Hepatitis B virus (HBV) core protein assembles viral pre-genomic (pg) RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs) and sulfamoylbenzamides (SBAs), have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or “empty” capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc) DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B.

乙型肝炎病毒（HBV）核心蛋白将病毒前基因组（pg）RNA和DNA聚合酶组装成核衣壳，以进行逆转录DNA复制。已发现小分子的几种化学型，包括杂芳基二氢嘧啶（HAP）和氨磺酰基苯甲酰胺（SBA），可以变构地调节核心蛋白的结构，从而改变核心蛋白组装的动力学和途径，导致形成不规则形状的核心蛋白聚集体或“空的衣壳，不含前基因组RNA和病毒DNA聚合酶。有趣的是，除了抑制核壳装配和随后的病毒基因组复制外，我们现在还证明了HAP和SBA差异性调节了从头开始共价闭合环状（ccc）DNA的生物合成。诱导病毒颗粒的核衣壳以及细胞质中含双链DNA的子代核衣壳的解体，从而感染和胞内扩增途径。具体而言，错误地提示衣壳脱壳的提示可防止从头开始形成cccDNA。感染肝细胞，同时从细胞质子代核衣壳中瞬时加速cccDNA的合成。我们的研究表明，正链DNA的延伸会诱导核衣壳的结构变化，从而赋予成熟的核衣壳结合CpAM的能力并触发其拆卸。了解核心蛋白变构调节剂对核衣壳组装和拆卸的双重作用的分子机制将有助于发现靶向核心蛋白的新型抗病毒剂，该抗病毒剂可以更有效地抑制cccDNA的合成并治愈慢性乙型肝炎。