title: Evaluation of novel Streptococcus pyogenes vaccine candidates incorporating multiple conserved sequences from the C-repeat region of the M-protein.
abstract: A major challenge for Streptococcus pyogenes vaccine development is the identification of epitopes that confer protection from infection by multiple S. pyogenes M-types. Here we have identified and characterised the distribution of common variant sequences from individual repeat units of the C-repeat region (CRR) of M-proteins representing 77 different M-types. Three polyvalent fusion vaccine candidates (SV1, SV2 and SV3) incorporating the most common variants were subsequently expressed and purified, and demonstrated to be alpha-helical by Circular Dichroism (CD), a secondary conformational characteristic of the CRR in the M-protein. Antibodies raised against each of these constructs recognise M-proteins that vary in their CRR, and bind to the surface of multiple S. pyogenes isolates. Antibodies raised against SV1, containing five variant sequences, also kill heterologous S. pyogenes isolates in in vitro bactericidal assays. Further structural characterisation of this construct demonstrated the conformation of SV1 was stable at different pHs, and thermal unfolding of SV1 is a reversible process. Our findings demonstrate that linkage of multiple variant sequences into a single recombinant construct overcomes the need to embed the variant sequences in foreign helix promoting flanking sequences for conformational stability, and demonstrates the viability of the polyvalent candidates as global S. pyogenes vaccine candidates.
authors.affiliation: Bacterial Pathogenesis Laboratory, The Queensland Institute of Medical Research, Brisbane, Queensland 4029, Australia.
authors.forename: Michelle J
authors.initials: MJ
authors.lastname: Bauer
authors.forename: Melina M
authors.initials: MM
authors.lastname: Georgousakis
authors.forename: Therese
authors.initials: T
authors.lastname: Vu
authors.forename: Anna
authors.initials: A
authors.lastname: Henningham
authors.forename: Andreas
authors.initials: A
authors.lastname: Hofmann
authors.forename: Mandy
authors.initials: M
authors.lastname: Rettel
authors.forename: Louise M
authors.initials: LM
authors.lastname: Hafner
authors.forename: Kadaba S
authors.initials: KS
authors.lastname: Sriprakash
authors.forename: David J
authors.initials: DJ
authors.lastname: McMillan
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001770:Blood Bactericidal Activity; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D002942:Circular Dichroism; D017124:Conserved Sequence; D000939:Epitopes / Q000235:genetics / Q000276:immunology*; D005260:Female; D051379:Mice; D050296:Microbial Viability / Q000187:drug effects; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D020449:Repetitive Sequences, Amino Acid / Q000235:genetics / Q000276:immunology; D022541:Streptococcal Vaccines / Q000008:administration & dosage* / Q000235:genetics / Q000276:immunology*; D013297:Streptococcus pyogenes / Q000235:genetics / Q000276:immunology*; D014614:Vaccines, Synthetic / Q000235:genetics / Q000276:immunology
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D000939:Epitopes; D011993:Recombinant Fusion Proteins; D022541:Streptococcal Vaccines; D014614:Vaccines, Synthetic; C014689:streptococcal M protein