Overcoming Symmetry Mismatch in Vaccine Nanoassembly through Spontaneous Amidation

Abstract

Matching of symmetry at interfaces is a fundamental obstacle in molecular assembly. Virus-like particles (VLPs) are important platforms to generate vaccines against pathogenic threats including Covid-19. However, symmetry mismatch between multimeric antigens and VLP subunits can prohibit vaccine nanoassembly. Here we explore spontaneous amidation to establish principles for coupling VLPs to diverse antigen symmetries. SpyTag003/SpyCatcher003-mediated decoration enabled efficient VLP conjugation and extreme thermal resilience. Many people showed pre-existing antibodies to SpyTag:SpyCatcher, but decreased antibodies against 003 variants. We coupled efficiently to the computationally-designed VLP, built from twenty trimers, not only monomers (SARS-CoV-2) but also cyclic dimers (Newcastle Disease Virus or Lyme disease), trimers (Influenza Hemagglutinins), and tetramers (Influenza Neuraminidases). Even small antigens with dihedral symmetry could be displayed. For the global challenge of Influenza, Spy-display enabled powerful induction of neutralizing antibodies to trimeric and tetrameric antigens. SpyCatcher003 conjugation is compatible with nanodisplay of diverse symmetries, towards generation of potent vaccines.

Nanoparticle and virus-like particle vaccine approaches against SARS-CoV-2

“…This "plug-and-display" system can link the SpyTagantigen to the SpyCatcher-scaffold protein (or vice versa) by spontaneous isopeptide bond formation. Recently, another modification was introduced to SpyTag/SpyCatcher to achieve a more efficient antigen display and enhanced stability (Rahikainen et al, 2021). Fc-tagged SARS-CoV-2 RBD has also been coupled with a protein A-expressing VLP scaffold (Geng et al, 2021).…”

“…There is a demand for the development of a novel linking system as people may possess linker-specific immune responses or develop such responses by repeated immunizations. Due to its bacterial origin, 98% of the tested human samples were positive for antibodies specific for SpyTag/ SpyCatcher protein (Rahikainen et al, 2021). It is not clear if linker-specific antibodies affect the immunogenicity of NP and VLP vaccines.…”

“…This self-assembling NP scaffold can display 20 trimeric antigens (60-mer). The platform is thermally stable and, therefore, useful for vaccine supply, where cold-chain storage is problematic (Liu et al, 2021b;Rahikainen et al, 2021). mi3 has been used to develop vaccines against influenza virus (Cohen et al, 2021b), classical swine fever virus (Liu et al, 2021b(Liu et al, , 2021c, and malaria (Bruun et al, 2018) in addition to vaccine candidates for SARS-CoV-2 (Cohen et al, 2021a;Guo et al, 2021a;Halfmann et al, 2021;Kang et al, 2021;Tan et al, 2021).…”

N‐Terminal Modification of Gly‐His‐Tagged Proteins with Azidogluconolactone

“…(A,C) Scheme for biosynthesis of nanoparticles (PDB 2MS2, SpyCatcher-mi3 trimer was modelled with AlphaFold v2.0 [34] and PDB 7B3Y was used for N-terminal GH3-tagging the coat proteins of the bacteriophages MS2 [35] , PQ-465 [36] , Shahe1 [37] , and AP205d (single-chain dimer) [38] permitted VLP assembly, IMAC purification, and (6A)GDL labelling. GSSH6-tagged "Plug-and-Display" SpyCatcher-mi3 [39,40] protein nanoparticles also reacted (Figure 3, Figure S25). However, not all VLP-fusions were Gly-His compatible in our hands.…”

“…[66] It is yet unknown if pre-existing Abs in the human population against SpyCatcher and/or HBsAg could be problematic, and whether this could be overcome by increasing dose. [39] The emerging threat of vaccine escape mutants may require additional approaches. [40,74] Possible synergy, also for other antigens, may be obtained by heterologous boosting to circumvent any carrier-induced suppression.…”