背景:结合适当的细胞受体是任何病毒生命周期的关键步骤。在未来的药物设计项目开始之前,必须确定病毒表面蛋白的受体结合域的结构。
目的:研究pH诱导的具有功能缺失突变的衣壳肽二级结构的变化可以为进入机制提供一些启示。
方法:光谱法伴随着电泳,超滤,和计算生物化学。
结果:在这项研究中,我们表明,在0.01M磷酸盐缓冲液中,细小病毒B19VP1衣壳(残基13-31)的受体结合域的肽在pH=7.4时是β结构,但根据圆二色性(CD)光谱结果,在pH=5.0时α-螺旋。红外(IR)光谱的结果表明,在pH=7.4和pH=5.0的部分脱水条件下,相同的肽以α-螺旋和β-结构构象存在。相比之下,具有Y20W突变的肽,已知可以阻断病毒的内化,在pH=7.4的部分脱水条件下,主要形成α-螺旋构象。根据我们的假设,在pH=7.4时,由野生型肽在其四聚体中形成的分子间反平行β结构是由细小病毒B19受体结合域的相应部分与其细胞受体(AXL)形成的类似分子间反平行β结构的原型.
结论:VP1衣壳蛋白中Y20W取代功能的丧失通过α螺旋稳定和其转变为无序状态的能力的降低防止了向β结构状态的转变。
BACKGROUND: Binding appropriate cellular receptors is a crucial step of a lifecycle for any virus. Structure of receptor-binding domain for a viral surface protein has to be determined before the start of future drug design projects.
OBJECTIVE: Investigation of pH-induced changes in the secondary structure for a capsid peptide with loss of function mutation can shed some light on the mechanism of entrance.
METHODS: Spectroscopic methods were accompanied by electrophoresis, ultrafiltration, and computational biochemistry.
RESULTS: In this
study, we showed that a peptide from the receptor-binding domain of Parvovirus B19 VP1 capsid (residues 13-31) is beta-structural at pH=7.4 in 0.01 M phosphate buffer, but alpha- helical at pH=5.0, according to the circular dichroism (CD) spectroscopy results. Results of infra- red (IR) spectroscopy showed that the same peptide exists in both alpha-helical and beta-structural conformations in partial dehydration conditions both at pH=7.4 and pH=5.0. In contrast, the peptide with Y20W mutation, which is known to block the internalization of the virus, forms mostly alpha-helical conformation in partial dehydration conditions at pH=7.4. According to our hypothesis, an intermolecular antiparallel beta structure formed by the wild-type peptide in its tetramers at pH=7.4 is the prototype of the similar intermolecular antiparallel beta structure formed by the corresponding part of Parvovirus B19 receptor-binding domain with its cellular receptor (AXL).
CONCLUSIONS: Loss of function Y20W substitution in VP1 capsid protein prevents the shift into the beta-structural state by the way of alpha helix stabilization and the decrease of its ability to turn into the disordered state.