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Epitope-distal effects accompany the binding of two distinct antibodies to hepatitis B virus capsids.

ABSTRACT: Infection of humans by hepatitis B virus (HBV) induces the copious production of antibodies directed against the capsid protein (Cp). A large variety of anticapsid antibodies have been identified that differ in their epitopes. These data, and the status of the capsid as a major clinical antigen, motivate studies to achieve a more detailed understanding of their interactions. In this study, we focused on the Fab fragments of two monoclonal antibodies, E1 and 3120. E1 has been shown to bind to the side of outward-protruding spikes whereas 3120 binds to the "floor" region of the capsid, between spikes. We used hydrogen-deuterium exchange coupled to mass spectrometry (HDX-MS) to investigate the effects on HBV capsids of binding these antibodies. Conventionally, capsids loaded with saturating amounts of Fabs would be too massive to be readily amenable to HDX-MS. However, by focusing on the Cp protein, we were able to acquire deuterium uptake profiles covering the entire 149-residue sequence and reveal, in localized detail, changes in H/D exchange rates accompanying antibody binding. We find increased protection of the known E1 and 3120 epitopes on the capsid upon binding and show that regions distant from the epitopes are also affected. In particular, the ?2a helix (residues 24-34) and the mobile C-terminus (residues 141-149) become substantially less solvent-exposed. Our data indicate that even at substoichiometric antibody binding an overall increase in the rigidity of the capsid is elicited, as well as a general dampening of its breathing motions.

SUBMITTER: Bereszczak JZ

PROVIDER: S-EPMC3658141 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature

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Epitope-distal effects accompany the binding of two distinct antibodies to hepatitis B virus capsids.

Bereszczak Jessica Z JZ Rose Rebecca J RJ van Duijn Esther E Watts Norman R NR Wingfield Paul T PT Steven Alasdair C AC Heck Albert J R AJ

Journal of the American Chemical Society 20130418 17

Infection of humans by hepatitis B virus (HBV) induces the copious production of antibodies directed against the capsid protein (Cp). A large variety of anticapsid antibodies have been identified that differ in their epitopes. These data, and the status of the capsid as a major clinical antigen, motivate studies to achieve a more detailed understanding of their interactions. In this study, we focused on the Fab fragments of two monoclonal antibodies, E1 and 3120. E1 has been shown to bind to the ...[more]

PMID: 23597076

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Project description:BackgroundAround 30% of the HCV infected patients can spontaneously clear the virus. Cumulative evidence suggests the role of neutralizing antibodies in such spontaneous resolution. Understanding the epitope specificity of such antibodies will inform the rational vaccine design as such information is limited to date. In addition to conformational epitope targeted antibodies, linear epitope specific antibodies have been identified that are broadly cross reactive against diverse HCV strains. In this study, we have characterized the potential role of three conserved linear epitopes in the spontaneous clearance of HCV.MethodsWe tested the reactivity of sera from chronic patients (CP) and spontaneous resolvers (SR) with linear peptides corresponding to three conserved regions of HCV envelope protein E2 spanning amino acids 412-423, 523-532 and 432-443 using ELISA. Subsequently, we characterized the dependency of HCV neutralization by the reactive serum samples on the antibodies specific for these epitopes using pseudoparticle-based neutralization assay. In ELISA most of the CP sera showed reactivity to multiple peptides while most of the SR samples were reactive to a single peptide suggesting presence of more specific antibodies in the SR sera. In most of the HCVpp neutralizing sera of particular peptide reactivity the neutralization was significantly affected by the presence of respective peptide. HCV neutralization by CP sera was affected by multiple peptides while 75% of the HCVpp neutralizing SR sera were competed by the 432 epitope.ConclusionsThese findings suggest that individuals who spontaneously resolve HCV infection at the acute phase, can produce antibodies specific for conserved linear epitopes, and those antibodies can potentially play a role in the spontaneous viral clearance. The epitope present in the 432-443 region of E2 was identified as the primary neutralizing epitope with potential role in spontaneous viral clearance and this epitope potentiates for the design of immunogen for prophylactic vaccine.

Project description:UnlabelledHepatitis C virus (HCV) neutralization occurring at the E2 region 412-426 (EP-I) could be enhanced when antibodies directed specifically to the E2 region 434-446 (EP-II) were removed from serum samples of persistently infected patients and vaccinated chimpanzees, a phenomenon of so-called antibody interference. Here, we show that this type of interference can be observed in individuals after immunization with recombinant E1E2 proteins. One hundred twelve blinded serum samples from a phase I, placebo-controlled, dose escalation trial using recombinant HCV E1E2 with MF59C.1 adjuvant in healthy HCV-negative adults were tested in enzyme-linked immunosorbent assay for binding reactivity to peptides representing the E2 regions 412-426 (EP-I) and 434-446 (EP-II). All samples were subsequently tested for neutralizing activity using cell-culture HCV 1a(H77)/2a chimera, HCV pseudotype particles (HCVpp) H77, and HCVpp HCV-1 after treatment to remove EP-II-specific antibodies or mock treatment with a control peptide. Among the 112 serum samples, we found 22 double positive (EP-I and EP-II), 6 EP-II positive only, 14 EP-I positive only, and 70 double negative. Depleting EP-II antibodies from double-positive serum samples increased 50% inhibitory dose (ID50) neutralizing antibody titers (up to 4.9-fold) in up to 72% of samples (P ≤ 0.0005), contrasting with ID50 neutralization titer increases in 2 of 70 double-negative samples (2.9%; P > 0.5). In addition, EP-I-specific antibody levels in serum samples showed a significant correlation with ID50 neutralization titers when EP-II antibodies were removed (P < 0.0003).ConclusionThese data show that antibodies to the region 434-446 are induced during immunization of individuals with recombinant E1E2 proteins, and that these antibodies can mask effective neutralizing activity from EP-I-specific antibodies. Elicitation of EP-II-specific antibodies with interfering capacity should be avoided in producing an effective cross-neutralizing vaccine aimed at the HCV envelope proteins.

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Epitope-distal effects accompany the binding of two distinct antibodies to hepatitis B virus capsids.

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Epitope-distal effects accompany the binding of two distinct antibodies to hepatitis B virus capsids.

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