1 Fibrinogen is a ligand for the S. aureus MSCRAMM Bbp (Bone sialoprotein-binding protein ) Vanessa Vazquez ‡, ¶ , Xiaowen Liang ‡ , Jenny K. Horndahl ‡ , Vannakambadi K. Ganesh ‡ , Emanuel Smeds ‡ , Timothy J. Foster § , Magnus Hook ‡** ‡ Center for Infectious and Inflammatory Diseases, Institute of Biosciences and Technology, Texas A&M University System Health Science Center, Houston, TX 77030. ¶ Graduate School of Biomedical Sciences, University of Texas Health Science Center, Houston, TX 77030. § Moyne Institute of Preventive Medicine, Department of Microbiology, Trinity College, Dublin, Ireland. ** To whom correspondence should be addressed: Center for Infectious and Inflammatory Diseases, Texas A&M University System Health Science Center, Institute of Biosciences and Technology, 2121 West Holcombe Blvd., Suite 603, Houston, TX 77030, Tel.: 713-677-7551; Fax: 713-677-7576; E-mail: mhook@ibt.tamhsc.edu . Running Title: Fibrinogen is a ligand for the S. aureus MSCRAMM Bbp ABSTRACT MSCRAMMs (microbial surface components recognizing adhesive matrix molecules) are bacterial surface proteins mediating adherence of the microbes to components of the extracellular matrix of the host. On Staphylococci the MSCRAMMs often have multiple ligands. Consequently we hypothesized that the S. aureus MSCRAMM Bbp (bone sialoprotein-binding protein) might recognize host molecules other than the identified bone protein. A ligand screen revealed that Bbp binds human fibrinogen (Fg) but not Fg from other mammals. We have characterized the interaction between Bbp and Fg. The binding site of Bbp was mapped to residues 561-575 in the Fg Aα chain using recombinant Fg chains and truncation mutants in Far Western blots and solid phase binding assays. Surface plasmon resonance was used to determine the affinity of Bbp for Fg. The interaction of Bbp with Fg peptides corresponding to the mapped residues was further characterized using isothermal titration calorimetry. In addition, Bbp expressed on the surface of bacteria mediated adherence to immobilized Fg Aα . Also, Bbp interferes with thrombin-induced Fg coagulation. Together these data demonstrate that human Fg is a ligand for Bbp and that Bbp can manipulate the biology of the Fg ligand in the host. INTRODUCTION Staphylococcus aureus uses secreted and cell-surface associated virulence factors to cause disease ranging from mild skin infections like folliculitis and impetigo to life-threatening illnesses like sepsis, and pneumonia (1). MSCRAMMs (Microbial surface components recognizing adhesive matrix molecules) are surface proteins used by bacteria to interact with host molecules such as collagen, fibronectin, and fibrinogen (Fg). The Sdr proteins are a subset of putative staphylococcal MSCRAMMs, covalently anchored to the cell- wall and characterized by a segment composed of repeated serine-aspartate (SD) dipeptides. The Sdr proteins have similar structural organization where the N-terminal ligand-binding A region can be further divided into three sub-domains (N1, N2, and N3), where N2 and N3 adopt IgG-like folds. The A-region is often followed by a B region that consists of repeated β-sandwich domains. The carboxy-terminal section of the proteins contains the serine-aspartate repeats followed by motifs required for cell-wall anchoring (2). A dynamic ligand binding mechanism called the “dock, lock and latch” was revealed by biochemical and structural studies of the fibrinogen binding S. epidermidis MSCRAMM SdrG (3). SdrG binds to a linear sequence in the N-terminus of the Bβ chain of human Fg The SdrG binding sequence includes the thrombin cleavage site, and the MSCRAMM inhibits thrombin-catalyzed release of fibrinopeptide B and fibrin formation (3, 4). Binding is initiated by the “docking” of the ligand peptide into the trench formed between the N2 and N3 IgG-domains. http://www.jbc.org/cgi/doi/10.1074/jbc.M110.214981 The latest version is at JBC Papers in Press. Published on June 3, 2011 as Manuscript M110.214981 Copyright 2011 by The American Society for Biochemistry and Molecular Biology, Inc. at IReL (Trinity College Dublin), on June 15, 2011 www.jbc.org Downloaded from