文献类型：期刊文献

中文题名：原癌蛋白MDM2分离纯化及结晶条件的初步筛选

英文题名：Purification and optimization of crystallization conditions of oncogenic protein MDM2

作者：王玥[1,2];齐心洁[1,2];郭红霞[1,2];刘玥[1,2];胡贤达[3];黄迎春[1,2]

第一作者：王玥

机构：[1]北京联合大学生物化学工程学院;[2]生物质废弃物资源化利用北京市重点实验室;[3]中国藏学研究中心北京藏医院

第一机构：北京联合大学生物化学工程学院

年份：2016

卷号：36

期号：6

起止页码：41-44

中文期刊名：中国生化药物杂志

外文期刊名：Chinese Journal of Biochemical and Pharmaceutics

语种：中文

中文关键词：MDM2;原核表达;分离纯化;结晶

外文关键词：MDM2; prokaryotic expression; purification; crystallization;

摘要：目的探讨原癌蛋白MDM2分离纯化的条件,获得高纯度的目的蛋白,筛选该蛋白的结晶条件,获得MDM2蛋白结晶。方法利用大肠杆菌表达系统,诱导表达MDM2蛋白,采用Ni-NTA亲和柱和分子筛对该蛋白进行分离纯化,通过圆二色光谱法对其二级结构进行分析,采用坐滴法对结晶条件进行初步的筛选。结果获得了高纯度的MDM2蛋白,其二级结构有序,在低pH条件下有形成晶体结构的倾向。结论 MDM2蛋白结晶的最适pH为5.5,最适的MDM2蛋白质浓度为10 mg/m L。
Objective To investigate and optimize the condition of purification and crystallization of oncogenic protein MDM2. Methods MDM2 was expressed in E. coli expression system,and purified by Ni-NTA chelating affinity chromatography and molecular sieve chromatography. The secondary structure of purified protein was analyzed by circular dichroism spectroscopy（ CD）. Then the crystallization condition of MDM2 was screened and optimized by sitting-drop vapor-diffusion method. Results High purity of MDM2 was obtained by Ni-NTA chelating affinity chromatography and molecular sieve chromatography purification. CD analysis indicated the secondary structure of MDM2 was ordered. Protein-crystallisation experiments illustrated that MDM2 was prone to crystallization under lower pH. Conclusion The optimum pH of MDM2 protein crystallization is 5. 5,the optimum protein concentration is 10 mg /mL.