Structural proteomics, electron cryo-microscopy and structural modeling approaches in bacteria–human protein interactions

Chowdhury, Sounak; Happonen, Lotta; Khakzad, Hamed; Malmström, Lars; Malmström, Johan

doi:10.1007/s00430-020-00663-5

Cited by 15 publications

(13 citation statements)

References 103 publications

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“…To cause invasive infections, GAS expresses a wide range of virulence factors to evade human defense mechanisms. These virulence factors mostly consist of secreted or surface-associated proteins that target proteins and protein complexes of the innate and adaptive immune systems [3][4][5].…”

Section: Introductionmentioning

confidence: 99%

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

et al. 2021

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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.

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Section: Introductionmentioning

confidence: 99%

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

et al. 2021

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“…Here, we elucidate how in vivo cross-linking of human plasma to the surface of live bacteria can provide detailed information of protein-protein complexes in their native environment, with possible implications for the studies of conformational differences between in vitro crystallized and electron cryo-microscopically imaged and reconstructed macromolecular complexes and in vivo cross-linked ones. The approach detailed here should also open up new avenues for the future integration of XL-MS data with in vivo prokaryotic electron cryo-tomographic imaging and model reconstruction of host-pathogen complexes (Nans et al, 2014(Nans et al, , 2015Chowdhury et al, 2020).…”

Section: Discussionmentioning

confidence: 99%

In vivo Cross-Linking MS of the Complement System MAC Assembled on Live Gram-Positive Bacteria

Khakzad

Happonen²,

Nhieu³

et al. 2021

Front. Genet.

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Protein–protein interactions are central in many biological processes, but they are challenging to characterize, especially in complex samples. Protein cross-linking combined with mass spectrometry (MS) and computational modeling is gaining increased recognition as a viable tool in protein interaction studies. Here, we provide insights into the structure of the multicomponent human complement system membrane attack complex (MAC) using in vivo cross-linking MS combined with computational macromolecular modeling. We developed an affinity procedure followed by chemical cross-linking on human blood plasma using live Streptococcus pyogenes to enrich for native MAC associated with the bacterial surface. In this highly complex sample, we identified over 100 cross-linked lysine–lysine pairs between different MAC components that enabled us to present a quaternary model of the assembled MAC in its native environment. Demonstrating the validity of our approach, this MAC model is supported by existing X-ray crystallographic and electron cryo-microscopic models. This approach allows the study of protein–protein interactions in native environment mimicking their natural milieu. Its high potential in assisting and refining data interpretation in electron cryo-tomographic experiments will be discussed.

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“…Bacterial pathogens have evolved to express a multitude of virulence factors on their surface to establish versatile host-pathogen protein-protein interactions (HP-PPI) 1 . These interactions range from binary interactions between two proteins to the formation of multimeric interspecies protein complexes that enable bacterial pathogens to hijack and re-wire molecular host systems to circumvent host immune defenses.…”

Section: Introductionmentioning

confidence: 99%

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

Chowdhury

Khakzad

Bergdahl

et al. 2021

Preprint

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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.

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Structural proteomics, electron cryo-microscopy and structural modeling approaches in bacteria–human protein interactions

Cited by 15 publications

References 103 publications

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

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

In vivo Cross-Linking MS of the Complement System MAC Assembled on Live Gram-Positive Bacteria

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

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