A structural model for maturation of the hepatitis B virus core

Abstract

Hepatitis B virus, a widespread and serious human pathogen, replicates by reverse transcription of an RNA intermediate. The virus consists of an inner nucleocapsid or core, surrounded by a lipid envelope containing virally encoded surface proteins. Using electron cryomicroscopy, we compare the structures of the bacterially expressed RNA-containing core particle and the mature DNA-containing core particle extracted from virions. We show that the mature core contains 240 subunits in a T ‫؍‬ 4 arrangement similar to that in expressed core (T is the triangulation number and the icosahedral shell contains 60 T subunits). During the infective cycle, the core assembles in an immature state around a complex of viral pregenomic RNA and polymerase. After reverse transcription with concomitant degradation of the RNA, the now mature core buds through a cellular membrane containing the surface proteins to become enveloped. Envelopment must not happen before reverse transcription is completed, so it has been hypothesized that a change in capsid structure may signal maturation. Our results show significant differences in structure between the RNAand DNA-containing cores. One such difference is in a hydrophobic pocket, formed largely from residues that, on mutation, lead to abnormal secretion. We suggest that the changes we see are related to maturation and control of envelopment, and we propose a mechanism based on DNA synthesis for their triggering.cryomicroscopy ͉ envelopment ͉ nucleocapsid ͉ viral maturation H epatitis B virus (HBV) is a major human pathogen and member of the Hepadnaviridae, a family of viruses that replicate by reverse transcription in the hepatoctyes of their hosts (1). During virus assembly, the core protein polymerizes around a complex consisting of pregenomic mRNA and viral polymerase to form an immature core. The RNA is reverse transcribed into single-stranded DNA and concomitantly degraded, then the partial second DNA strand is synthesized. Cores are then mature and can interact with the cytoplasmic parts of the surface proteins, already inserted into an inner cellular membrane, through which the core buds to form a virion (see Bruss, ref. 2).

Cryo-electron microscopy of hepatitis B virions reveals variability in envelope capsid interactions

“…Escherichia coli-derived core protein (HBc), devoid of any HBV-specific nucleic acid forms large (240 subunits, T ¼ 4 arrangement) and small (180 subunits, T ¼ 3 arrangement) core particles (Crowther et al, 1994;Böttcher et al, 1997;Conway et al, 1997;Wynne et al, 1999). In virions, mainly the large nucleocapsid particles are found (Roseman et al, 2005;Dryden et al, 2006). The building blocks of the cores are HBc-dimers with a spike in the center that is composed of a four helix bundle to which each monomer contributes two helices.…”

Three-dimensional structure of the hepatitis B core antigen particle truncated at residue 154

“…The RNA density was not observed inside the capsid while a strong shell of RNA was present between radii 90-125 Å in HBcAg-183 [9]. As one kind of HBcAg particle constructed by the entire core protein, HBcAg-183 encapsidates large amounts of RNA in the capsid.…”

“…Its inner space is empty without observing RNA for the lack of a C-terminal tail. The capsid structure of the full-length HBcAg particle (HBcAg-183) is similar to that of HBcAg-149, while HBcAg-183 contains large amounts of RNA [8,9]. The difference of RNA content indicates the significance of the C-terminal tail of the core protein in packaging RNA.…”

The Leeuwenhoek lecture 2006. Microscopy goes cold: frozen viruses reveal their structural secrets

“…These observations suggest a model (Roseman et al 2005) in which DNA synthesis inside the core causes changes in the core protein shell that propagate to the outside and alter the hydrophobic pocket in the spike. In the immature RNA-containing core, the pocket is in a closed state that cannot interact with the surface proteins, but after DNA synthesis the open pocket is able to interact with the surface proteins, and envelopment and secretion ensue.…”

“…In order to investigate the nature of the maturation signal, it was necessary to compare the structure of immature RNA-containing cores, made by expressing the full-length core protein in bacteria, with the structure of mature DNA-containing cores, extracted from viruses isolated from infected blood samples (Roseman et al 2005). The two structures (figure 7) are very similar to each other and to the empty core particle made from truncated protein (figure 4).…”