Isolation,identification and enzymatic properties of the strain producing β-mannosidase
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摘要: 本研究通过初筛（魔芋粉为唯一碳源并结合刚果红染色法）和复筛（利于对硝基苯基-β-D-吡喃甘露糖苷法测定β-甘露糖苷酶活力）,从采集的森林土壤中筛选产β-甘露糖苷酶的菌株,获得1株酶高产菌株B19。通过显微形态、革兰氏染色、生理生化特征、16S r DNA序列及系统发育进化树分析等方法,将其鉴定为肠杆菌属（Enterobacter sp.）。菌株B19所产的β-甘露糖苷酶为胞内酶,在37℃、200 r/min条件下培养48 h后,测得的酶活力为1.26 U/m L。初步酶学性质研究表明:该酶的最适反应p H和最适反应温度分别为7.5和50℃,且在p H6.08.0和温度4550℃稳定性较高,浓度为10 mmol/L的Mn2+和Mg2+对该β-甘露糖苷酶具有激活作用,但K+、Fe2+、Ca2+、Cu2+、Co2+及Zn2+抑制酶活力。该菌株可作为产β-甘露糖苷酶的潜在菌株。Abstract: The strain B19 with high yield of β- mannosidase was isolated from forest soil sample using first screening( Konjac powder as the sole carbon source and Congo red staining method) and secondary screening( determination of β-mannosidase enzyme activity by p-Nitrophenyl-β-D-Mannopyranoside method).The strain was identified as Enterobacter sp. by electron microscopy,Gram- staining,physiological and biochemical examination,16 S r DNA sequence and phylogenetic tree analysis. The strain B19 could produce intracellularβ- mannosidase,and its activity could be up to 1.26 U / m L when the strain was cultivated 48 h under the condition of 37 ℃,200 r / min.The enzymatic properties analysis showed that the optimal p H and temperature of this enzyme were 7.5 and 50 ℃,respectively.The enzyme exhibited high thermal stability when the temperature ranged from 45 to 50 ℃,or the p H value ranged from 6.0 to 8.0. The enzyme activity was activated by Mg2 +and Mn2 +in concentration of 10 mmol / L,but was inhibited by K+,Fe2 +,Ca2 +,Cu2 +,Co2 +and Zn2 +. It might be a good candidate strain in studying β- mannosidase.
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Keywords:
- β-mannosidase /
- isolation /
- identification /
- Enterobacter sp /
- enzymatic properties
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