天花粉蛋白的定点突变及聚乙二醇修饰-生物探索.PDFVIP

2008 3 21 3 Chin J Biologicals March 2008, Vo 1. 21 No. 3 # 194 # Q78 A 2008 1 2 1 1 1 1 1 1 1 2 安群星 雷迎峰 穆士杰 张献清 陈蕤 夏爱军 陈晨 易静 吴原茹 徐志凯 (Trichosanthin,T CS) ( PEG) , DNA TCS ( YFF8183 KR173174) , (TCSYFF8183ACS TCSKR173174CG) , 82 173 , PEG TCS , PEG T CS DNA TCS , , 30 000, PEG , 40 000 , TCS , PEG TCS DNA TCS PEG , TCS ; ; ; Sitedirected Mutation of Trichosanthin and Chemica Modification of Mutant by Po yethy ene G yco AN Qunxing, LEI Yingf ng, MU Shiji , t al ( Dep artment of Blood Transf usion, X ij ing H osp ital, Fourth Military Med i cal University , Xi ! an 710032, China) Abstract Objective To induc sit dir ct d mutation of trichosanthin (T CS) , modify th mutant by poly thyl n glycol ( PEG) and analyz its DNas lik activity. Methods Two pot ntial antig nic d t rminants of TCS, YFF8183 and KR173174, w r s l ct d by comput r mod ling , bas d on which th g n s ncoding two sit dir ct d mutants of TCS, TCSYFF8183ACS and TCSKR173174CG, w r amplifi d by PCR, ins rt d into pRSETA v ctor and transform d to E coli DH5. Th r combinant plasmid was xtract d, id ntifi d by r striction analysis and s qu ncing and transform d to E coli BL21( DE3) for xpr ssion und r induction of IPTG. Th xpr ss d product was id ntifi d by SDSPAGE and W st rn blot and modifi d by PEG. Th DNas lik activiti s of th two PEGmodifi d mutants w r analyz d and compar d with thos of unmodifi d mutants. Resu ts Both r striction analysis and s qu ncing r sult show d that r combinant plasmids w r construct d corr ctly.Th r lativ mol cular mass s of th xpr ss d products b for and aft r modification w r about 30 000 and about 40 000 r sp ctiv ly .Lik unmod ifi d TCS mutants, th two mutants modifi d by PEG show d DNas lik