NIH迁移是酶催化苯环上羟化反应时一种分子内的重排现象，多种芳烃代谢过程中伴随着NIH迁移，但对其机理研究甚少。侧孢短芽孢杆菌PHB-7a通过伴有NIH迁移的龙胆酸途径将4-羟基苯甲酸彻底降解，但该代谢途径在任何微生物菌株中无遗传和生化水平的研究报道。本研究拟以侧孢短芽孢杆菌PHB-7a为研究对象，通过基因组测序、转座子插入突变等方法获得完整的4-羟基苯甲酸代谢基因簇，结合基因敲除互补、体外表达、酶活测定和产物鉴定等方法鉴定其生理学和生化功能；纯化代谢途径中NIH迁移的关键酶，进行酶学特性研究。本研究将首次从分子生物学、生化和遗传学水平上阐明微生物代谢4-羟基苯甲酸时伴随羧基NIH迁移的龙胆酸途径，并揭示NIH迁移的分子机理，进一步阐明微生物分解代谢的多样性及其适应性进化策略。

An NIH shift is a chemical rearrangement where an atom (or a group) on an aromatic ring undergoes an intramolecular migration primarily during a hydroxylation reaction catalyzed by an enzyme. However, the mechanism study of this interesting observation is very limited. Brevibacillus laterosporus PHB-7a degrades 4-hydroxybenzoate via gentisate pathway involving an NIH shift. But to date, neither the 4-hydroxybenzoate catabolic genes nor the corresponding enzymes have been identified in any microbial strains. In this proposed study, the complete 4-hydroxybenzoate catabolic gene cluster in strain PHB-7a will be identified by genome sequencing together with transposon mutagenesis. The physiological and biochemical functions of catabolic genes involved in this pathway will be characterized through gene knockout and complementation, expression in vitro, product identification and enzyme assays. The functions and properties of the key enzymes involved in the NIH shift will be investigated in great details. The findings from this study will hopefully reveal, for the first time, the pathway for 4-hydroxybenzoate catabolism via gentisate involving an NIH shift at molecular, biochemical, and genetic levels. In particular, the mechanism of NIH shift will also be elucidated. It is also of great help to explore the catabolic diversity for the microbial aromatic degradation and illustrate the adaptive evolution strategies of microbes in their catabolism.