乙型肝炎病毒衣壳蛋白二聚体的内含子界面的完整性调节衣

StephenW

The Integrity of the Intradimer Interface of the Hepatitis B Virus Capsid Protein Dimer Regulates Capsid Self-Assembly
Zhongchao Zhao, Joseph Che-Yen Wang, Carolina Perez Segura, Jodi A Hadden-Perilla, Adam Zlotnick

    PMID: 32459465 DOI: 10.1021/acschembio.0c00277

Abstract

During the hepatitis B virus lifecycle, 120 copies of homodimeric capsid protein assemble around a copy of reverse transcriptase and viral RNA and go on to produce an infectious virion. Assembly needs to be tightly regulated by protein conformational change to ensure symmetry, fidelity and reproducibility. Here we show that structures at the intradimer interface regulate conformational changes at the distal interdimer interface and so regulate assembly. A pair of interacting charged residues, D78 from each monomer, conspicuously located at the top of a four-helix bundle that forms the intradimer interface, were mutated to serine to disrupt communication between the two monomers. The mutation slowed assembly and destabilized dimer to thermal and chemical denaturation. Mutant dimers showed evidence of transient partial unfolding based on appearance of new proteolytically-sensitive sites. Though mutant dimer was less stable, the resulting capsids were as stable as wildtype, based on assembly and thermal denaturation studies. Cryo-EM image reconstructions of capsid indicated that the subunits adopted an "open" state more usually associated with free dimer and that the spike tips were either disordered or highly flexible. Molecular dynamics simulations provide mechanistic explanations for these results, suggesting that D78 stabilizes helix 4a, which forms part of the intradimer interface, by capping its N-terminus and hydrogen-bonding to nearby residues, whereas the D78S mutation disrupts these interactions, leading to partial unwinding of helix 4a. This in turn weakens the connection from helix 4 and the intradimer interface to helix 5, which forms the interdimer interface. .

StephenW

乙型肝炎病毒衣壳蛋白二聚体的内含子界面的完整性调节衣壳的自组装。
赵忠超，王若-，卡罗来纳州的佩雷斯·塞古拉，乔迪·阿·哈登·佩雷利亚，亚当·兹洛特尼克

    PMID：32459465 DOI：10.1021 / acschembio.0c00277

抽象

在乙型肝炎病毒的生命周期中，120个拷贝的同二聚体衣壳蛋白围绕一个拷贝的逆转录酶和病毒RNA聚集，并继续产生感染性病毒体。组装需要通过蛋白质构象变化严格控制，以确保对称性，保真度和可重复性。在这里，我们显示了二聚体内界面的结构调节了二聚体远端界面的构象变化，从而调节了组装。来自每个单体的一对相互作用的带电残基D78，显着地位于形成二聚体界面的四螺旋束的顶部，被突变为丝氨酸以破坏两个单体之间的通讯。该突变减慢了组装，并使二聚体不稳定，从而热和化学变性。突变的二聚体显示出基于新的蛋白水解敏感位点出现的瞬时部分展开的证据。尽管突变二聚体的稳定性较差，但根据组装和热变性研究，所得衣壳与野生型一样稳定。衣壳的冷冻-EM图像重建表明，亚基采用“开放”状态，通常与游离二聚体相关，并且尖峰尖端无序或高度灵活。分子动力学模拟为这些结果提供了机理解释，表明D78通过封闭其N端并与附近残基氢键合来稳定构成内二聚体界面一部分的螺旋4a，而D78S突变破坏了这些相互作用，导致部分相互作用。展开螺旋线4a。这进而削弱了从螺旋4和二聚体内界面到螺旋5的连接，后者形成了二聚体界面。 。