核心蛋白二聚体-二聚体界面处的氨基酸残基调节乙型肝炎病

StephenW

核心蛋白二聚体-二聚体界面处的氨基酸残基调节乙型肝炎病毒复制和 HBeAg 生物发生的多个步骤
Hui Liu 1 2 , Junjun Cheng 2 , Usha Viswanathan 2 , Jinhong Chang 2 , Fengmin Lu 1 , Ju-Tao Guo 2
隶属关系
隶属关系

    1
    北京大学医学部基础医学院微生物与传染病中心，北京，中国。
    2
    Baruch S. Blumberg Institute，Doylestown，宾夕法尼亚，美利坚合众国。

    PMID：34752483 DOI：10.1371/journal.ppat.1010057

抽象的

乙型肝炎病毒 (HBV) 的核心蛋白 (Cp) 将前基因组 RNA (pgRNA) 和病毒 DNA 聚合酶组装在一起，形成核衣壳，在那里进行逆转录病毒 DNA 复制。核心蛋白变构调节剂 (CpAM) 通过与 Cp 二聚体-二聚体界面处的疏水性“HAP”口袋结合，将 Cp 二聚体组装成没有 pgRNA 的异常或形态“正常”衣壳，从而抑制 HBV 复制。我们在此报告，一组 CpAM 抗性 Cp 在二聚体-二聚体界面处具有单个氨基酸取代残基，不仅破坏了 pgRNA 包装，而且还破坏了核衣壳包膜、病毒粒子感染性和共价闭合环状 (ccc) DNA 生物合成。有趣的是，这些突变也显着减少了 HBeAg 的分泌。生化分析表明，前核心蛋白（p25）背景下的CpAM抗性突变不影响信号肽酶去除N端19个氨基酸残基产生的p22水平，但显着降低了由弗林蛋白酶切割产生的p17 p22 的 C 端富含精氨酸的结构域，并以 HBeAg 的形式分泌。有趣的是，p22 以未磷酸化和磷酸化两种形式存在。虽然未磷酸化的 p22 位于膜状分泌细胞器和 HBeAg 的前体中，但细胞质和细胞核中的 p22 在 C 末端富含精氨酸的结构域中过度磷酸化，并与 Cp 相互作用以破坏衣壳组装和病毒 DNA 复制。因此，结果表明，除了核衣壳组装外，Cp 在二聚体-二聚体界面处的相互作用还通过调节核衣壳包膜和脱壳在后代病毒粒子的产生和感染中发挥重要作用。在减少的 p17 二聚体-二聚体界面处的类似相互作用似乎对其代谢稳定性和对 CpAM 抑制 HBeAg 分泌的敏感性很重要。

StephenW

Amino acid residues at core protein dimer-dimer interface modulate multiple steps of hepatitis B virus replication and HBeAg biogenesis
Hui Liu  1   2 , Junjun Cheng  2 , Usha Viswanathan  2 , Jinhong Chang  2 , Fengmin Lu  1 , Ju-Tao Guo  2
Affiliations
Affiliations

    1
    Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China.
    2
    Baruch S. Blumberg Institute, Doylestown, Pennsylvania, United States of America.

    PMID: 34752483 DOI: 10.1371/journal.ppat.1010057

Abstract

The core protein (Cp) of hepatitis B virus (HBV) assembles pregenomic RNA (pgRNA) and viral DNA polymerase to form nucleocapsids where the reverse transcriptional viral DNA replication takes place. Core protein allosteric modulators (CpAMs) inhibit HBV replication by binding to a hydrophobic "HAP" pocket at Cp dimer-dimer interfaces to misdirect the assembly of Cp dimers into aberrant or morphologically "normal" capsids devoid of pgRNA. We report herein that a panel of CpAM-resistant Cp with single amino acid substitution of residues at the dimer-dimer interface not only disrupted pgRNA packaging, but also compromised nucleocapsid envelopment, virion infectivity and covalently closed circular (ccc) DNA biosynthesis. Interestingly, these mutations also significantly reduced the secretion of HBeAg. Biochemical analysis revealed that the CpAM-resistant mutations in the context of precore protein (p25) did not affect the levels of p22 produced by signal peptidase removal of N-terminal 19 amino acid residues, but significantly reduced p17, which is produced by furin cleavage of C-terminal arginine-rich domain of p22 and secreted as HBeAg. Interestingly, p22 existed as both unphosphorylated and phosphorylated forms. While the unphosphorylated p22 is in the membranous secretary organelles and the precursor of HBeAg, p22 in the cytosol and nuclei is hyperphosphorylated at the C-terminal arginine-rich domain and interacts with Cp to disrupt capsid assembly and viral DNA replication. The results thus indicate that in addition to nucleocapsid assembly, interaction of Cp at dimer-dimer interface also plays important roles in the production and infectivity of progeny virions through modulation of nucleocapsid envelopment and uncoating. Similar interaction at reduced p17 dimer-dimer interface appears to be important for its metabolic stability and sensitivity to CpAM suppression of HBeAg secretion.

StephenW

https://pubmed.ncbi.nlm.nih.gov/34752483/