伯克霍尔德菌IDO3降解吲哚途径及关键基因解析

Indole is a common N-heterocyclic aromatic compound, which plays an important role in the microbial physiological and biochemical properties, and human diseases and health. High concentrations of indole have certain toxicity to organisms and should endanger the ecological environment and human health. Indole biotransformation study is an important part to clarify its biological significance in a complex system, and it is also the key for indole pollution remediation. Although the study of indole transformation has lasted for nearly a century, the bacterial resources for indole degradation are still lacking, and the degradation pathways and molecular mechanisms remain unclear. In this project, Burkholderia sp. IDO3, a novel and highly efficient indole-degrading bacterium previously obtained in our laboratory, is used as the research object. The degradation product and chemical degradation pathway of indole in strain IDO3 will be determined in detail. Then the functional genes (clusters) involved in indole metabolism are mapped by genome and transcriptome sequencing. Finally, the functional genes responsible for indole oxidation, isatin hydrolysis and anthranilic acid metabolism will be elucidated in-depth by gene mutation, heterologous expression and enzymatic property studies. This project aims to reveal the degradation pathway and key functional genes of indole degradation in Burkholderia strains, which will perfect the indole molecular metabolic mechanism and promote its biological function and remediation studies.

吲哚是一种常见的氮杂环芳香化合物，在微生物生理生化及人类疾病与健康等方面扮演着重要的角色，而高浓度的吲哚对生物具有一定毒性，危害生态环境和人类健康。吲哚的生物转化研究是阐明其在复杂体系中生物学意义的重要组成部分，也是吲哚污染修复的关键。虽然吲哚转化研究已持续近百年，但仍存在降解菌株资源缺乏、降解途径和分子机制不明晰等研究现状。本项目以实验室前期获得的高效吲哚降解菌Burkholderia sp. IDO3为研究对象，将详细解析菌株IDO3降解吲哚的中间产物，推测其降解途径。进一步通过对菌株进行基因组和转录组测序分析，定位参与吲哚代谢的关键功能基因(簇)。利用基因突变、异源表达和酶学特性研究阐明菌株IDO3中负责吲哚加氧、靛红水解和邻氨基苯甲酸代谢的功能基因及其特性。本项目旨在揭示Burkholderia菌属降解吲哚的途径及关键基因，补充和完善吲哚分子代谢机制，促进其生物学功能及修复研究。

吲哚是一种常见的氮杂环芳香化合物，在人体及环境中广泛存在。目前关于吲哚的研究大多集中于信号分子作用研究，吲哚转化研究虽也开展较早，但降解分子机制尚不明晰。本项目以实验室前期筛选到的Burkholderia sp. IDO3为研究对象，通过条件优化确定该菌具有较强的吲哚降解能力，利用高效液相色谱质谱联用鉴定靛红和邻氨基苯甲酸为重要中间产物；通过基因组克隆文库技术找到一个含吲哚加氧酶的基因簇iif1，该酶能催化吲哚合成靛蓝，在进化上较为独立；进一步通过全基因组测序和比较基因组分析发现菌株IDO3含有第二个吲哚加氧酶基因簇iif2，通过基因异源表达、基因敲除等实验揭示iif2是菌株IDO3降解吲哚的关键基因簇；在iif2基因簇中找到一个新颖的基因iifF，通过基因表达和纯酶分析，证实该基因编码靛红水解酶，催化靛红生成靛红酸，首次为吲哚降解的靛红途径提供分子证据；最后我们还发现菌株IDO3能催化吲哚衍生物—粪臭素的降解，优化了降解性能，并通过投加活性污泥和高通量测序实验，证实该菌具有生物强化应用潜力，能长期定殖于活性污泥中。本项目系统揭示了革兰氏阴性菌降解吲哚的分子机制，促进了吲哚的生物转化和环境行为研究。