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Conserved glycine 33 residue in flexible domain I of hepatitis C virus core protein is critical for virus infectivity.

ABSTRACT: Hepatitis C virus core protein forms the viral nucleocapsid and plays a critical role in the formation of infectious particles. In this study, we demonstrate that the highly conserved residue G33, located within domain 1 of the core protein, is important for the production of cell culture-infectious virus (HCVcc). Alanine substitution at this position in the JFH1 genome did not alter viral RNA replication but reduced infectivity by ?2 logs. Virus production by this core mutant could be rescued by compensatory mutations located immediately upstream and downstream of the original G33A mutation. The examination of the helix-loop-helix motif observed in the core protein structure (residues 15 to 41; Protein Data Bank entry 1CWX) indicated that the residues G33 and F24 are in close contact with each other, and that the G33A mutation induces a steric clash with F24. Molecular simulations revealed that the compensatory mutations increase the helix-loop-helix flexibility, allowing rescue of the core active conformation required for efficient virus production. Taken together, these data highlight the plasticity of core domain 1 conformation and illustrate the relationship between its structural tolerance to mutations and virus infectivity.

SUBMITTER: Angus AG

PROVIDER: S-EPMC3255827 | biostudies-literature | 2012 Jan

REPOSITORIES: biostudies-literature

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Conserved glycine 33 residue in flexible domain I of hepatitis C virus core protein is critical for virus infectivity.

Angus Allan G N AG Loquet Antoine A Stack Séamus J SJ Dalrymple David D Gatherer Derek D Penin François F Patel Arvind H AH

Journal of virology 20111109 2

Hepatitis C virus core protein forms the viral nucleocapsid and plays a critical role in the formation of infectious particles. In this study, we demonstrate that the highly conserved residue G33, located within domain 1 of the core protein, is important for the production of cell culture-infectious virus (HCVcc). Alanine substitution at this position in the JFH1 genome did not alter viral RNA replication but reduced infectivity by ∼2 logs. Virus production by this core mutant could be rescued b ...[more]

PMID: 22072760

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Project description:The baculovirus VP39 protein is a major nucleocapsid protein essential for viral propagation. However, the critical domains or residues of the VP39 protein have not yet been identified. Here, we performed mutagenesis experiments with Bombyx mori nucleopolyhedrovirus (BmNPV) using 5-bromo-2'-deoxyuridine and isolated a BmNPV mutant that produced fewer occlusion bodies than the wild-type virus. This mutant also produced fewer infectious budded viruses (BVs) than the wild-type virus in both cultured cells and B. mori larvae. Marker rescue experiments using genomic libraries identified a single nucleotide mutation in the vp39 gene. This mutation resulted in an amino acid substitution at glycine 276 (Gly-276) to serine, which was required for all the defective phenotypes observed in the mutant. Sequence comparison revealed that this residue is completely conserved among the VP39 proteins of the sequenced alphabaculoviruses, betabaculoviruses, and gammabaculoviruses. Although early viral gene expression was not significantly affected, the level of expression of a late gene, vcath, was reduced. In addition, two of the very late genes were markedly downregulated in cells infected with this mutant. Western blot and quantitative PCR analyses revealed that the BVs produced from cells infected with this mutant contained smaller amounts of the VP39 protein and viral genomic DNA than those produced from wild-type virus-infected cells. Combined with the results of transmission electron microscopy, VP39 Gly-276 can be concluded to be essential for correct nucleocapsid assembly, viral DNA packaging, and viral gene expression, especially of very late genes.IMPORTANCE The major nucleocapsid protein gene vp39 is one of the most well-known baculovirus genes. Although several viral and host proteins that interact with the VP39 protein have been identified, the functionally important domains or residues of this protein remain unknown. The present study revealed that the glycine residue at residue 276, which is completely conserved among sequenced alphabaculoviruses, betabaculoviruses, and gammabaculoviruses, is important for the VP39 function, i.e., structural assembly of nucleocapsids and viral DNA packaging. Moreover, our results provide evidence for the link between nucleocapsid formation and the transcription of viral very late genes.

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Conserved glycine 33 residue in flexible domain I of hepatitis C virus core protein is critical for virus infectivity.

Publications

Conserved glycine 33 residue in flexible domain I of hepatitis C virus core protein is critical for virus infectivity.

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