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A Self‐Adjuvanting Vaccine Platform: Optimization of Site‐Specific Sortase A Mediated Conjugation of Toll‐Like Receptor 2 Ligands onto the Carboxyl or Amino terminus of Recombinant Protein Antigens
Author(s) -
Xu Zhenghui,
Moyle Peter Michael
Publication year - 2020
Publication title -
chempluschem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.801
H-Index - 61
ISSN - 2192-6506
DOI - 10.1002/cplu.201900687
Subject(s) - sortase a , lipopeptide , chemistry , sortase , antigen , agonist , recombinant dna , ligation , tlr2 , receptor , biochemistry , microbiology and biotechnology , biology , immunology , innate immune system , bacterial protein , gene , genetics , bacteria
Self‐adjuvanting vaccines, consisting of recombinant protein antigens and covalently attached Toll‐like receptor (TLR) agonists, have the ability to simultaneously and efficiently deliver antigen and TLR adjuvant to antigen presenting cells (APCs). Here, an enzyme‐mediated ligation approach was used to overcome difficulties in producing homogeneous, molecularly defined self‐adjuvanting vaccine products under native conditions. This process was optimized to allow the incorporation of the lipopeptide TLR2 agonist fibroblast‐stimulating lipopeptide (FSL)‐1 onto the N‐ or C‐termini of recombinant protein antigens, employing the enzyme Staphylococcus aureus sortase A (SrtAsa) penta mutant. In addition, because SrtAsa‐mediated ligations are reversible, a tryptophan zipper derived sequence was introduced into both reactants, which was demonstrated to improve ligation efficiency through the formation of a β‐hairpin structure that hinders the reverse reaction. Finally, it was demonstrated that N‐ or C‐terminal conjugation, and the incorporation of the β‐hairpin structure, did not affect the TLR2‐agonist activities of protein antigen TLR agonist conjugates. Overall, this SrtAsa‐mediated ligation platform enabled production of antigen TLR2 agonist conjugates with enhanced ligation efficiency, with the conjugates demonstrating potent TLR2 signaling activation (EC 50 <1nM).