Ⅱ型抗癌晶体蛋白抗肝癌作用的关键芳香族氨基酸  

Key aromatic amino acids of anti-hepatoma activity on Parasporin-2

作　　者：廖利民[1] 林淑芳[1] 田凌[1] 陈爱明[1] 林毅[1] 

机构地区：[1]华侨大学化工学院生物工程与技术系,福建厦门361021

出　　处：《生物工程学报》2013年第6期823-835,共13页Chinese Journal of Biotechnology

基　　金：国家自然科学基金(No.40601046);教育部科学技术研究重点项目(No.211205);福建省高等学校新世纪优秀人才支持计划(2006年度);福建省自然科学基金(No.2011J01221)资助~~

摘　　要：为探讨Ⅱ型抗癌晶体蛋白(Parasporin-2)上与抗肝癌作用相关的关键氨基酸,利用5-BU对Parasporin-2活性区编码DNA(P2Y)进行PCR诱变,之后在大肠杆菌中表达,产物纯化后经MTT法检测其对肝癌细胞系和正常肝细胞系的作用。获得的9个突变体其抗肝癌活性差异极大,其中P2M1和P2M8对两种肝癌细胞系SMMC7721和Bel7402均有较强的细胞毒杀作用而不影响正常肝细胞系Chang-liver。比较了P2M1、P2M8和P2Y的二级结构与三级结构,发现二级结构上的变化如β折叠变长或α螺旋增加影响着Ⅱ型抗癌晶体蛋白的抗肝癌活性。基于突变体间氨基酸序列比对、突变体与受体间分子对接以及模拟突变等研究的结果表明,位点52、56、58和208上的氨基酸残基特别是芳香族氨基酸在Parasporin-2与受体间的互作中可能起着重要作用。Nine mutants （P2M1-9） were obtained using PCR with 5-BU based on DNA template （P2Y） encoding the activeregion of Parasporin-2. Mutant proteins were purified after expressing in E. coli BL21 cells, followed by assayed against hepatoma cells and normal liver cells by MTT. They showed diverse anti-hepatoma activities, in which two mutant proteins, P2M1 and P2M8, exhibited high cytotoxicity against hepatoma cell lines SMMC7721 and Be17402, meanwhile leaving normal liver cells Chang-liver unaffected. Structural comparison among P2Y, P2M1 and P2M8 showed that the length of 13-sheet or 13-fold, and the amount of ct helix greatly affected the anti-hepatoma activity of Parasporin-2. Results based on amino acid alignment, molecular docking between P2Y, P2M1 or P2M8 and receptor, and mimic mutation demonstrated that amino acid residues at the sites of 52, 56, 58 and 208 on P2Y, especially the aromatic amino acids such as Trp, Phe, and Tyr were involved in the interactions.

关 键 词：Ⅱ型抗癌晶体蛋白 突变体 抗肝癌 芳香族氨基酸 

分 类 号：R735.7[医药卫生—肿瘤]