小肽在原核和真核细胞中抑制α-synuclein异常折叠的研究  

Study of Screening Method for Inhibiting the Abnormal α-synuclein Aggregation with Small Peptides

作　　者：王俊卿[1] 付烈[1] 周稳[1] 熊平[1] 汪浩勇[1] 

机构地区：[1]湖北工业大学湖北省工业微生物重点实验室,湖北武汉430068

出　　处：《生物技术》2011年第3期29-33,共5页Biotechnology

基　　金：国家自然科学基金项目("促进α-synuclein正确折叠多肽的筛选";30870798)资助

摘　　要：目的:研究在大肠杆菌中能够有效抑制α-synuclein异常折叠的小肽,在人神经母细胞瘤SH-SY5Y中的抑制作用。方法:基于a-Syn疏水区域设计的5种透膜小肽在大肠杆菌中能够有效抑制α-synuclein异常折叠;构建真核表达质粒CMV-SynGFP-IRES-Pn并转入SH-SY5Y细胞,通过测定细胞荧光强度分析小肽抑制α-synuclein异常折叠情况,由此评价大肠杆菌筛选系统确定的小肽在真核SH-SY5Y细胞中的有效性。结果:在大肠杆菌中能有效抑制α-Syn(129A)异常折叠的三种小肽R7P1、R7P3、R7P4,在SH-SY5Y中的作用效果也十分明显,分别比对照肽ASI1D提高了114%、98%和98%。对于α-Syn(53T),R7P1和R7P2在SH-SY5Y中分别比对照提高了90%和44%;然而在两种细胞中,5种透膜小肽对野生型α-Syn(WT)的作用与对照相比差距不明显。Objective： Small peptides screened from E. coli can inhibit the abnormal α- synuclein aggregation in neuroblastoma SH - SY5Y cells. Method： Five membrane permeational peptides RTP1 - R7P5 were designed based on hydrophobic regions; Eukaryotic expression vector CMV -SynGFP-IRES-Pn were transfected into SH -SY5Y cell,the inhibition of α-synuclein from abnormal aggregation of the small peptide was measured by cell fluorescence strength, to evaluate the effectiveness of E. coli screening system. Result： The small peptides （ RTPI , RTP3, RTP4） can inhibit α- Syn （S129A） abnormal folding in E. coli, also have significant effect in SH - SY5Y. Compared with ASI1D, R7P1, R7P3, R7P4 were 114% ,98% and 98% more effective in inhibiting abnormal aggregation of α- Syn （S129A） respectively. RTP1 and R7P2 were 90% and 44% more effective in inhibiting abnormal an＆negation of α- Syn （A53T） respectively. However, the difference between the effectiveness of five small peptides and ASI1D for the wildtype α-Syn （WT） is not obvious.

关 键 词：Α-SYNUCLEIN 透膜小肽 SH—SY5Y 相对荧光强度 帕金森病 

分 类 号：Q74[生物学—分子生物学]