乙酰短杆菌中核苷磷酸化酶的性质研究  

Study of the Properties of Nucleoside Phosphorylase in Brevibacterium Acetylicum

作　　者：张春艳 许彦梅 王玥 丁庆豹[2] 欧伶[2] 

机构地区：[1]上海斯贝生物科技有限公司,上海200235 [2]华东理工大学生物反应器重点实验室,上海200237

出　　处：《现代生物医学进展》2013年第23期4419-4423,共5页Progress in Modern Biomedicine

基　　金：上海市宝山区产学研合作专项基金项目(CXY-2010-28)

摘　　要：以乙酰短杆菌完整细胞为酶源,研究不同条件下核苷磷酸化酶的性质。方法:将乙酰短杆菌湿菌体置于不同保藏温度及在不同种类缓冲溶液中考察其稳定性;在有或无核保护下核苷磷酸化酶对热的稳定性;并设计核苷的磷酸解反应或合成反应,测定核苷磷酸化酶的活力及酶促反应的表观米氏常数。结果:乙酰短杆菌中的核苷磷酸化酶经低温保藏可以保持较长时间的稳定性;菌体在60℃处理1小时即失去核苷磷酸化酶的活力,但是添加胸腺嘧啶有明显的保护作用;菌体中核苷磷酸化酶的合成能力明显大于磷酸解能力;对尿苷和5-甲基尿苷的表观米氏常数和最大反应速率分别为16.7、11.4 mmol/L,0.0063、0.0041 mmol/L.min。结论:含核苷磷酸化酶的乙酰短杆菌完整细胞作为酶源,在低温下可以长时间保藏,反应中的碱基对核苷磷酸化酶的抗热性有益,该菌种可以作为工业上核苷磷酸化酶的来源。Objective： The properties of nucleoside phosphorylase in the intact Brevibacterium acetylicum cells were studied tra- der different conditions. Methods： The stability of nucleoside phosphorylase was inspected when wet cells of Brevibacterium acetylicum were stored in different temperature or in different buffered solution. The thermal stability of enzyme was studied with or without protec- tion. The phosphorolysis or synthesis reaction was designed to determine activity of nucleoside phosphorylase and apparent Michaelis-Menten constants. Results： Nucleoside phosphorylase in Brevibacteriurn acetylicum could keep stability when the wet cells were stored long in low temperature. Activity of nucleoside phosphorylase in cells was lost after heated at 60~C for 1 hour, however, the introduction of thymine and analogues could provide some protection. The ability of synthesis was obviously higher than that of phospho- rolysis. The apparent Michaelis-Menton constant and Maximal reaction rate to uridine and 5-methyluridine were 16.7, 11.4 mmol/L; and 0.0063, 0.0041 mmoFL.min respectively. Conclusion： Wet cells of Brevibacterium acetylicum could be stored long in low temperature. Some base in the reaction could provide some protection to nucleoside phosphorylase. Brevibacterium acetylicum could be used as the resource ofnucleoside phosphorylase in industry.

关 键 词：核苷磷酸化酶 乙酰短杆菌 酶学性质 5-甲基尿苷 尿苷 

分 类 号：Q814[生物学—生物工程] Q939.97