一株海洋假单胞菌降解有机卤代物特性及酶学性质

Degradation of organic halogenated compounds by a marine Pseudomonas sp.strain and its enzymatic properties

以有机卤代物的生物降解为导向,从海洋中分离筛选有机卤代物降解菌株56,根据其形态、生理生化特征及16S rRNA序列鉴定其种属,考察其以卤代乙酸、卤代丙酸、卤代丁酸和卤代酰胺等15种有机卤代物作为唯一碳源时的生长特性、降解能力及其细胞中卤代酸脱卤酶催化特性。结果表明:菌株56为昆明假单胞菌(Pseudomonas kunmingensis),可以耐受体积分数为1%~5%的甲醇、乙醇、丙酮与乙腈。该菌株能够以除2-氟丙酸之外的14种有机卤代物作为唯一碳源。其中对单溴乙酸、2-溴丙酸、2-溴-2-甲基丙酸和二氯乙酸降解能力较高,96 h内对初始摩尔浓度为10 mmol/L的卤代酸去除率为96%~100%。该菌株中脱卤酶可以催化多种有机卤代物的脱卤,对2-氯丙酸的脱卤效果最好,其催化D-2-氯丙酸与L-2-氯丙酸脱氯的最适反应温度均为40℃,最适pH均为9.0;脱卤酶在低于40℃环境中,具有良好的热稳定性,随着温度的升高,酶活性迅速降低;pH在5.0~9.0,酶活性比较稳定,当pH<5.0以及pH>9.0时,酶活性降低。综上,该菌株具有应用于降解有机卤代物的潜力。

To explore the biodegradation of organic halogenated compounds,strain 56 was isolated from marine environment.Strain 56 was identified based on its morphology,physiological and biochemical characteristics and 16S rRNA sequence analysis.The growth characteristics and degradation ability were investigated by culturing strain 56 with fifteen organic halogenated compounds including haloacetic acid,halopropionic acid,halobutyric acid and haloamides as the sole carbon source.Furthermore,dehalogenase produced by strain 56 was characterized.Strain 56 was identified as Pseudomonas kunmingensis and showed tolerance against methonal,ethanol,acetone and acetonitrile with volume fraction from 1%to 5%.Fifteen halogenated compounds could be utilized as sole carbon source by Pseudomonas kunmingensis except 2-fluopropionic acid.High degradation ability was observed when 10 mmol/L monobromoacetic acid,2-bromopropionic acid,2-bromo-2-methylpropionic acid and dichloroacetic acid acted as the carbon source and 96%to 100%of them were removed within 96 h.The dehalogenase could act on many halogenated acids,among which 2-chloropropionic acid was the best substrate.The optimal temperature and pH of catalyzing D-and L-2-chlorpropionic acid were 40℃and 9.0,respectively.Excellent thermal stability was observed as the temperature was below 40℃,and the enzyme activity decreased rapidly with the increase of temperature.High enzyme stability was observed from pH 5.0 to pH 9.0,while the enzyme activity decreased when pH<5.0 and pH>9.0.In conclusion,Pseudomonas kunmingensis has the potential to be applied in the biodegradation of organic halogenated compounds.