GST融合蛋白在杆状病毒系统中表达的可溶性研究

Solubilization Analysis of GST Fusion Protein Expressed in Baculovirus System

将构建好的可表达GST融合蛋白的重组病毒AcMNPV OCC- GST 6xHis Etp2 8感染Sf9细胞 ,一定时间后取感染了病毒的细胞裂解物上清液进行SDS PAGE分析 ,结果显示 5 3kDa的融合蛋白 (GST 6xHis Etp2 8)呈不溶状态。在原有裂解液的基础上 ,加固体十二烷基肌氨酸钠至终浓度 1.5 % ,并将TritonX 10 0的比例由 1%提高到 2 %。SDS PAGE结果显示至少有 1/ 3的GST 6xHis Etp2 8处于溶解状态 ,可溶性GST 6xHis Etp2 8经亲合层析 ,5

Insect Sf 9 cells were infected with constructed recombinant virus AcMNPV OCC - GST 6xHis Etp 28, which could produce GST fusion protein and the supernatant of 72 h pi cell lysate was examined by SDS PAGE. The results showed that GST 6xHis Etp 28 fusion protein of 53 kDa was expressed in insoluble status. After the sodium salt of the alkylanionic detergent sarkosyl was added to insect cell lysis buffer to a final concentration of 1.5%, and the final concentration of TritonX 100 was increased from 1% to 2%, at least 1/3 of GST 6xHis Etp 28 appeared to be soluble, which was examined by SDS PAGE. Then soluble GST 6xHis Etp 28 was purified by affinity chromatography using glutathione agarose.