人肝再生增强因子CXXC结构域与生物学活性的关系

Relationship between CXXC motif and biologic activity of hALR

目的:初步探讨人肝再生增强因子蛋白(human augmenter of liver regeneration protein,hALRp)CXXC结构域与生物学活性的关系。方法:根据酶催化反应中巯基浓度的变化,测定目的蛋白巯基氧化酶活性;采用GST-pulldown方法测定目的蛋白与GST-Na+,K+-ATPase融和蛋白体外相互作用;用MTT法检测目的蛋白促进成人肝细胞HL-7702增殖情况;探讨hALR与hALR-C65A(hALRp的CXXC结构域突变)蛋白活性差异。结果:在巯基氧化酶活性实验中,酶活性以转换数(TN)表示,hALR组TN=1.25±0.21,而hALR-C65A组TN=0,与hALR组比较差异有显著性(P<0.05)。但hALR-C65A与GST-Na+,K+-ATPase融和蛋白体外相互作用和促进肝细胞增殖活性与hALRp比较无明显变化。结论:hALRp的CXXC结构在巯基氧化酶活性方面有重要作用,但该结构改变并不影响hALRp与Na+,K+-ATPase的相互作用和促进肝细胞增殖活性。

Objective To study the relationship between CXXC motif and biologic activity of human augmenter of liver regeneration protein （hALRp）. Methods The function of CXXC motif and its relationship with bioactivity were investigated between hALRp and hALRp-C65A groups by detecting the sulfhydryl oxidase activity; the interaction of target protein with GST-Na^＋ , K^＋-ATPase fusion protein was examined by GST-Pulldown and MTT was used to study the ability of hALRp-C65A to promote HL-7702 hepatic cells proliferation in vitro. Results The turnover number （TN） of hALRp in sulfhydryl oxidase activity assay was 1. 254-0. 21, while TN was 0 of hALRp- C65A, there was significant difference between them （P〈0.05）. The interaction between GST-Na^＋ , K^＋-ATPase fusion protein and hALRp-C65A still remained. And the ability to promote HL-7702 proliferation of hALRp-C65A didn＇t have marked change when compared with hALRp. Conclusion The CXXC motif is essential to the activity of sulfhydryl oxidase. But this motif contributes less to the interaction between hALRp and Na^＋ , K^＋-ATPase and the ability to stimulate HL-7702 hepatic cell growth in vitro.