Enhanced stability of a chimeric hepatitis B core antigen virus-like-particle (HBcAg-VLP) by a C-terminal linker-hexahistidine-peptide

Abstract

BackgroundVirus-like-particles (VLPs) are attractive nanoparticulate scaffolds for broad applications in material/biological sciences and medicine. Prior their functionalization, specific adaptations have to be carried out. These adjustments frequently lead to disordered particles, but the particle integrity is an essential factor for the VLP suitability. Therefore, major requirements for particle stabilization exist. The objective of this study was to evaluate novel stabilizing elements for functionalized chimeric hepatitis B virus core antigen virus-like particles (HBcAg-VLP), with beneficial characteristics for vaccine development, imaging or delivery.ResultsThe effects of a carboxy-terminal polyhistidine-peptide and an intradimer disulfide-bridge on the stability of preclinically approved chimeric HBcAg-VLPs were assessed. We purified recombinant chimeric HBcAg-VLPs bearing different modified C-termini and compared their physical and chemical particle stability by quantitative protein-biochemical and biophysical techniques. We observed lower chemical resistance of T = 3- compared to T = 4-VLP (triangulation number) capsids and profound impairment of accessibility of hexahistidine-peptides in assembled VLPs. Histidines attached to the C-terminus were associated with superior mechanical and/or chemical particle stability depending on the number of histidine moieties. A molecular modeling approach based on cryo-electron microscopy and biolayer interferometry revealed the underlying structural mechanism for the strengthening of the integrity of VLPs. Interactions triggering capsid stabilization occur on a highly conserved residue on the basis of HBcAg-monomers as well as on hexahistidine-peptides of adjacent monomers. This new stabilization mechanism appears to mimic an evolutionary conserved stabilization concept for hepadnavirus core proteins.ConclusionsThese findings establish the genetically simply transferable C-terminal polyhistidine-peptide as a general stabilizing element for chimeric HBcAg-VLPs to increase their suitability.Electronic supplementary materialThe online version of this article (10.1186/s12951-018-0363-0) contains supplementary material, which is available to authorized users.

Plant-Produced Antigen Displaying Virus-Like Particles Evokes Potent Antibody Responses against West Nile Virus in Mice

“…For downstream processing, HBcAg-wDIII VLPs were easily purified to homogeneity with a one-step process, which is routinely used for VLP vaccine production in the pharmaceutical industry and is in full compliance with regulations of current Good Manufacturing Practice (cGMP) [ 47 ]. Capsid protein-based VLPs including HBcAg-derived VLPs have been shown to have superior stability over other protein-based subunit vaccines under diverse physical stress conditions [ 48 ]. The combined advantages of rapid high-level production, facile purification, and increased stability of HBcAg-wDIII VLPs suggest the feasibility of using plants to produce HBcAg-wDIII VLP-based vaccines at a large scale and low cost.…”

Ensembles of Hydrophobicity Scales as Potent Classifiers for Chimeric Virus-Like Particle Solubility – An Amino Acid Sequence-Based Machine Learning Approach

“…Chimeric HBcAg VLP constructs were based on His-tagged C-terminally truncated HBcAg (Schumacher et al, 2018). The molecules were created using 82 different peptide inserts and eight different insertion strategies, a total of 691 chimeric VLP constructs, which were experimentally tested for solubility.…”

Cost‐effective purification process development for chimeric hepatitis B core (HBc) virus‐like particles assisted by molecular dynamic simulation

“…For example, Schumacher et al. claimed that self‐aggregation of HBc VLP was formed over time in Tris‐buffer [17].…”

“…In addition, easily formation of mis-folding and aggregation was found after insertion of epitope to HBc VLP [15,16]. These challenges can lead to failure to induce epitope-specific immune response [17].…”