【作者】 陈香君；

【导师】 邱广亮；

【作者基本信息】 内蒙古师范大学 ， 植物学， 2008， 硕士

【摘要】 从烟气生物脱硫系统的好氧产硫磁性稳态流化床反应器中取磁性多孔珠固定化生物膜,采用排硫硫杆菌培养基在好氧条件下富集、反复分离纯化获得分离菌,经过细菌、菌落形态的观察以及一系列生理生化指标,对应伯杰氏细菌鉴定手册,初步鉴定该分离菌为硫杆菌属（thiobacillus）的排硫硫杆菌（thiobacillus thioparus）。将分离纯化得到的排硫硫杆菌细胞超声波破碎、脱盐浓缩、DEAE-52离子交换层析、Sephadex G-150分子排阻层析分离纯化得到亚硫酸盐受体氧化还原酶,并对其进行性质表征。光谱分析表明该酶除了在280nm外无其他特征吸收峰;SDS-PAGE结果显示该酶是一种αβ异二聚体,由分子量33.9KDa和42.2KDa的两个亚基构成;活力分析发现有92%的酶活存在于细胞膜部分,该酶属于膜蛋白。其最适pH为7.5,最适温度为40℃,在3-7.5较宽的pH范围内酶活力具有很好的稳定性,温度在3℃-50℃之间酶活力能保持在70%以上。该酶对亚硫酸钠的Vmax为22.15μM/min,Km为86.09μM;底物专一性强,该酶基本不受Na⁺、K⁺、Mg²⁺、Fe²⁺、Fe³⁺、Ca²⁺、Cu²⁺、Zn²⁺、Mn²⁺和Ba²⁺等金属离子的影响,EDTA和氢氧化钠对该酶有明显的抑制作用。其氧化产物为硫酸盐。对可能参与分离菌硫氧化代谢途径中的酶采用酶活分析方法进行了测定,并参考相关文献,提出了分离菌硫氧化代谢途径的可能模型。更多还原

【Abstract】 The biofilm immobilized on the magnetic porous beads obtained from the aerobic magnetically stabilized bed (aMSFB) was enriched cultured under the aerobic condition by using culture medium for thiobacillus thioparus, and isolated and purified repeatedly to get the isolated bacteria. The isolated bacteria was identified as thiobacillus thioparus of thiobacillus according to《Bergey’s Manual of Systemaic Bacteriology》through the observe of the bacteria and the morphology of the bacteria clone and a series of physiological and biochemical index.The isolated and purified thiobacillus thioparus bacteria were broken by using ultrasonic,and then desalted condensed. sulfite acceptor oxidoreductase was obtained through isolation and purification by using DEAE-52 ion-exchange chromatography and Sephadex G-150 molecular-exclusion chromatography and was characterized for its properties. The spectra analysis indicated that the enzyme had no other characteristic adsorption peak except in 280nm. SDS-PAGE showed that the enzyme was aαβheterodimer constituted by two subunits with molecular weights 33.9kDa and 42.2 kDa. Activity analysis indicated that 92% of enzyme existed in the cell membrane and the enzyme was a membrane protein. The optimum pH of the enzyme was 7.5, the optimum temperature of the enzyme was 40℃and the enzyme activity had good stability and the enzyme activity could maintain up to 70% within the temperature range of 3℃-50℃. The Vmax and Km of the enzyme to sodium sulfite as substrate were 22.15μM/min and 86.09μM respectively. The enzyme had high substrate specificity and could not be influenced by metal ions. EDTA and NaOH had obvious inhibition to the enzyme and the oxidative product was sulphate.According to the latest study of sulphur oxidative bacteria, the cytoplasm extracts and the membrane extracts of the isolated bacteria were fetched and the activity of the key enzymes in the physiological and biochemical metabolism was determined to judge the location of the enzymes. The possible sulphur metabolic path in thiobacillus thioparus was conferred through the analysis of the reactive paths catalyzed by different enzymes and the reference to the relative literature.The sulphur oxidative system model of thiobacillus thioparus was put forward based on the above facts.更多还原