Microbiology & Immunology - Theses

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Subject: vaccine × Start date: 2010 × End date: 2019 × Author: McCaffrey, Kathleen ×

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    Hepatitis C virus envelope glycoprotein E2: isolation and characterisation of a functional core domain
    McCaffrey, Kathleen ( 2010)
    Hepatitis C virus (HCV) is a major indicator of liver disease and over 200 million chronic infections are estimated worldwide. No vaccine is available and current treatments report limited efficacy. A strong neutralizing antibody response is a key determinant in clearance of acute HCV infection although often appears delayed as broadly neutralizing antibodies do not appear until after chronic infection is established. Recent studies suggest that HCV evades neutralization through the accumulation of immune escape variants within a highly variable, immunodominant sequence of the viral envelope glycoprotein E2. The E1 and E2 envelope glycoproteins are transmembrane proteins that are embedded in the viral membrane and mediate virus attachment and entry into liver cells. The E2 glycoprotein has been shown to interact with the cellular receptors scavenger receptor class B type 1 (SRB1) and CD81 during virus entry and is a major target for neutralizing antibodies. There is currently no high-resolution structure of the HCV E2 glycoprotein to further understand its mechanism of viral entry or immune evasion. However, a soluble E2 ectodomain fragment has been identified that can be efficiently secreted from cells and displays CD81 receptor-binding function. The E2 ectodomain has three discrete variable regions interspersed between conserved CD81-binding motifs: the immunodominant hypervariable region 1 (HVR1), hypervariable region 2 (HVR2) and the intergenotypic variable region (igVR). In this study, simultaneous substitution of these variable regions (VRs) with short, flexible linker motifs within diverse E2 ectodomain sequences was described with the retention of both native folding and CD81 binding function. This indicated that the E2 VRs are excluded from the functional core domain of the glycoprotein. The conserved E2 core domain, lacking all three VRs, was further shown to elicit higher titers of broadly neutralizing antibodies than the unmodified E2 ectodomain suggesting that the VRs occlude conserved neutralization-sensitive epitopes within the underlying core domain. The E2 VRs were demonstrated to modulate CD81 binding in a manner consistent with solvent-exposed structures and illustrated a potential mechanism by which the VRs could mediate immune evasion during acute infection. Therefore, the conserved E2 core domain was proposed to represent a novel and improved antigen for redirecting the immune response towards conserved, neutralization-sensitive epitopes within the E2 glycoprotein. Further biochemical and functional characterization of the E2 core domain is also presented towards optimization of this antigen as viable candidate for future vaccine trials.