用至少两个先前存在的途径中的酶将一个功能不良的降解PCP的途径拼凑在一起。这个实验系统为我们在这个过程的早期阶段提供了一个新的代谢途径的进化窗口。本应用程序侧重于途径中限制通过PCP降解途径的通量的初始三种酶。PCP羟化酶催化PCP羟基化生成有毒中间体四氯苯醌（TCBQ）。TCBQ还原酶催化TCBQ还原为四氯对苯二酚（TCHQ）。TCHQ脱卤酶催化两个连续的还原脱卤反应，去除氯，使环被切割。我们的目标是：1)获得PCP羟化酶和TCHQ脱卤酶的结构；2)探索TCBQ还原酶的进化起源和可能的附加功能；3)进化出这三种酶的改良版本；4)确定改良酶如何影响细菌降解PCP的能力。这项工作的结果将是更好地了解新酶可以产生的来源，以及在新酶进化的初始阶段实现的催化质量。公共卫生相关性：我们将更好地了解降解有毒污染物途径的进化前景，以及通过招募解毒抗生素的酶与抗生素耐药性的发展相关的问题。此外，这项工作将为设计酶和细菌以生产化学品和药品或在无害环境条件下降解污染物的努力提供信息。

DESCRIPTION (provided by applicant): The evolution of enzymes was a critical element of the emergence of life on Earth and its subsequent divergence to occupy a vast range of ecological niches. Most enzymes have impressive catalytic capabilities that have been honed over 3.8 billion years of evolution. We are interested in the early stages of evolution of new enzymes. This application addresses enzymes that appear to have been recently recruited to serve new functions in the pathway for degradation of pentachlorophenol (PCP) in Sphingobium chlorophenolicum, a bacterium isolated from soil heavily contaminated with PCP. S. chlorophenolicum appears to have patched together a poorly functioning pathway for degradation of PCP using enzymes from at least two previously existing pathways. This experimental system provides us with a window on the evolution of a new metabolic pathway at a very early stage in the process. This application focuses on the initial three enzymes in the pathway that limit the flux through the PCP degradation pathway. PCP hydroxylase catalyzes the hydroxylation of PCP to form the toxic intermediate, tetrachlorobenzoquinone (TCBQ). TCBQ reductase catalyzes the reduction of TCBQ to tetrachlorohydroquinone (TCHQ). TCHQ dehalogenase catalyzes two successive reductive dehalogenation reactions that remove chlorines, allowing the ring to be cleaved. Our aims are 1) to obtain structures of PCP hydroxylase and TCHQ dehalogenase; 2) to explore the evolutionary origin and a possible additional function of TCBQ reductase; 3) to evolve improved versions of all three enzymes; and 4) to determine how the improved enzymes affect the ability of the bacterium to degrade PCP. The outcome of this work will be a better understanding of the sources from which new enzymes can arise, and the quality of catalysis achieved during the initial stages of evolution of a new enzyme. PUBLIC HEALTH RELEVANCE: We will better understand the prospects for evolution of pathways to degrade toxic pollutants and the problems associated with development of antibiotic resistance by recruitment of enzymes that detoxify antibiotics. In addition, this work will inform efforts to engineer enzymes and bacteria to produce chemicals and pharmaceuticals or to degrade pollutants under environmentally benign conditions.