Active but inoperable thrombin is accumulated in a plasma protein layer surrounding Streptococcus pyogenes

Summary

Activation of thrombin is a critical determinant in many physiological and pathological processes including haemostasis and inflammation. Under physiological conditions many of these functions are involved in wound healing or eradication of an invading pathogen. However, when activated systemically, thrombin can contribute to severe and life-threatening conditions by causing complications such as multiple multi-organ failure and disseminated intravascular coagulation. In the present study we investigated how the activity of thrombin is modulated when it is bound to the surface of Streptococcus pyogenes. Our data show that S. pyogenes bacteria become covered with a proteinaceous layer when incubated with human plasma, and that thrombin is a constituent of this layer. Though the coagulation factor is found attached to the bacteria with a functional active site, thrombin has lost its capacity to interact with its natural substrates and inhibitors. Thus, the interaction of bacteria with human plasma renders thrombin completely inoperable at the streptococcal surface. This could represent a host defense mechanism to avoid systemic activation of coagulation which could be otherwise induced when bacteria enter the circulation and cause systemic infection.

Keywords

Thrombin - fibrinogen - proteolysis - coagulation - Streptococcus pyogenes

• References

• 1 Engelmann B, Massberg S. Thrombosis as an intravascular effector of innate immunity. Nat Rev Immunol 2013; 13: 34-45.
• 2 Adams TE, Huntington JA. Thrombin-cofactor interactions: structural insights into regulatory mechanisms. Arterioscl Thromb Vasc Biol 2006; 26: 1738-1745.
• 3 Huntington JA. Molecular recognition mechanisms of thrombin. J Thromb Haemost 2005; 3: 1861-1872.
• 4 Rose T, Di Cera E. Three-dimensional modeling of thrombin-fibrinogen interaction. J Biol Chem 2002; 277: 18875-18880.
• 5 Hall SW, Nagashima M, Zhao L. et al. Thrombin interacts with thrombomodulin, protein C, and thrombin-activatable fibrinolysis inhibitor via specific and distinct domains. J Biol Chem 1999; 274: 25510-25516.
• 6 Huntington JA. Thrombin inhibition by the serpins. J Thromb Haemost 2013; 11 (Suppl. 01) 254-264.
• 7 Rehman AA, Ahsan H, Khan FH. alpha-2-Macroglobulin: a physiological guardian. J Cell Physiol 2013; 228: 1665-1675.
• 8 Cole JN, Barnett TC, Nizet V. et al. Molecular insight into invasive group A streptococcal disease. Nature Rev Microbiol 2011; 9: 724-736.
• 9 Semeraro N, Ammollo CT, Semeraro F. et al. Sepsis, thrombosis and organ dys-function. Thromb Res 2012; 129: 290-295.
• 10 Marx PF, Brondijk TH, Plug T. et al. Crystal structures of TAFI elucidate the inactivation mechanism of activated TAFI: a novel mechanism for enzyme auto-regulation. Blood 2008; 112: 2803-2809.
• 11 Nordin SL, Andersson C, Bjermer L. et al. Midkine is part of the antibacterial activity released at the surface of differentiated bronchial epithelial cells. J Innate Immun 2013; 5: 519-530.
• 12 Herwald H, Mörgelin M, Dahlbäck B. et al. Interactions between surface proteins of Streptococcus pyogenes and coagulation factors modulate clotting of human plasma. J Thromb Haemost 2003; 1: 284-291.
• 13 Marx PF, Plug T, Havik SR. et al. The activation peptide of thrombin-activatable fibrinolysis inhibitor: a role in activity and stability of the enzyme?. J Thromb Haemost 2009; 7: 445-452.
• 14 Sjöholm K, Karlsson C, Linder A. et al. A comprehensive analysis of the Streptococcus pyogenes and human plasma protein interaction network. Mol Bio Syst 2014; 10: 1698-1708.
• 15 Kihlberg BM, Cooney J, Caparon MG. et al. Biological properties of a Streptococcus pyogenes mutant generated by Tn916 insertion in mga. Microb Pathog 1995; 19: 299-315.
• 16 Loof TG, Mörgelin M, Johansson L. et al. Coagulation, an ancestral serine pro-tease cascade, exerts a novel function in early immune defense. Blood 2011; 118: 2589-2598.
• 17 Wood JP, Silveira JR, Maille NM. et al. Prothrombin activation on the activated platelet surface optimizes expression of procoagulant activity. Blood 2011; 117: 1710-1718.
• 18 Krishnaswamy S. The transition of prothrombin to thrombin. J Thromb Hae-most 2013; 11 (Suppl. 01) 265-276.
• 19 Smith SA. The cell-based model of coagulation. J Vet Emerg Crit Care 2009; 19: 3-10.
• 20 Nickel KF, Renne T. Crosstalk of the plasma contact system with bacteria. Thromb Res 2012; 130 (Suppl. 01) S78-83.
• 21 Martin FA, Murphy RP, Cummins PM. Thrombomodulin and the vascular endothelium: insights into functional, regulatory, and therapeutic aspects. Am J Physiol Heart Circ Physiol 2013; 304: H1585-1597.
• 22 Myles T, Le Bonniec BF, Betz A. et al. Electrostatic steering and ionic tethering in the formation of thrombin-hirudin complexes: the role of the thrombin anion-binding exosite-I. Biochemistry 2001; 40: 4972-4979.
• 23 Baglin TP, Carrell RW, Church FC. et al. Crystal structures of native and thrombin-complexed heparin cofactor II reveal a multistep allosteric mechanism. Proc Natl Acad Sci USA 2002; 99: 11079-11084.
• 24 O’Keeffe D, Olson ST, Gasiunas N. et al. The heparin binding properties of heparin cofactor II suggest an antithrombin-like activation mechanism. J Biol Chem 2004; 279: 50267-50273.
• 25 Li W, Johnson DJ, Esmon CT. et al. Structure of the antithrombin-thrombin-heparin ternary complex reveals the antithrombotic mechanism of heparin. Nature Struct Mol Biol 2004; 11: 857-862.
• 26 Kettner C, Shaw E. D-Phe-Pro-ArgCH2C1-A selective affinity label for thrombin. Thromb Res 1979; 14: 969-973.
• 27 Carapetis JR, Steer AC, Mulholland EK. et al. The global burden of group A streptococcal diseases. Lancet Infect Dis 2005; 5: 685-694.
• 28 Boyd JH, Russell JA, Fjell CD. The meta-genome of sepsis: host genetics, pathogens and the acute immune response. J Innate Immun 2014; 6: 272-283.
• 29 Herwald H, Egesten A. A farewell to arms: streptococcal strategies to cope with innate immunity. J Innate Immun 2014; 6: 561-562.
• 30 de Jong HK, van der Poll T, Wiersinga WJ. The systemic pro-inflammatory response in sepsis. J Innate Immun 2010; 2: 422-430.
• 31 Pahlman LI, Mörgelin M, Kasetty G. et al. Antimicrobial activity of fibrinogen and fibrinogen-derived peptides--a novel link between coagulation and innate immunity. Thromb Haemost 2013; 109: 930-939.
• 32 Ly D, Taylor JM, Tsatsaronis JA. et al. Plasmin(ogen) acquisition by group A Streptococcus protects against C3b-mediated neutrophil killing. J Innate Immun 2014; 6: 240-250.
• 33 Courtney HS, Hasty DL, Dale JB. Anti-phagocytic mechanisms of Streptococcus pyogenes: binding of fibrinogen to M-related protein. Mol Microbiol 2006; 59: 936-947.
• 34 Downing MR, Bloom JW, Mann KG. Comparison of the inhibition of thrombin by three plasma protease inhibitors. Biochemistry 1978; 17: 2649-2653.
• 35 Harpel PC. Immunoassay of alpha 2-macroglobulin-enzyme complexes. Ann NY Acad Sci 1983; 421: 134-142.
• 36 Pochon F, Steinbuch M. Interaction of alpha 2-macroglobulin-bound thrombin with hirudin. FEBS Lett 1984; 177: 109-111.
• 37 Pochon F, Steinbuch M, Lambin P. et al. Structural arrangement in the alpha 2-macroglobulin--thrombin complex. FEBS Lett 1983; 161: 51-54.
• 38 Doyle MF, Haley PE. Meizothrombin: active intermediate formed during prothrombinase-catalysed activation of prothrombin. Methods Enzymol 1993; 222: 299-312.
• 39 Fischer BE, Schlokat U, Himmelspach M. et al. Binding of hirudin to meizothrombin. Protein Engin 1998; 11: 715-721.