Dataset Information

Importin ? Can Bind Hepatitis B Virus Core Protein and Empty Core-Like Particles and Induce Structural Changes.

ABSTRACT: Hepatitis B virus (HBV) capsids are found in many forms: immature single-stranded RNA-filled cores, single-stranded DNA-filled replication intermediates, mature cores with relaxed circular double-stranded DNA, and empty capsids. A capsid, the protein shell of the core, is a complex of 240 copies of core protein. Mature cores are transported to the nucleus by a complex that includes both importin ? and importin ? (Imp? and Imp?), which bind to the core protein's C-terminal domains (CTDs). Here we have investigated the interactions of HBV core protein with importins in vitro. Strikingly, empty capsids and free core protein can bind Imp? without Imp?. Cryo-EM image reconstructions show that the CTDs, which are located inside the capsid, can extrude through the capsid to be bound by Imp?. Imp? density localized on the capsid exterior near the quasi-sixfold vertices, suggested a maximum of 30 Imp? per capsid. However, examination of complexes using single molecule charge-detection mass spectrometry indicate that some complexes include over 90 Imp? molecules. Cryo-EM of capsids incubated with excess Imp? shows a population of damaged particles and a population of "dark" particles with internal density, suggesting that Imp? is effectively swallowed by the capsids, which implies that the capsids transiently open and close and can be destabilized by Imp?. Though the in vitro complexes with great excess of Imp? are not biological, these results have implications for trafficking of empty capsids and free core protein; activities that affect the basis of chronic HBV infection.

SUBMITTER: Chen C

PROVIDER: S-EPMC4982637 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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Importin β Can Bind Hepatitis B Virus Core Protein and Empty Core-Like Particles and Induce Structural Changes.

Chen Chao C Wang Joseph Che-Yen JC Pierson Elizabeth E EE Keifer David Z DZ Delaleau Mildred M Gallucci Lara L Cazenave Christian C Kann Michael M Jarrold Martin F MF Zlotnick Adam A

PLoS pathogens 20160812 8

Hepatitis B virus (HBV) capsids are found in many forms: immature single-stranded RNA-filled cores, single-stranded DNA-filled replication intermediates, mature cores with relaxed circular double-stranded DNA, and empty capsids. A capsid, the protein shell of the core, is a complex of 240 copies of core protein. Mature cores are transported to the nucleus by a complex that includes both importin α and importin β (Impα and Impβ), which bind to the core protein's C-terminal domains (CTDs). Here we ...[more]

PMID: 27518410

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Project description:A vaccine to prevent hepatitis C virus (HCV) infection is urgently needed for use alongside direct-acting antiviral drugs to achieve elimination targets. We have previously shown that a soluble recombinant form of the glycoprotein E2 ectodomain (residues 384 to 661) that lacks three variable regions (Δ123) is able to elicit a higher titer of broadly neutralizing antibodies (bNAbs) than the parental form (receptor-binding domain [RBD]). In this study, we engineered a viral nanoparticle that displays HCV glycoprotein E2 on a duck hepatitis B virus (DHBV) small surface antigen (S) scaffold. Four variants of E2-S virus-like particles (VLPs) were constructed: Δ123-S, RBD-S, Δ123A7-S, and RBDA7-S; in the last two, 7 cysteines were replaced with alanines. While all four E2-S variant VLPs display E2 as a surface antigen, the Δ123A7-S and RBDA7-S VLPs were the most efficiently secreted from transfected mammalian cells and displayed epitopes recognized by cross-genotype broadly neutralizing monoclonal antibodies (bNMAbs). Both Δ123A7-S and RBDA7-S VLPs were immunogenic in guinea pigs, generating high titers of antibodies reactive to native E2 and able to prevent the interaction between E2 and the cellular receptor CD81. Four out of eight animals immunized with Δ123A7-S elicited neutralizing antibodies (NAbs), with three of those animals generating bNAbs against 7 genotypes. Immune serum generated by animals with NAbs mapped to major neutralization epitopes located at residues 412 to 420 (epitope I) and antigenic region 3. VLPs that display E2 glycoproteins represent a promising vaccine platform for HCV and could be adapted to large-scale manufacturing in yeast systems. IMPORTANCE There is currently no vaccine to prevent hepatitis C virus infection, which affects more than 71 million people globally and is a leading cause of progressive liver disease, including cirrhosis and cancer. Broadly neutralizing antibodies that recognize the E2 envelope glycoprotein can protect against heterologous viral infection and correlate with viral clearance in humans. However, broadly neutralizing antibodies are difficult to generate due to conformational flexibility of the E2 protein and epitope occlusion. Here, we show that a VLP vaccine using the duck hepatitis B virus S antigen fused to HCV glycoprotein E2 assembles into virus-like particles that display epitopes recognized by broadly neutralizing antibodies and elicit such antibodies in guinea pigs. This platform represents a novel HCV vaccine candidate amenable to large-scale manufacture at low cost.

Dataset Information

Importin ? Can Bind Hepatitis B Virus Core Protein and Empty Core-Like Particles and Induce Structural Changes.

Publications

Importin β Can Bind Hepatitis B Virus Core Protein and Empty Core-Like Particles and Induce Structural Changes.

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