The effect of substituted laminin A chain-derived peptides on the conformation and activation kinetics of plasminogen.
Conversion of the zymogen plasminogen (Pg) to the active enzyme plasmin is catalyzed by proteinases such as tissue-type plasminogen activator (t-PA). Interaction of Pg with small ligands such as lysine or macromolecular ligands such as fibrin induces a dramatic conformational change in the zymogen which enhances its efficacy as a t-PA substrate, thereby increasing catalytic efficiency of the activation reaction. We have previously demonstrated that a synthetic peptide derived from amino acids 2091-2108 of the laminin A chain (designated LamA2091-2108) can significantly enhance t-PA-catalyzed Pg activation. To probe the mechanism of this stimulatory reaction, we have determined the effect of substituted LamA2091-2108 derivatives on Pg activation by t-PA. Substitution of charged residues in LamA2091-2108 with neutral amino acids decreases the kcat/Km observed in the presence of native LamA2091-2108. Furthermore, fluorescence-quenching experiments demonstrate that whereas LamA2091-2108 alters the solvent accessibility of Pg Trp residues, charge-substituted peptides have little effect on Pg conformation. These data suggest that LamA2091-2108 stimulates Pg activation by inducing a conformational change in the zymogen similar to that observed upon binding of other ligands such as lysine and fibrin.
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Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710.
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D000595:Amino Acid Sequence; D005341:Fibrinolysin / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D007797:Laminin / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D010958:Plasminogen / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D013329:Structure-Activity Relationship; D010959:Tissue Plasminogen Activator / Q000378:metabolism
D007797:Laminin; D010446:Peptide Fragments; C084098:laminin A; D010958:Plasminogen; D010959:Tissue Plasminogen Activator; D005341:Fibrinolysin