CAJ | 학술논문

目的 ADS-J1是通过虚拟筛选从20 000个化合物中获得的靶向HIV gp41的小分子HIV进入抑制剂.该研究探讨ADS-J1与gp41的结合位点和作用机制.方法采用酸性天然聚丙烯酰胺凝胶电泳技术(AN-PAGE),研究ADS-J1与衍生于gp41不同功能区的多肽的结合.结果此前采用的天然聚丙烯酰胺凝胶电泳(N-PAGE)等方法,由于不能显示衍生于gp41的N-端多肽,未能确定ADS-J1的作用位点.该研究建立的AN-PAGE技术,能直接显示N-端多肽的条带,证实ADS-J1能与gp41的N-端螺旋重复序列(NHR)复合螺旋核中的深穴结合,从而抑制gp41六螺旋束结构的形成,而且深穴中第574位的赖氨酸残基(K574)与ADS-J1的抑制作用密切相关.结论 ADS-J1通过与gp41 NHR靶穴中的K574结合,抑制gp41六螺旋束结构的形成,从而抑制HIV进入靶细胞.此外,该研究建立的AN-PAGE技术,为研究靶向Ⅰ型包膜病毒的病毒进入抑制剂的作用机制提供了一个简便有效的实验方法.
목적 ADS-J1시통과허의사선종20 000개화합물중획득적파향HIV gp41적소분자HIV진입억제제.해연구탐토ADS-J1여gp41적결합위점화작용궤제.방법 채용산성천연취병희선알응효전영기술(AN-PAGE),연구ADS-J1여연생우gp41불동공능구적다태적결합.결과 차전채용적천연취병희선알응효전영(N-PAGE)등방법,유우불능현시연생우gp41적N-단다태,미능학정ADS-J1적작용위점.해연구건립적AN-PAGE기술,능직접현시N-단다태적조대,증실ADS-J1능여gp41적N-단라선중복서렬(NHR)복합라선핵중적심혈결합,종이억제gp41륙라선속결구적형성,이차심혈중제574위적뢰안산잔기(K574)여ADS-J1적억제작용밀절상관.결론 ADS-J1통과여gp41 NHR파혈중적K574결합,억제gp41륙라선속결구적형성,종이억제HIV진입파세포.차외,해연구건립적AN-PAGE기술,위연구파향Ⅰ형포막병독적병독진입억제제적작용궤제제공료일개간편유효적실험방법.
Aim ADS-J1 is a low molecular HIV entry inhibitor targeting HIV transmembrane subunit gp41 through virtual screening from a compound library containing 20 000 molecules.This study is to investigate the binding sites of ADS-J1 on gp41.Methods Acid native polyacrylamide gel electrophoresis (AN-PAGE) assay was applied to test the binding ability of ADS-J1 with the peptides derived from gp41 N-terminal heptad repeat (NHR) region.Results It was reported previously that ADS-J1 could block the gp41 six-helix bundle (6-HB) formation using native polyacrylamide gel electrophoresis (N-PAGE).However,the binding sites could not be found because positive charged N-peptides derived from gp41 NHR could not show bands on the gel.In the present study,the AN-PAGE assay which could show N-peptides in the gel was established,and it was found that ADS-J1 could inhibit the gp41 6-HB formation.Moreover,ADS-J1 bound directly to the gp41 cavity region of NHR.The positively charged residue (K574) located in this region was critical for the binding of ADS-J1.Conclusions ADS-J1 inhibits HIV entry by targeting the cavity region of gp41 NHR,whereas K574 in the cavity plays a critical role for the binding.Furthermore,the AN-PAGE assay provides a simple method for studying the mechanism of action of virus entry inhibitors targeting the transmembrane protein of type I enveloped virus.