title: Kringle 2 mediates high affinity binding of plasminogen to an internal sequence in streptococcal surface protein PAM.
abstract: Many cells express receptors for plasminogen (Pg), although the responsible molecules in most cases are poorly defined. In contrast, the group A streptococcal surface protein PAM contains a domain with two 13-amino acid residue long repeated sequences (a1 and a2) responsible for Pg binding. Here we identify the region in Pg that interacts with PAM. A radiolabeled proteolytic plasminogen fragment containing the first three kringles (K1-K3) interacted with streptococci expressing PAM or a chimeric surface protein harboring the a1a2 sequence. In contrast, plasminogen fragments containing kringle 4 or kringle 5 and the activable serine proteinase domain failed to bind to PAM-expressing group A streptococci. A synthetic and a recombinant polypeptide containing the a1a2 sequence both bound to immobilized recombinant K2 (rK2) but not to rK1 or rK3. The interaction between the a repeat region and rK2 was reversible, and rK2 completely blocked the binding of Pg to the a1a2 region. The binding of the a repeat containing polypeptide to K2 occurred with an equilibrium association constant of 4.5 x 10(7) M-1, as determined by surface plasmon resonance, a value close to that (1.6 x 10(7) M-1) calculated for the a1a2-Pg interaction. Inhibition experiments suggested involvement of the lysine-binding site of K2 in the interaction. These data demonstrate that K2 contains the major Pg-binding site for PAM, providing the first well defined example of an interaction between an internal Pg-binding region in a protein and a single kringle domain.
authors.affiliation: Department of Laboratory Medicine, University of Lund, S-22362 Lund, Sweden.
authors.forename: A C
authors.initials: AC
authors.lastname: Wistedt
authors.forename: H
authors.initials: H
authors.lastname: Kotarsky
authors.forename: D
authors.initials: D
authors.lastname: Marti
authors.forename: U
authors.initials: U
authors.lastname: Ringdahl
authors.forename: F J
authors.initials: FJ
authors.lastname: Castellino
authors.forename: J
authors.initials: J
authors.lastname: Schaller
authors.forename: U
authors.initials: U
authors.lastname: Sjöbring
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010958:Plasminogen / Q000737:chemistry* / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013297:Streptococcus pyogenes / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D017931:DNA Primers; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C084529:plasminogen-binding protein, bacteria; D010958:Plasminogen