title: Interactions between factor XIII and the alphaC region of fibrinogen.
abstract: Fibrinogen αC residues 242-424 have been shown to have a major regulatory role in the activation of factor XIII-A(2)B(2) (FXIII-A(2)B(2)); however, the interactions underpinning this enhancing effect have not been determined. Here, we have characterized the binding of recombinant (r)FXIII-A subunit and FXIII-A(2)B(2) with fibrin(ogen) and fibrin αC residues 233-425. Using recombinant truncations of the fibrin αC region 233-425 and surface plasmon resonance, we found that activated rFXIII-A bound αC 233-425 (K(d) of 2.35 ± 0.09 μM) which was further localized to αC 389-403. Site-directed mutagenesis of this region highlighted Glu396 as a key residue for binding of activated rFXIII-A. The interaction was specific for activated rFXIII-A and depended on the calcium-induced conformational change known to occur in rFXIII-A during activation. Furthermore, nonactivated FXIII-A(2)B(2), thrombin-cleaved FXIII-A(2)B(2), and activated FXIII-A(2)B(2) each bound fibrin(ogen) and specifically αC region 371-425 with high affinity (K(d) < 35 nM and K(d) < 31 nM, respectively), showing for the first time the potential involvement of the αC region in binding to FXIII-A(2)B(2). These results suggest that in addition to fibrinogen γ' chain binding, the fibrin αC region also provides a platform for the binding of FXIII-A(2)B(2) and FXIII-A subunit.
authors.affiliation: Division of Cardiovascular and Diabetes Research, Leeds Institute for Genetics Health and Therapeutics, Faculty of Medicine and Health, University of Leeds, Leeds, United Kingdom.
authors.forename: Kerrie A
authors.initials: KA
authors.lastname: Smith
authors.forename: Penelope J
authors.initials: PJ
authors.lastname: Adamson
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Pease
authors.forename: Jane M
authors.initials: JM
authors.lastname: Brown
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Balmforth
authors.forename: Paul A
authors.initials: PA
authors.lastname: Cordell
authors.forename: Robert A S
authors.initials: RA
authors.lastname: Ariëns
authors.forename: Helen
authors.initials: H
authors.lastname: Philippou
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Grant
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D002118:Calcium / Q000378:metabolism; D005176:Factor XIII / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005340:Fibrinogen / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs / Q000235:genetics; D025941:Protein Interaction Mapping; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D050505:Mutant Proteins; D010446:Peptide Fragments; D020033:Protein Isoforms; D011994:Recombinant Proteins; C098000:fibrinogen alphaC; D005340:Fibrinogen; D005176:Factor XIII; D002118:Calcium