Thermomonospora amylolytica来源嗜碱β-葡萄糖苷酶的克隆、异源表达及其酶学性质研究

Cloning,heterologous expression and characterization of a basophilic β-glucosidase from Thermomonospora amylolytica

β-葡萄糖苷酶能催化水解纤维二糖为葡萄糖,在医药、农业、食品、饲料等领域有着非常广泛的应用。本研究从嗜热放线菌(Thermomonospora amylolytica)中获得一个碱性β-葡萄糖苷酶基因,命名为tabg12a。该基因在大肠杆菌中成功进行克隆和异源表达,并对其酶学性质进行研究。结果表明:tabg12a的最适反应p H和温度分别为8.0和45℃,其在p H 8.0~9.6下保持50%以上相对活性,在p H为4.0~10.0缓冲液孵育24 h后仍保持80%以上的相对酶活,在30℃下孵育2 h仍保持70%以上的相对酶活,在35℃下孵育1 h仍保持50%以上的相对酶活。1 mmo/L的K+和Ca2+对其表现出促进作用,提高至120%;0.1%的SDS和1 mmo/L Cu^(2+)下仍保持100%的相对活性。以上酶学性质预示着该酶在食品、饲料和纤维素乙醇生产等领域具有潜在利用价值。

β-glucosidase can catalyze the hydrolysis of cellobiose to glucose,which has been widely used in medicine,agriculture,food,feed and other fields.Here,a basophilicβ-glucosidase gene(denominated tabg 12a)was obtained from Thermomonospora amylolytica.The gene was successfully cloned and heterologous expressed in E.coli,and its enzymatic properties were characterized.The results showed that tabg12a exhibited optimal activity at pH 8.0 and 45℃,respectively.It maintained more than 50%relative activity at pH 8.0~9.6,more than 80%relative enzyme activity after 24 h incubation in pH 4.0~10.0,more than 70%relative enzyme activity after incubation at 30℃ for 2 h,and more than 50%relative enzyme activity after incubation at 35℃ for 1 h.1 mmol/L of K^(+) and Ca^(2+)could promote its activity to 120%.The relative activity remained 100% in the presence of 0.1%SDS and 1 mmol/L Cu^(2+).These properties indicated that tabg I2a had potential applications in food,feed and cellulosic ethanol production etc.