Streptococcus pyogenesforms serotype and local environment-dependent inter-species protein complexes

Abstract

Streptococcus pyogenes is known to cause both mucosal and systemic infections in humans. In this study, we used a combination of quantitative and structural mass spectrometry techniques to determine the composition and structure of the interaction network formed between human plasma proteins and the surface of different S. pyogenes serotypes. Quantitative network analysis revealed that S. pyogenes form serotype-specific interaction networks that are highly dependent on the domain arrangement of the surface-attached M protein. Subsequent structural mass spectrometry analysis and computational modelling on one of the M proteins, M28 revealed that the network structure changes across different host microenvironments. We report that M28 binds secretory IgA via two separate binding sites with high affinity in saliva. During vascular leakage mimicked by increasing plasma concentrations in saliva, the binding of secretory IgA was replaced by binding of monomeric IgA and C4BP. This indicates that an upsurge of C4BP in the local microenvironment due to damage of the mucosal membrane drives binding of C4BP and monomeric IgA to M28. The results suggest that S. pyogenes has evolved to form microenvironment-dependent host-pathogen protein complexes to combat the human immune surveillance during both mucosal and systemic infections.

Distinct Serotypes of Streptococcal M Proteins Mediate Fibrinogen-Dependent Platelet Activation and Proinflammatory Effects

“…The serotypes that do not bind fibrinogen via the M protein might acquire fibrinogen to the surface via another surface protein in the M-related protein family ( 12 ). Recent work has shown that the emm 28, -49, and -89 bacterial serotypes from pattern E also bind significantly less fibrinogen to the bacterial surface than the emm 1, -3, and -5 serotypes ( 39 ) from pattern A-C. Distinct human plasma-interactome networks were observed for the serotypes from pattern A-C that acquired mainly fibrinogen and complement proteins, while pattern E serotypes acquired mainly apolipoproteins, immunoglobulins, and coagulation proteins.…”

“…The cloning, expression, and purification of the recombinant proteins used in this study have been described previously ( 39 ). For this study, the proteins were further purified using StrepTactin Sepharose (Sigma-Aldrich), washed with phosphate-buffered saline (PBS), and eluted with 5 mM biotin in PBS (Sigma-Aldrich).…”

“…The samples were centrifuged at 12,000 × g for 1 min, and the pellets were washed and resuspended in 50 μL D-PBS. These samples were prepared for data-independent acquisition mass spectrometry (DIA-MS) as previously described ( 39 ). In short, the proteins were denatured with 8 M urea and 100 mM ammonium bicarbonate (Sigma-Aldrich) at room temperature for 30 min, and the disulfide bonds were reduced using 0.5 mM Tris(2-carboxyethyl)phosphine hydrochloride (TCEP) (Sigma-Aldrich) at 37°C for 60 min and alkylated with 20 mM 2-iodoacetamide (IAA) (Sigma-Aldrich) for 30 min in the dark at room temperature.…”

“…All data analyses were stored and managed using openBIS ( 49 ). Raw data files were converted to mzXML using msconvert and analyzed using OpenSWATH 2.0.1 (revision c23217e) and a previously established spectral library ( 39 ). RT was calibrated using iRT peptides ( 48 ).…”