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proEMAPⅡ(又称p43蛋白)是哺乳动物氨酰tRNA合成酶的辅助因子,近年来发现,p43具有细胞因子活性以及抗新生血管生成的作用,具有潜在的抗肿瘤疗效.p43的C端结构域即为EMAPⅡ,其结构和功能都已知,具有细胞因子和抗血管生成的双重功能.但是N端的结构还不清楚,研究表明全长的p43比EMAPⅡ具有更高的生物学活性,但是p43的结构和作用机制尚不明确.此外,作为蛋白质药物,更小的分子能够减少免疫原性和副作用,从而发挥更好的活性.本文利用生物信息学方法,对p43 N端的二级结构进行预测,并且在不破坏二级结构的前提下,构建了10个p43的缺失突变体,在体外对10个缺失突变体与全长的p43蛋白进行抗新生血管生成活性验证比较,最终我们获得了3个活性高的p43缺失突变体,并且发现N端的1~16,1~79位氨基酸和C端的264~312位氨基酸不是p43发挥抗血管生成功能所必需的,而且它们的删除使得活性位点更好地呈现并发挥活性.通过我们的研究,有助于揭示p43蛋白结构和功能的关系以及其作用机制,同时为临床筛选肿瘤治疗候选药物奠定基础.
ProEMAPⅡ (aka p43 protein) is a cofactor for mammalian aminoacyl-tRNA synthetase. In recent years, it has been found that p43 has cytotoxic activity and anti-angiogenic activity and has potential anti-tumor effect. The C-terminal domain ofp43 Is known as EMAPII, its structure and function are known, with dual functions of cytokines and anti-angiogenesis.But the N-terminal structure is unclear, the study showed that full-length p43 than EMAPII has higher biological activity, but the structure of p43 And the mechanism of action is not clear.In addition, as a protein drug, smaller molecules can reduce the immunogenicity and side effects, and thus play a better activity.This paper uses bioinformatics method to predict the secondary structure of p43 N-terminal, and Without disrupting the secondary structure, 10 p43 deletion mutants were constructed, and 10 deletion mutants were compared with the full-length p43 protein in anti-angiogenic activity verification. Finally, we obtained 3 actives High p43 deletion mutant and found that amino acids 1 to 16 and 1 to 79 at the N terminal and amino acids 264 to 312 at the C terminal are not necessary for p43 to exert antiangiogenic function and their deletion Was active site appear better and play activities. Through our research will help reveal the relationship between structure and function of the p43 protein and its mechanism of action, while laying the foundation for the clinical treatment of cancer screening drug candidates.