酵母双杂交技术筛选人PIF1解螺旋酶相互作用蛋白  

Screening interaction proteins of human PIF1helicase by yeast-two-hybridization

作　　者：王攀[1,2] 王建校[2] 李珊珊[2] 刘晓丹 王豫[1] 周平坤[1] 顾永清[1,2] 

机构地区：[1]军事医学科学院放射与辐射医学研究所,北京100850 [2]石河子大学医学院,新疆石河子832003

出　　处：《西安交通大学学报（医学版）》2013年第5期595-598,共4页Journal of Xi’an Jiaotong University（Medical Sciences）

基　　金：国家自然科学基金资助项目(No.30960093;31160183;31270894)~~

摘　　要：目的 应用酵母双杂交技术研究筛选与人类PIF1解螺旋酶相互作用的蛋白。方法 应用酵母双杂交技术，以人PIF1蛋白PINT功能域（1～180氨基酸）和解螺旋酶模序（167～926氨基酸）为诱饵，与HeLa细胞cDNA文库杂交，筛选能与人PIF1蛋白不同功能域相互作用的蛋白。结合生物信息学分析，一对一酵母回复性杂交及β-galactosidase实验等确定阳性克隆。结果 PIF1蛋白PINT功能域杂交共获得17个阳性克隆，生物信息学分析有3个阳性基因，它们分别是CCNDBP1、OTUD5、CAP1。而PIF1蛋白解螺旋酶模序未获得阳性克隆。结论 PIF1蛋白的PINT功能域对调控PIF1的生理功能具有非常重要的作用。Objective To screen interaction proteins of human PIF1 helicase by yeast-two-hybridization. Methods The proteins interacting with different domains of human PIF1 were screened by using PINT domain （1-180 amino acids） and helicase motifs （167-641 amino acids） of PIF1 helicase as baits respectively to hybridize a HeLa cDNA library. The positive clones were confirmed by one-to-one yeast-two-hybridization, β-galactosidase assay and bioinformatic analysis. Results Totally 17 positive clones interacting with PINT domain of PIF1 were obtained, and 3 positive genes, namely, CCNDBP1, OTUD5 and CAP1, were verified by bioinformatic analysis. However, no positive clone interacting with helicase motifs of PIF1 was obtained. Conclusion The PINT domain of human PIF1 plays a very important role in regulating physiological functions of PIF1 helicase.

关 键 词：人PIF1解螺旋酶 PINT功能域 酵母双杂交 蛋白质相互作用 

分 类 号：Q78[生物学—分子生物学]