热稳定性高β-甘露聚糖酶产生菌的筛选、鉴定及酶学性质研究

为开发耐高温、热稳定性高的β-甘露聚糖酶,以魔芋胶为唯一碳源,采用透明圈法,从土壤中筛选产β-甘露聚糖酶的菌株;通过16S rDNA序列及分子发育树分析对菌株进行鉴定;采用DNS法测定β-甘露聚糖酶活性并对酶学性质进行研究。结果表明,该菌能够水解魔芋胶,水解圈D/d比值平均为1.67;鉴定该菌为地衣芽孢杆菌(Bacillus licheniformis)KD-1;该菌所产β-甘露聚糖酶最适pH6.0,最适反应温度60℃;在pH5.0~9.0和60~80℃,酶的稳定性良好,60、70和80℃酶的半衰期(T1/2)分别为5.5、4.3和4.2 h;10 mmol/L的Cu2+和Mg2+明显促进β-甘露聚糖酶活性,而Mn2+明显抑制酶活性。本研究筛选到一株地衣芽孢杆菌KD-1,其所产β-甘露聚糖酶,高温下(如80℃)热稳定性高于目前报道的β-甘露聚糖酶。

Isolation,Identification and Enzyme Properties of a β-Mannanase Producing Strain with High Thermal Stability

To develop a thermostable β-mannanase,a high β-mannanase producing strain was isolated from soil based on the sole carbon source cultivation and clearing zone screening on konjac powder plates. The strain was identified by 16S rDNA phylogenetic analysis. The β-mannanase activity and the enzyme properties were assayed by DNS method. The results showed that the strain could hydrolyze konjac powder and the clearing zone D/d value was about 1.67. The strain was assigned to Bacillus licheniformis KD-1.The optimal pH of the enzyme was pH6.0 and the optimal temperature was 60℃. The β-mannanase was stable at pH5.0~9.0 and 60~80℃. The half-life time(T1/2)of the enzyme activity was 5.5,4.3 and 4.2 h at 60,70 and 80℃. The β-mannanase activity was promoted by 10 mmol/L Cu2+ and Mg2+,whereas the enzyme activity was inhibited by 10 mmol/L Mn2+. In this study,a strain of Bacillus licheniformis KD-1 was screened. The thermostability of β-mannanase produced by KD-1 was higher than that of β-mannanase reported at present.