Abstract
No vaccine exists against group A Streptococcus (GAS), a leading cause of worldwide morbidity and mortality. A severe hurdle is the hypervariability of its major antigen, the M protein, with >200 different M types known. Neutralizing antibodies typically recognize M protein hypervariable regions (HVRs) and confer narrow protection. In stark contrast, human C4b-binding protein (C4BP), which is recruited to the GAS surface to block phagocytic killing, interacts with a remarkably large number of M protein HVRs (apparently ∼90%). Such broad recognition is rare, and we discovered a unique mechanism for this through the structure determination of four sequence-diverse M proteins in complexes with C4BP. The structures revealed a uniform and tolerant ‘reading head’ in C4BP, which detected conserved sequence patterns hidden within hypervariability. Our results open up possibilities for rational therapies that target the M–C4BP interaction, and also inform a path towards vaccine design.
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References
Carapetis, J. R., Steer, A. C., Mulholland, E. K. & Weber, M. The global burden of group A streptococcal diseases. Lancet Infect. Dis. 5, 685–694 (2005).
Cole, J. N., Barnett, T. C., Nizet, V. & Walker, M. J. Molecular insight into invasive group A streptococcal disease. Nat. Rev. Microbiol. 9, 724–736 (2011).
Dale, J. B. et al. Group A streptococcal vaccines: paving a path for accelerated development. Vaccine 31 (Suppl. 2), B216–B222 (2013).
Good, M. F., Pandey, M., Batzloff, M. R. & Tyrrell, G. J. Strategic development of the conserved region of the M protein and other candidates as vaccines to prevent infection with group A streptococci. Expert Rev. Vaccines 14, 1459–1470 (2015).
McNamara, C. et al. Coiled-coil irregularities and instabilities in group A Streptococcus M1 are required for virulence. Science 319, 1405–1408 (2008).
Ghosh, P. The nonideal coiled coil of M protein and its multifarious functions in pathogenesis. Adv. Exp. Med. Biol. 715, 197–211 (2011).
Sandin, C., Carlsson, F. & Lindahl, G. Binding of human plasma proteins to Streptococcus pyogenes M protein determines the location of opsonic and non-opsonic epitopes. Mol. Microbiol. 59, 20–30 (2006).
Penfound, T. A., Ofek, I., Courtney, H. S., Hasty, D. L. & Dale, J. B. The NH2-terminal region of Streptococcus pyogenes M5 protein confers protection against degradation by proteases and enhances mucosal colonization of mice. J. Infect. Dis. 201, 1580–1588 (2010).
Lannergard, J. et al. The hypervariable region of Streptococcus pyogenes M protein escapes antibody attack by antigenic variation and weak immunogenicity. Cell Host Microbe 10, 147–157 (2011).
Dale, J. B., Penfound, T. A., Chiang, E. Y. & Walton, W. J. New 30-valent M protein-based vaccine evokes cross-opsonic antibodies against non-vaccine serotypes of group A streptococci. Vaccine 29, 8175–8178 (2011).
McMillan, D. J. et al. Updated model of group A Streptococcus M proteins based on a comprehensive worldwide study. Clin. Microbiol. Infect. 19, E222–E229 (2013).
Persson, J., Beall, B., Linse, S. & Lindahl, G. Extreme sequence divergence but conserved ligand-binding specificity in Streptococcus pyogenes M protein. PLoS Pathog. 2, e47 (2006).
Ermert, D. & Blom, A. M. C4b-binding protein: the good, the bad and the deadly. Novel functions of an old friend. Immunol. Lett. 169, 82–92 (2016).
Lambris, J. D., Ricklin, D. & Geisbrecht, B. V. Complement evasion by human pathogens. Nat. Rev. Microbiol. 6, 132–142 (2008).
Blom, A. M., Hallstrom, T. & Riesbeck, K. Complement evasion strategies of pathogens—acquisition of inhibitors and beyond. Mol. Immunol. 46, 2808–2817 (2009).
Carlsson, F., Berggard, K., Stalhammar-Carlemalm, M. & Lindahl, G. Evasion of phagocytosis through cooperation between two ligand-binding regions in Streptococcus pyogenes M protein. J. Exp. Med. 198, 1057–1068 (2003).
Ermert, D. et al. Virulence of group A Streptococci is enhanced by human complement inhibitors. PLoS Pathog. 11, e1005043 (2015).
Fremont, D. H., Matsumura, M., Stura, E. A., Peterson, P. A. & Wilson, I. A. Crystal structures of two viral peptides in complex with murine MHC class I H-2Kb. Science 257, 919–927 (1992).
Madden, D. R., Gorga, J. C., Strominger, J. L. & Wiley, D. C. The three-dimensional structure of HLA-B27 at 2.1 Å resolution suggests a general mechanism for tight peptide binding to MHC. Cell 70, 1035–1048 (1992).
Jenkins, H. T. et al. Human C4b-binding protein, structural basis for interaction with streptococcal M protein, a major bacterial virulence factor. J. Biol. Chem. 281, 3690–3697 (2006).
Accardo, P., Sanchez-Corral, P., Criado, O., Garcia, E. & Rodriguez de Cordoba, S. Binding of human complement component C4b-binding protein (C4BP) to Streptococcus pyogenes involves the C4b-binding site. J. Immunol. 157, 4935–4939 (1996).
Blom, A. M. et al. Human C4b-binding protein has overlapping, but not identical, binding sites for C4b and streptococcal M proteins. J. Immunol. 164, 5328–5336 (2000).
Andre, I. et al. Streptococcal M protein: structural studies of the hypervariable region, free and bound to human C4BP. Biochemistry 45, 4559–4568 (2006).
Lawrence, M. C. & Colman, P. M. Shape complementarity at protein/protein interfaces. J. Mol. Biol. 234, 946–950 (1993).
Sanderson-Smith, M. et al. A systematic and functional classification of Streptococcus pyogenes that serves as a new tool for molecular typing and vaccine development. J. Infect. Dis. 210, 1325–1338 (2014).
Perkins, S. J., Chung, L. P. & Reid, K. B. Unusual ultrastructure of complement-component-C4b-binding protein of human complement by synchrotron X-ray scattering and hydrodynamic analysis. Biochem. J. 233, 799–807 (1986).
Berggard, K. et al. Binding of human C4BP to the hypervariable region of M protein: a molecular mechanism of phagocytosis resistance in Streptococcus pyogenes. Mol. Microbiol. 42, 539–551 (2001).
Gustafsson, M. C. et al. Factor H binds to the hypervariable region of many Streptococcus pyogenes M proteins but does not promote phagocytosis resistance or acute virulence. PLoS Pathog. 9, e1003323 (2013).
Ermert, D. et al. Binding of complement inhibitor C4b-binding protein to a highly virulent Streptococcus pyogenes M1 strain is mediated by protein H and enhances adhesion to and invasion of endothelial cells. J. Biol. Chem. 288, 32172–32183 (2013).
Zhou, T. et al. Structural definition of a conserved neutralization epitope on HIV-1 gp120. Nature 445, 732–737 (2007).
Sui, J. et al. Structural and functional bases for broad-spectrum neutralization of avian and human influenza A viruses. Nat. Struct. Mol. Biol. 16, 265–273 (2009).
Ekiert, D. C. et al. Antibody recognition of a highly conserved influenza virus epitope. Science 324, 246–251 (2009).
McLellan, J. S. et al. Structure of HIV-1 gp120 V1/V2 domain with broadly neutralizing antibody PG9. Nature 480, 336–343 (2011).
Steer, A. C., Law, I., Matatolu, L., Beall, B. W. & Carapetis, J. R. Global emm type distribution of group A streptococci: systematic review and implications for vaccine development. Lancet Infect. Dis. 9, 611–616 (2009).
Doublie, S. Production of selenomethionyl proteins in prokaryotic and eukaryotic expression systems. Methods Mol. Biol. 363, 91–108 (2007).
Kabsch, W. XDS. Acta Crystallogr. D 66, 125–132 (2010).
Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997).
Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D 66, 213–221 (2010).
Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D 60, 2126–2132 (2004).
Chen, V. B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D 66, 12–21 (2010).
Schneider, C. A., Rasband, W. S. & Eliceiri, K. W. NIH Image to ImageJ: 25 years of image analysis. Nat. Methods 9, 671–675 (2012).
Pearlman, D. A. et al. Amber, a package of computer-programs for applying molecular mechanics, normal-mode analysis, molecular-dynamics and free-energy calculations to simulate the structural and energetic properties of molecules. Comput. Phys. Commun. 91, 1–41 (1995).
Wang, J. M., Wolf, R. M., Caldwell, J. W., Kollman, P. A. & Case, D. A. Development and testing of a general amber force field. J. Comput. Chem. 25, 1157–1174 (2004).
Case, D. A. et al. AMBER 2015 (Univ. California, San Francisco, 2015).
Olsson, M. H. M., Sondergaard, C. R., Rostkowski, M. & Jensen, J. H. PROPKA3 consistent treatment of internal and surface residues in empirical pKa predictions. J. Chem. Theory Comput. 7, 525–537 (2011).
Sondergaard, C. R., Olsson, M. H. M., Rostkowski, M. & Jensen, J. H. Improved treatment of ligands and coupling effects in empirical calculation and rationalization of pKa values. J. Chem. Theory Comput. 7, 2284–2295 (2011).
Li, P. F., Roberts, B. P., Chakravorty, D. K. & Merz, K. M. Rational design of particle mesh Ewald compatible Lennard-Jones parameters for +2 metal cations in explicit solvent. J. Chem. Theory Comput. 9, 2733–2748 (2013).
Darden, T., York, D. & Pedersen, L. Particle mesh Ewald—an N log(N) method for Ewald sums in large systems. J. Chem. Phys. 98, 10089–10092 (1993).
Essmann, U. et al. A smooth particle mesh Ewald method. J. Chem. Phys. 103, 8577–8593 (1995).
Nelson, M. T. et al. NAMD: a parallel, object oriented molecular dynamics program. Int. J. Supercomput. Appl. 10, 251–268 (1996).
Phillips, J. C. et al. Scalable molecular dynamics with NAMD. J. Comput. Chem. 26, 1781–1802 (2005).
Izaguirre, J. A., Catarello, D. P., Wozniak, J. M. & Skeel, R. D. Langevin stabilization of molecular dynamics. J. Chem. Phys. 114, 2090–2098 (2001).
Jiang, W. et al. High-performance scalable molecular dynamics simulations of a polarizable force field based on classical Drude oscillators in NAMD. J. Phys. Chem. Lett. 2, 87–92 (2011).
Berendsen, H. J. C., Postma, J. P. M., Vangunsteren, W. F., Dinola, A. & Haak, J. R. Molecular-dynamics with coupling to an external bath. J. Chem. Phys. 81, 3684–3690 (1984).
Caves, L. S. D., Evanseck, J. D. & Karplus, M. Locally accessible conformations of proteins: multiple molecular dynamics simulations of crambin. Protein Sci. 7, 649–666 (1998).
Roe, D. R. & Cheatham, T. E. PTRAJ and CPPTRAJ software for processing and analysis of molecular dynamics trajectory data. J. Chem. Theory Comput. 9, 3084–3095 (2013).
Humphrey, W., Dalke, A. & Schulten, K. VMD visual molecular dynamics. J. Mol. Graph. Model. 14, 33–38 (1996).
Levine, B. G., Stone, J. E. & Kohlmeyer, A. Fast analysis of molecular dynamics trajectories with graphics processing units—radial distribution function histogramming. J. Comput. Phys. 230, 3556–3569 (2011).
Acknowledgements
We thank O. Ghosh for help on the project. This works was supported by National Institutes of Health (NIH) grant T32 GM007240 (C.Z.B.), American Heart Association Predoctoral Fellowship 14PRE18320032 (C.Z.B.), NIH R01 AI096837 (P.G. and V.N.) and NIH R01 AI077780 (V.N.). The work was also funded in part by the National Biomedical Computation Resource, NIH P41 GM103426, NIH Director's New Innovator Award Program DP2-OD007237 and through the National Science Foundation XSEDE Supercomputer Resources Grant RAC CHE060073N to R.E.A. S.P.H. was supported by the Interfaces Multi-Scale Analysis of Biological Structure and Function training grant NIH T32 EB009380.
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C.Z.B., V.N. and P.G. conceived the experiments. C.Z.B., A.J.B.-S. and T.B. carried out the structure determinations. C.Z.B. and A.J.B.-S. carried out the binding studies. S.P.H. and R.E.A. carried out and analysed the MD simulations. C.Z.B. and P.G. wrote the paper with input from all the authors.
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Supplementary information
Supplementary Information
Supplementary Tables 1–3, Supplementary Figures 1–16, legends for Supplementary Videos 1–5, Supplementary References (PDF 10446 kb)
Supplementary Video 1
R39 nook in M2-C4BPα1-2. (MOV 9313 kb)
Supplementary Video 2
R39 nook in M2 (F75A)-C4BPα1-2. (MOV 9328 kb)
Supplementary Video 3
C4BPα2 contacts in M2-C4BPα1-2. (MOV 13863 kb)
Supplementary Video 4
C4BPα2 contacts in M2 (K65A)-C4BPα1-2. (MOV 13986 kb)
Supplementary Video 5
C4BPα2 contacts in M2 (N66D)-C4BPα1-2. (MOV 13809 kb)
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Buffalo, C., Bahn-Suh, A., Hirakis, S. et al. Conserved patterns hidden within group A Streptococcus M protein hypervariability recognize human C4b-binding protein. Nat Microbiol 1, 16155 (2016). https://doi.org/10.1038/nmicrobiol.2016.155
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DOI: https://doi.org/10.1038/nmicrobiol.2016.155
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