A recombinant hepatitis B core antigen polypeptide with the protamine-like domain deleted self-assembles into capsid particles but fails to bind nucleic acids

Abstract

Escherichia coli both the complete core gene of hepatitis B virus and a truncated version of it, leading to the synthesis of high levels of a core-antigen-equivalent polypeptide (r-p22) and of an e-antigen-equivalent polypeptide (r-p16), respectively. We then compared the structural and antigenic properties of the two polypeptides, as well as their ability to bind viral nucleic acids. r-p16 was found to self-assemble into capsid-like particles that appeared similar, when observed under the electron microscope, to those formed by r-p22. In r-p16 particles, disulfide bonds linked the truncated polypeptides in dimers, assembled in the particle by noncovalent interactions. In r-p22 capsids, further disulfide bonds, conceivably involving the carboxy-terminal cysteines of r-p22 polypeptides, joined the dimers together, converting the structure into a covalently closed lattice. The protamine-like domain was at least partly exposed on the surface of r-p22 particles, since it was accessible to selective proteolysis. Finally, r-p22, but not r-p16, was shown to bind native and denatured DNA as well as RNA. Taken together, these results suggest that the protamine-like domain in core polypeptides is a nucleic acid-binding domain and is dispensable for the correct folding and assembly of amino-terminal and central regions.

Morphogenesis of hepatitis B virus and its subviral envelope particles

“…While the N-terminus of the capsid protein is important for the dimerization process, the basic C-terminal domain contains a nuclear localization sequence (NLS) and an Arg-enriched area. Formed by the last 34 aa, this basic area is probably involved in the pgRNA/reverse-transcriptase complex encapsidation (Gallina et al, 1989;Nassal, 1992). Finally, the HBV core protein structure was solved by crystallization at a resolution of 3.3 Å (Wynne et al, 1999).…”

Extensive Mutagenesis of the Hepatitis B Virus Core Gene and Mapping of Mutations That Allow Capsid Formation

“…These recombinant particles are morphologically and immunologically indistinguishable from natural capsids. Two kinds of particles are formed; one type, composed of 90 dimers, has a Tϭ3 symmetry and a diameter of 32 nm, and the other type, formed by 120 dimers, has a Tϭ4 symmetry and a diameter of 36 nm (10,32). During the assembly process in heterologous systems, nonspecific host RNA is packaged into the particles by interaction with the protamine-like R-rich C-terminal domain of 30 aa (3).…”

Functional Characterization of Naturally Occurring Variants of Human Hepatitis B Virus Containing the Core Internal Deletion Mutation

“…In addition to viral factors, host factors have been hypothesized to be involved in hepadnaviral replication (28,29,49). The core protein has a nucleic acid binding domain rich in arginine residues near the C terminus (9,21,24,43). Serial deletions from the C terminus of the core protein have shown that the arginine-rich domain is not required for multimerization of the core protein, although it might increase the stability of the nucleocapsid (21,43).…”

“…The core protein has a nucleic acid binding domain rich in arginine residues near the C terminus (9,21,24,43). Serial deletions from the C terminus of the core protein have shown that the arginine-rich domain is not required for multimerization of the core protein, although it might increase the stability of the nucleocapsid (21,43). The deletion found in CID variants is located outside the nucleic acid binding domain.…”