核苷水解酶是一类催化核苷生成D-核糖及嘌呤或嘧啶的酶,广泛存在于除哺乳动物以外的生物中。该研究表达并纯化了3个来源于谷氨酸棒杆菌Corynebacterium glutamicum ATCC 13032的核苷水解酶Cgl1364、Cgl1977和Cgl2835并考察其酶学特性。酶学实验结果表明,重组Cgl1364、Cgl1977和Cgl2835均具有核苷水解酶活性;能够催化腺苷、胞苷、鸟苷、黄苷、肌苷和尿苷生成D-核糖和相应的嘌呤或嘧啶,但底物特异性有所差异;重组酶在15~55 ℃以及pH 4~10条件下均具有活性,其最适温度和pH分别为35 ℃和7.0;Cgl1977的热稳定性优于Cgl1364和Cgl2835。利用BL-cgl1364静息细胞为酶源催化水解尿苷,底物转化率达到89.5%。该研究首次报道了谷氨酸棒杆菌核苷水解酶的功能和性质,可为D-核糖、嘌呤及嘧啶的“一锅法”生物合成提供重要参考。
The nucleoside hydrolase is the enzyme that catalyzes nucleoside to D-ribose as well as purine or pyrimidine. Nucleoside hydrolases have been widely found in organisms except mammals. In this study, three nucleoside hydrolases (Cgl1364, Cgl1977, and Cgl2835) from Corynebacterium glutamicum ATCC 13032 were expressed and purified. The three recombinant enzymes all exhibited activities of nucleoside hydrolases, which could catalyze adenosine, cytidine, guanosine, xanthoside, inosine and uridine to D-ribose and correspond purine or pyrimidine. However, the specificity to substrates of the recombinant enzymes were different. These enzymes exhibited activities at 15-55 ℃ and pH 4-10, and the optimum temperature was 35 ℃ and the optimum pH was 7.0. Moreover, the thermal stability of Cgl1977 was higher than that of Cgl1364 and Cgl2835. The BL-cgl2835 resting cells were used as the enzyme source to hydrolyze uridine, resulting in the yield of 89.5%.This is the first work reported the function and characteristics of nucleoside hydrolases from C. glutamicum, which will provide reference for biosynthesis of D-ribose, purine, and pyrimidine.
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