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幽门螺旋杆菌中的肿瘤坏死因子α诱导蛋白(Tipα)被鉴定为幽门螺旋杆菌致病感染中的新型致癌因子.Tipα通过NF-κB的激活诱导肿瘤坏死因子α(TNF-α)的大量表达,从而促使宿主的炎症反应以及肿瘤发生、发展的进程.Tipα的同源二聚体为其发挥生物学功能的活性形式,此二聚体以两个单体间N端半胱氨酸形成的二硫键(Cys25-Cys25与Cys27-Cys27)共价连接.Tipα(25-192)的基因克隆至载体pET22b中,并且在大肠杆菌菌株BL21(DE3)中以可溶形式高水平表达.重组蛋白经过Ni2+金属亲和层析、阳离子交换层析和凝胶阻滞层析进行分离纯化.Tipα蛋白样品分别通过悬滴和microbatch的方法进行结晶搜索和优化.母体和硒代晶体分别衍射到2.2和2.6,均属于C2空间群,并且具有相似的晶胞参数.母体晶体的晶胞参数为a=127.01,b=47.57,c=96.5,α=γ=90°,β=127.5°.
Tumor necrosis factor alpha-induced protein (Tipα) in Helicobacter pylori has been identified as a novel oncogenic factor in the pathogenic infection of Helicobacter pylori.Tipα induces the massive expression of tumor necrosis factor α (TNF-α) through the activation of NF-κB, Thereby promoting the host’s inflammatory response and tumorigenesis and progression.Tipα homodimers for its biological activity of the active form, the dimer to two monomers N-terminal cysteine form of bis (Cys25-Cys25 and Cys27-Cys27) .Tipα (25-192) was cloned into vector pET22b and expressed in soluble form at high level in E. coli strain BL21 (DE3) Ni2 + metal affinity chromatography, cation exchange chromatography and gel retardation chromatography were used for the isolation and purification of the proteins.Tipα protein samples were respectively subjected to crystal search and optimization by hanging drop and microbatch methods.The precursors and selenium crystals were diffracted to 2.2 and 2.6 all belong to the C2 space group and have similar unit cell parameters.The unit cell parameters of the parent crystal are a = 127.01, b = 47.57, c = 96.5, α = γ = 90 °, β = 127.5 ° .