Engineering the PP7 Virus Capsid as a Peptide Display Platform

Abstract

As self-assembling polyvalent nanoscale structures that can tolerate substantial genetic and chemical modification, virus-like particles are useful in a variety of fields. Here we describe the genetic modification and structural characterization of the Leviviridae PP7 capsid protein as a platform for the presentation of functional polypeptides. This particle was shown to tolerate the display of sequences from 1 kDa (a cell penetrating peptide) to 14 kDa (the Fc-binding double Zdomain) on its exterior surface as C-terminal genetic fusions to the coat protein. In addition, a dimeric construct allowed the presentation of exogenous loops between capsid monomers and the simultaneous presentation of two different peptides at different positions on the icosahedral structure. The PP7 particle is thereby significantly more tolerant of these types of polypeptide additions than Qβ and MS2, the other Leviviridae-derived VLPs in common use.

Viral nanoparticles for drug delivery, imaging, immunotherapy, and theranostic applications

“…With genetic fusion however, typically only short peptide fragments can be fused to avoid disrupting capsid assembly. Finn et al overcame this shortcoming with a PP7 phage-derived VLP platform with Cterminal fused peptides [84]. PP7 is amenable to C-terminus display as it is signficantly less crowded around its threefold axes increasing its tolerance to peptide and protein display.…”

Bacteriophage MS2 displays unreported capsid variability assembling T = 4 and mixed capsids

“…On the observation of divergent symmetry VLPs, we created an otherwise identical wild-type (wt) construct with monomeric CP and prepared cryo-EM grids to confirm or deny that the presence of the CP-His-CP dimer was not solely responsible for the formation of mixed and large capsids as previously reported for non-wild-type constructs (Peabody and Chakerian, 1999;Plevka et al, 2008Plevka et al, , 2009Asensio et al, 2016;Zhao et al, 2019). We confirmed similar observations in the micrographs of the wild-type setting; in the higher molecular weight fractions of the wt-CP sample, the majority of the particles belonged to the smaller class, ~85% of the total on sorting by 2D classification, while several varieties of 'large' particles accounted for the remaining ~15%, of which ~6% later proved to be large icosahedral particles ( Supplementary Fig.…”

Virus‐Like Particles as Theranostic Platforms

“…[ 44 ] In some cases, these changes may have little impact on the assembly and stability of the shell, whereas in other cases, they may prevent particle assembly, seriously damage the stability of the particle, or damage some shell functions. [ 38,74 ] Insertion in an exposed target peptide loop usually limits the conformational integrity of the inserted peptide segment, because both ends of the peptide segment remain in the specified position because they are covalently bonded to the structural subunit of the viral protein. [ 61 ] In some cases, this restriction may prevent or impair the functional conformation of the inserted peptide.…”

“…Peptide or protein sequences can be added directly to the primary amino acid sequences of the coat proteins (CPs), as either insertions or extensions, to allow their presentation on either the interior or exterior surface of the VLP. [ 38–41 ] Alternatively, reactive amino acids can be used to couple cargo molecules to the interior or ligands to the exterior of the capsids, in repeated and consistent orientations. [ 42–44 ] VLPs are much less toxic after parenteral administration than metal NPs, are more stable than liposomes, and are more uniform than polymer NPs.…”