OsLEA5c在大肠杆菌中的表达、纯化及对蛋白质的保护效应研究  

Expression and Purification of OsLEA5c in Escherichia coli and Its Protective Effect on Proteins

作　　者：胡廷章[1] 陈再刚[1] 秦娟[1] 黄小云[1] 

机构地区：[1]重庆三峡学院生命科学与工程学院,重庆404100

出　　处：《河南农业科学》2015年第12期11-14,共4页Journal of Henan Agricultural Sciences

基　　金：重庆市万州区科学技术项目(201403063)

摘　　要：为了研究OsLEA5c蛋白对其他蛋白的保护效应,在大肠杆菌DL21中异源表达OsLEA5c蛋白,采用HiTrap^(TM) chelating HP层析柱纯化OsLEA5c蛋白。纯化的OsLEA5c蛋白用于高温、脱水和冻融胁迫下对乳酸脱氢酶的保护效应研究。结果表明,在PBS缓冲体系中,50℃处理10、20、30 min后,乳酸脱氢酶的活性分别残留43.20%、24.53%、17.55%;而按1∶1的质量比加入纯化的OsLEA5c蛋白后,乳酸脱氢酶活性分别保留85.77%、74.24%、65.27%。干燥脱水5 h后,乳酸脱氢酶活性仅保留2.55%~4.19%;按2∶1、5∶1和10∶1的质量比加入OsLEA5c蛋白,乳酸脱氢酶活性分别保留17.59%、24.73%、36.39%。反复冻融5次,乳酸脱氢酶活性残留9.41%~12.14%;而按2∶1、5∶1和10∶1的质量比加入纯化的OsLEA5c蛋白,乳酸脱氢酶活性分别保留28.09%、69.44%、95.16%。表明OsLEA5c蛋白对其他蛋白质具有保护作用,能减少高温、脱水和冻融胁迫对蛋白质的损伤。In order to investigate protective effect of OsLEA5c protein on other proteins,OsLEA5c protein was heterologously expressed in Escherichia coli DL21,and purified using HiTrap^（TM） chelating HP column.The purified OsLEA5c protein was used to study the protective effect of the OsLEA5c protein on lactate dehydrogenase（ LDH） activity under heating,drying,and freeze-thaw stresses. LDH was incubated in PBS buffer at 50 ℃ for 10,20,30 min,its activity was decreased to 43. 20%,24. 53% and 17. 55%,respectively. However,when OsLEA5c protein was added into LDH solution at a mass ratio of 1∶ 1（ OsLEA5c∶LDH）,enzyme activity remained at 85. 77%,74. 24% and 65. 27%,respectively,after treatment in the same way. After air-drying for 5 h,the residual activity of LDH were 2. 55% —4. 19%; while OsLEA5c was added into LDH solution at mass ratios of 2 ∶ 1,5 ∶ 1 and 10 ∶ 1,its residual activity were 17. 59%,24. 73% and 36. 39%,respectively. After 5 cycles of freeze-thaw treatments,the activity of residual LDH was decreased to 9. 41% —12. 14%,while OsLEA5c was added to LDH solution at mass ratios of 2 ∶ 1,5∶ 1 and 10 ∶ 1,its residual activity were 28. 09%,69. 44% and 95. 16 %,respectively,after freeze-thaw treatments. Therefore,the result suggested that OsLEA5c protein could protect other protein and decrease damage from heating,drying,and freeze-thaw stresses.

关 键 词：胚胎发育后期丰富蛋白 OsLEA5c蛋白 乳酸脱氢酶 蛋白质活性 

分 类 号：Q512[生物学—生物化学]