高盐环境下盐单胞菌（Halomonas）降解偶氮染料的机制研究

The azo dyes have the structural stability, which are considered as potential carcinogenic and mutagenic compounds discharged into water environment. The salt concentration of textile wastewater is generally high, and it is important to treat textile wastewater using the halophilic bacterium. There is little information about decolorization mechanism of halophilic bacterium. The present project will study the decolorization mechanism of halophilic bacterium Halomonas sp. strain GT. 1) Get the decoloration related genes and metabolites using high-through sequencing and mass spectrometry method, especially azo reductase gene, and then complete genome annotation to construct the new structure of gene networks and metabolic pathways. 2) Study the molecular mechanism of the salinity effect on the decolorization of azo dyes by Halomonas sp. strain GT using real time PCR and enzymatic activity. 3)The recombinant expression of azo reductase and characterization of the recombinant enzyme using prokaryotic expression vector. This project can deepen the understanding of the decolorization mechanism of azo dyes by halophilic bacterium, and provide new genetic resources and theoretical basis for bioremediation of azo dyes.

偶氮染料具有结构稳定性，进入水环境的染料残留具有潜在的致癌和致突变毒性。印染废水的含盐量一般都较高，利用嗜盐微生物处理印染废水具有重要的应用价值。目前对嗜盐微生物降解偶氮染料的机制研究较少，本项目拟对一株分离自印染废水活性污泥的嗜盐菌 Halomonas sp. Strain GT脱色偶氮染料的机制进行深入研究。1)通过高通量测序和质谱的方法获得嗜盐菌脱色偶氮染料的相关基因和代谢产物，尤其是偶氮还原酶基因，最终可完成全基因组注释，构建新的基因网络结构及代谢通路。2)应用实时定量PCR技术和酶学方法研究盐度影响Halomonas sp. Strain GT脱色偶氮染料的分子机制。3)利用原核表达载体对偶氮还原酶基因进行重组表达和重组酶的酶学性质研究。本项目可以深化对嗜盐菌催化偶氮染料脱色的机理认识，为偶氮染料的污染治理提供新的基因资源和理论基础。

偶氮染料具有结构稳定性，进入水环境的染料残留具有潜在的致癌和致突变毒性。印染废水的含盐量一般都较高，利用嗜盐微生物处理印染废水具有重要的应用价值。本项目以偶氮染料降解菌Halomonas sp. Strain GT为研究对象，通过三代测序技术，绘制了该菌的全基因图谱，并进行了基因注释。该菌含有编码偶氮还原酶、漆酶等降解偶氮染料的相关基因。本项目克隆并异源表达了该菌中的耐盐偶氮还原酶基因，并研究了其酶学性质。偶氮还原酶基因编码202个氨基酸，分子量21.68 KDa，偶氮还原酶活性随着盐度升高而降低。盐度通过抑制偶氮还原酶基因的表达，降低了偶氮还原酶的活性，导致脱色率降低。GC/MS和红外分析结果表明，酸性大红GR首先偶氮键断裂形成苯胺、苯二胺、1-氨基-2-萘酚-6,8-二磺酸，继续降解为二乙胺和乙酰胺。本项目可以深化对嗜盐菌催化偶氮染料脱色的机理认识，为高盐环境下降解偶氮染料提供新的基因资源和理论基础。