Structural determination of Streptococcus pyogenes M1 protein interactions with human immunoglobulin G using integrative structural biology

Streptococcus pyogenes (Group A streptococcus; GAS) is an important human pathogen responsible for mild to severe, life-threatening infections. GAS expresses a wide range of virulence factors, including the M family proteins. The M proteins allow the bacteria to evade parts of the human immune defenses by triggering the formation of a dense coat of plasma proteins surrounding the bacteria, including IgGs. However, the molecular level details of the M1-IgG interaction have remained unclear. Here, we characterized the structure and dynamics of this interaction interface in human plasma on the surface of live bacteria using integrative structural biology, combining cross-linking mass spectrometry and molecular dynamics (MD) simulations. We show that the primary interaction is formed between the S-domain of M1 and the conserved IgG Fc-domain. In addition, we show evidence for a so far uncharacterized interaction between the A-domain and the IgG Fc-domain. Both these interactions mimic the protein G-IgG interface of group C and G streptococcus. These findings underline a conserved scavenging mechanism used by GAS surface proteins that block the IgG-receptor (FcγR) to inhibit phagocytic killing. We additionally show that we can capture Fab-bound IgGs in a complex background and identify XLs between the constant region of the Fab-domain and certain regions of the M1 protein engaged in the Fab-mediated binding. Our results elucidate the M1-IgG interaction network involved in inhibition of phagocytosis and reveal important M1 peptides that can be further investigated as future vaccine targets.Author summary: Streptococcus pyogenes is a human specific pathogen causing both mild and invasive infections. It employs sophisticated mechanisms to evade and circumvent parts of the host’s immune defenses, in part via its major surface associated virulence factor, the family of M proteins. Of these, the M1 protein is the most prevalent serotype. The M1 protein creates a dense coat-like structure with multiple host proteins on the bacterial surface to disguise itself from opsonizing antibodies. It specifically interacts in a non-immune way with human immunoglobulin G (IgG) Fc-domains to disarm their receptor binding site. The molecular level details of this interaction have not been characterized. Here, we describe these interactions from minimally perturbed samples of human plasma adsorbed onto living bacteria using an integrative structural biology approach including cross-linking mass spectrometry, molecular modeling, and molecular dynamics simulations. We identify two distinct M1-peptides that bind IgGs and reveal the stability of these interactions. We show that both peptides block the Fc-receptor binding sites through capturing IgGs via their Fc-domains. These results highlight the importance of describing novel pathogen-derived peptides mediating host immune evasion as potential vaccine targets in future studies.