Structure And Assembly Of The Hepatitis B Nucleocapsid

背景：乙肝病毒感染是一个世界性的生物医学问题，对病毒的组装和结构的了解有助于开发新的抗病毒药物，并提供关于复杂大分子结构的基本信息。乙肝病毒核心基因编码前C蛋白(Pre-C)，它要么被部分加工形成一种被称为e抗原(HBeAg)的分泌型非颗粒蛋白(HBeAg)，要么被完全加工以产生核心抗原(HBcAg)。HBcAg是一种183个残基的蛋白质，包裹在RNA-逆转录酶复合体(乙肝病毒聚合酶)周围。如果HBcAg在细菌细胞质中组装成含有结合宿主核酸的二十面体衣壳，那么它就在大肠杆菌中得到了表达。多碱C-末端34个残基(鱼精蛋白结构域)的缺失也产生具有组装能力的蛋白。C端截短蛋白(Cp149：残基1-149)的衣壳不含核酸，其结构由低温电子显微镜、图像分析和X射线结晶学确定。天然HBeAg也在第149位被C-末端截断，此外还含有来自前-C部分处理的10个残基N-末端延伸。虽然HBeAg的功能和结构尚不清楚，但它是一种重要的血清学标志。结果：利用表面等离子共振(Biacore)，一组抗乙肝病毒核衣壳蛋白的单抗(MAbs)的运动亲和图显示了一系列的结合亲和力。用冰冻电子显微镜(A.Steven)对部分HBV型核衣壳抗体复合物进行了进一步表征。结果揭示了比之前描述的更多的不连续表位。这些发现有助于解释组装的HBcAg和未组装的HBeAg抗原之间的免疫学区别。对HBeAg的结构研究正在继续使用具有改进的物理特征的突变体。对大的乙肝病毒聚合酶蛋白的研究也在取得进展。在一种增量方法中，我们表达了聚合物的各种功能结构域，这些结构域将受到结晶轨迹的影响。摘要：乙肝病毒是世界范围内导致癌症的主要原因。虽然有疫苗可用，但慢性乙肝通常是在儿童时期感染的。病毒核衣壳在病毒的生命周期中起着重要的结构作用和代谢作用。了解乙肝病毒核衣壳的分子结构将有助于有针对性地发现药物，以防止病毒的组装和形成。

Background: Hepatitis B Virus (HBV) infection is a worldwide biomedical problem and an improved understanding of the assembly and structure of the virus may help develop new antiviral therapies as well as provide basic information on the structure of complex macromolecules. The HBV core gene codes for precore protein (pre-C) which is either partially processed to form a secreted non-particulate protein called e-antigen (HBeAg) or fully processed to produce core antigen (HBcAg). HBcAg is a 183-residue protein that encapsidate around a RNA-reverse transcriptase complex (HBV polymerase). HBcAg has been expressed in E.coli were it assembles in the bacterial cytoplasm into icosahedral capsids, which contain bound host nucleic acid. Deletion of the polybasic C-terminal 34 residues (protamine domain) also produces assembly competent protein. The capsids from C-terminal truncated protein (Cp149: residues 1-149) do not contain nucleic acid and their structure determined by cryo-electron microscopy and image analysis and by X-ray crystallography. Native HBeAg is also C-terminally truncated at position 149 and in addition contains a 10 residue N-terminal extension derived from partial processing of pre-C. Although the function and structure of HBeAg are unclear it is an important serological marker. Results: using surface plasmon resonance (Biacore) a kinetic-affinity map of a panel of monoclonal antibodies (mAbs) against HBV nucleocapsid proteins revealed a range of binding affinities. Some of the HBV nucleocapsid-antibody complexes were characterized further by cryo-electron microscopy (A.Steven). The results revealed a greater number of discontinuous epitopes than had been described previously. The findings help explain the immunological distinction between the assembled HBcAg and unassembled HBeAg antigens. Structural studies on HBeAg are continuing using mutants with improved physical characteristics. Work is also progressing on the large HBV polymerase protein. In an incremental approach we have expressed various functional domains of polymerse which will be subjected to crystallization trails. Summary: The Hepatitis B Virus (HBV) is the major worldwide cause of cancer. Although a vaccine is available, chronic HBV is often acquired in childhood. The HBV nucleocapsid plays an important structural role and metabolic role in the life cycle of the virus. An understanding of the molecular structure of the HBV nucleocapsid would allow targeted drug discovery with the aim of preventing the assembly and formation of the virus.