【摘要】 目的探索并鉴定β型血小板源性生长因子受体(platelet-derived growth facter recepterβ,PDGFRβ)与PDZ蛋白β2-syntrophin新的结合作用以及该相互作用的分子基础,为进一步研究PDGFRβ相关信号转导通路的调节提供线索。方法制备GST-PDGFRβ羧基端的融合蛋白,通过GST pull-down技术,检测PDGFRβ-CT与β2-syntrophin的PDZ结构域及完整的β2-syntrophin蛋白的相互作用。结果成功构建了重组质粒pGEX-PDGFRβ-CT。在大肠埃希菌中成功表达融合蛋白后,通过体外蛋白质结合实验证实了PDGFRβ-CT与β2-syntrophin的PDZ结构域和兔脑组织中内源性的β2-syntrophin蛋白可以相互作用。结论PDGFRβ-CT与β2-syntrophin之间存在相互作用,该相互作用是由PDGFRβ的羧基末端与β2-syntrophin的PDZ结构域结合而实现的,此结果为研究PDGFRβ介导的下游信号转导通路以及细胞特定调控受体功能的机制奠定了基础。更多还原

【Abstract】 Objective The aim of this study is to identify the interaction between the PDZ domain-containing protein β2-syntrophin and platelet-derived growth factor receptor-β (PDGFRβ), to further explore the underlying mechanism of PDGFRβ/β2-syntrophin association.Methods The carboxyl-terminal of PDGFRβ (PDGFRβ-CT) was fused to GST protein by gene cloning. The interactions between PDGFRβ-CT and PDZ-domain β2-syntrophin or full-lengh β2-syntrophin were characterized by GST pull-down assay. Results PDGFRβ-CT was cloned into pGEX-2TK and GST-fusion protein was over-expressed and purified in E.coli system. GST pull-down experiment revealed that carboxyl-terminal of PDGFRβ robustly pulled down PDZ domain of β2-syntrophin. The interaction with endogenous β2-syntrophin was confirmed in the native rabbit brain tissue.Conclusion PDGFRβ specifically interacts with β2-syntrophin through the carboxyl-terminal of PDGFRβ and the PDZ domain on β2-syntrophin,which may modulate PDGFRβ downstream signaling pathway.更多还原