文献类型：期刊文献

中文题名：双水相萃取与疏水层析分离基因工程人溶菌酶

英文题名：Purification of the Recombinant Human Lysozyme by Aqueous Two-Phase Extraction Coupled with Hydrophobic Interaction Chromatography

作者：张亚杰[1];夏杰[1];陆兵[1];徐殿胜[1]

机构：[1]华东理工大学生物反应器工程国家重点实验室,上海200237

年份：2008

卷号：34

期号：2

起止页码：193

中文期刊名：华东理工大学学报（自然科学版）

外文期刊名：Journal of East China University of Science and Technology

收录：CSTPCD;;EI(收录号：20081911245034);Scopus;北大核心:【北大核心2004】；CSCD:【CSCD2011_2012】；

语种：中文

中文关键词：双水相萃取;疏水层析;毕赤酵母

外文关键词：aqueous two-phase extraction; hydrophobic interaction chromatography; Pichia pastoris

摘要：采用双水相萃取与疏水层析分离纯化重组巴氏毕赤酵母表达的基因工程人溶菌酶。通过正交实验方法研究了聚合物浓度、盐浓度和pH对双水相体系萃取人溶菌酶过程的影响。结果表明:当双水相萃取的pH为4,硫酸钠、氯化钠、PEG4000的质量浓度为0.13、0.06和0.08时,人溶菌酶上相回收率达98.8%,纯化因子18.0,浓缩因子3.6。双水相萃取后选用高效疏水层析色谱纯化人溶菌酶,经苯基高取代基介质疏水层析后得到纯化因子为22.7,收率为88.4%的人溶菌酶纯化产品,电泳纯度检测为100%。
The purification of human lysozyme expressed and secreted by the recombinant yeast Pichia pastoris was studied. Extraction employing aqueous two-phase systems （ATPS） from PEG （polyethylene glycol） 4000 and sodium sulfate allowed direct processing of cell in yeast suspensions. The target protein was partially purified in the top phase while cells and cell debris were partitioned to the bottom phase of the system. The association of ATPS with hydrophobic interaction chromatography （HIC） for primary recovery of human lysozyme was evaluated. It was found that the target protein could be concentrated in the polymer phase with a purification factor of 18.0 and concentration factor of 3. 6 giving the yield of 98.8% after ATPE. In HIC, human lysozyme was obtained with enzyme recovery of up to 88.4% and a purification factor of 22.7. The purity of human lysozyme achieved 100% by SDS-PAGE analysis.