CAJ | 학술논문

胸苷磷酸化酶在核苷类物质合成中具有重要作用,本研究以短乳杆菌为胸苷磷酸化酶生产菌种,对短乳杆菌产胸苷磷酸化酶发酵培养基进行优化.首先通过 Plackett-Burman 设计筛选出影响短乳杆菌产胸苷磷酸化酶的3个较为重要因素:发酵时间(P=0.030)、接种量(P=0.033)和葡萄糖浓度(P=0.019).在此基础上采用最陡爬坡路径逼近最大响应区域,并利用响应面中心组合设计对影响显著因素进行优化,得到最适培养基组成成分和培养条件为:发酵初始pH 8.0,葡萄糖18 g/L,酵母膏15 g/L,NaCl 7.5 g/L,蛋白胨10 g/L,胸苷15 mmol/L,摇床转速110 r/min,发酵温度38℃,发酵时间10.57 h,接种量1.54%.在此优化条件下,短乳杆菌产胸苷磷酸化酶能力得到了很大提高,短乳杆菌胸苷磷酸化酶活从0.400 U/mg湿菌体提高到1.172 U/mg湿菌体,比优化前提高了2.93倍.蛋白质凝胶电泳分析显示经优化后每克湿菌体胸苷磷酸化酶的含量明显高于优化前.
흉감린산화매재핵감류물질합성중구유중요작용,본연구이단유간균위흉감린산화매생산균충,대단유간균산흉감린산화매발효배양기진행우화.수선통과 Plackett-Burman 설계사선출영향단유간균산흉감린산화매적3개교위중요인소:발효시간(P=0.030)、접충량(P=0.033)화포도당농도(P=0.019).재차기출상채용최두파파로경핍근최대향응구역,병이용향응면중심조합설계대영향현저인소진행우화,득도최괄배양기조성성분화배양조건위:발효초시pH 8.0,포도당18 g/L,효모고15 g/L,NaCl 7.5 g/L,단백동10 g/L,흉감15 mmol/L,요상전속110 r/min,발효온도38℃,발효시간10.57 h,접충량1.54%.재차우화조건하,단유간균산흉감린산화매능력득도료흔대제고,단유간균흉감린산화매활종0.400 U/mg습균체제고도1.172 U/mg습균체,비우화전제고료2.93배.단백질응효전영분석현시경우화후매극습균체흉감린산화매적함량명현고우우화전.
Thymidine phosphorylase plays a critical role in synthesis of nucleosides. In this study,fermentation medium of Lactobacillus brevis for producing thymidine phosphorylase was optimized. Firstly,three factors of fermentation time (P=0.032),inoculum size (P=0.037) and glucose concentrations (P=0.022) significantly affected thymidine phosphorylase were identified by Plackett-Burman experiment. Then,the three factors were further optimized through steepest ascent path approaching the maximum response region and response surface central composite design (CCD). The optimal fermentation medium were as follows :initial pH 8.0,glucose 18 g/L,yeast extract 15 g/L,NaCl 7.5 g/L,peptone 10 g/L,thymidine 15mmol/L. Culture conditions were as follows :rotating speed 110r/min,temperature 38 ℃,fermentation time 10.57 h,inoculum size 1.54%. Under optimum conditions, the capacity of Lactobacillus brevis to produce thymidine phosphorylase was greatly improved,thymidine phosphorylase activity reached 1.172 U/mg wet bacteria which was increased by 2.93 times more than that before optimization. Protein gel electrophoresis showed that thymidine phosphorylase content per gram wet bacteria with optimized fermentation medium was obviously higher than that before optimization.