海藻酸钠裂解酶高产菌株的筛选、鉴定及产酶条件的优化

Screening and Identification of Alginate Lyase-producing Strains andOptimization of Its Enzyme-production Conditions

以海藻酸钠为唯一碳源培养基,对广西北海涠洲岛采集的马尾藻进行微生物的筛选和分离,采用3,5-二硝基水杨酸法(DNS)进行海藻酸钠裂解酶相对酶活力测定,获得高产酶菌株M0101;依据形态、生理生化特征及16S rDNA序列分析对其进行鉴定,该菌属于弧菌属,命名为Vibrio gangliei M0101;通过单因素实验确定了菌株M0101的产酶优化方案:海藻酸钠50 g/L,(NH_(4))_(2)SO_(4)2 g/L,K_(2)HPO_(4)1 g/L,NaCl 100 g/L,MgSO_(4)·7H_(2)O 1 g/L,FeSO_(4)·7H_(2)O 0.01 g/L,培养温度30℃,初始pH9.0,200 r/min;优化后菌株M0101产酶最大活性可达221.90 U/mL,相较于优化前最大酶活36.32 U/mL,优化后酶活力是优化前的6.11倍。表明M0101能高效水解高浓度海藻酸钠,是强耐盐的菌株,具有大规模制备海藻酸钠裂解酶的潜力。

Using sodium alginate as the sole carbon source medium,the microorganisms in Sargassum collected from Weizhou Island,Guangxi,were screened and isolated.The relative enzyme activity was determined by 3,5-Dinitrosalicylic acid(DNS)method,and the high-yield strain,marked M0101,was obtained.According to the morphological,physiological and biochemical characteristics and 16S rDNA sequence analysis,the target strain was identified to belong to the genus of Vibrio,named as Vibrio gangliei M0101.The prioritization scheme of enzyme production of M0101 was confirmed by a single factor test:Alginate 50 g/L,(NH_(4))_(2)SO_(4)2 g/L,K2HPO41 g/L,NaCl 100 g/L,MgSO_(4)·7H_(2)O 1 g/L,FeSO_(4)·7H_(2)O 0.01 g/L,30℃,200 r/min,initial pH9.0;and the maximum enzyme activity of strain M0101 reached 221.90 U/mL after optimation.It was 6.11 times higher than before(36.32 U/mL).Its efficiently hydrolysis of high concentration alginate and high salt tolerance,which has potential for large-scale preparation of alginate lyase.