H2AX属组蛋白H2A家族成员,其磷酸化是细胞对DNA损伤做出反应的早期事件之一,在启动DNA修复过程中发挥重要功能,利用原核表达载体pGEX-4T-1构建了GST—H2AX融合蛋白表达载体,导入大肠杆菌DE3后经IPTG诱导和GST(Glutathione S—tmnsferase)磁珠纯化获得GST—H2AX融合蛋白,进一步利用GST和H2AX抗体对融合蛋白进行了验证.既建立了高效、稳定的GST蛋白纯化方法,也为进一步研究H2AX结构及生物学功能奠定了基础.
H2AX is a core histone H2A variant. Rapid H2AX phosphorylation is one of the key initiating events for DNA repair in response to DNA damage. In this study, an expression system for the glutathione S-transferase (GST)-H2AX fusion protein by using the prokaryotic expression vector pGEX-4T-lwas established. The fusion protein was expressed in E. coli (DE3) through IPTG induction, and purified using GST beads. Furthermore, the identity of the fusion protein was verified by Western blot using anti-GST and anti-H2AX antibodies. In conclusion, an efficient and stable method to purify GST-H2AX was established, which could serve as the foundation for elucidating the structure and function of H2AX.