利用蛋白质组学技术筛选与BIV基质蛋白MA相互作用蛋白

Identification of Cellular Protein Associated with Bovine Immunodeficiency Virus Matrix Protein by Proteomics Technology

牛免疫缺陷病毒BIV的基质蛋白MA是由gag基因编码病毒结构蛋白之一,参与病毒运输包装等多种过程.通过蛋白质组学方法,寻找与MA有相互作用的宿主蛋白.通过原核表达MA-CBP融合蛋白,将其结合在几丁质上,并与细胞裂解物混合,纯化特异结合的细胞蛋白.利用双向电泳以及MALDI-TOF质谱分析,筛选到8个与MA具有相互作用的候选蛋白,功能涉及运输、氧化还原、热激反应等.进而克隆所鉴定蛋白之一的PRDX4,利用体外结合实验进一步证实其与BIV MA蛋白存在相互作用.初步建立利用蛋白质组学技术研究病毒与宿主相互作用实验方法,所得结果为了解慢病毒结构蛋白在病毒生活周期中功能提供了线索,有助于探索过氧化物酶家族蛋白在病毒感染过程中的作用及其分子机制.

Matrix protein of Bovine immunodeficiency virus is one of the proteins encoded by gag gene among the structural proteins.We employed a proteomic approach in search of MA partners in cell.Cellular extracts were applied to MA-CBP or CBP immobilized on chitin resin and total bound cellular proteins were resolved by 2-DE.Analyses of interacting proteins by matrix-assisted laser desorption/ionization-time of flight mass spectrometry allowed identification of eight cellular proteins binding to BIV MA protein,which involved in various cellular pathways including transport,peroxide reduction,and heat shock response.Subsequently, PRDX4,one protein identified were cloned and its interaction with BIV MA were confirmed through binding assay in vitro.The present studies pave the way for elucidation of lentivirus structural protein regulation,as well as PRDXs function in virus life cycle,and may help provide a further understanding of interaction between virus and host.