Plasmin's peptide-binding specificity: characterization of ligand sites in alpha 2-antiplasmin.
The basis for specific binding of plasmin to alpha 2-antiplasmin (AP) was analyzed by preparing overlapping synthetic peptides of 11, 17, 18, 19, 26, 33, and 40 amino acid residues corresponding to the carboxy-terminal sequence of AP. Affinities of the peptides for plasmin were estimated by competitive inhibition of the association of AP with plasmin. Dissociation constants with increasing peptide length were: 200, 54, 19, 18, 9.8, 4.7, and 2.8 microM, respectively. Peptides blocked binding sites on plasmin, not the catalytic site, as evidenced by lack of effect on the hydrolysis of chromogenic substrates. Substituting arginine for lysine at the carboxy-terminus or the 17th residue from the carboxy-terminus decreased the affinity of peptides for plasmin 9-fold and 5-fold, respectively, implicating these lysine residues of AP as major ligand sites for plasmin. Several stepwise increases in affinity of peptides for plasmin as peptide length increased up to 40 residues suggest contributions by additional sites, possibly other lysine residues. A potential plasmin binding site in fibrin, analogous to that in AP, is identified by affinity for plasmin of synthetic peptides corresponding to part of the alpha-chain ending with residue 207. To explain these data, we propose that plasmin recognizes physiological ligands by binding two or more lysine residues which are optimally presented to favor simultaneous interaction with separate lysine-binding site in plasmin.
Hortin
G L
GL
Department of Pediatrics, Washington University School of Medicine, St. Louis MO 63110.
Trimpe
B L
BL
Fok
K F
KF
D001120:Arginine / Q000378:metabolism; D001665:Binding Sites; D005337:Fibrin / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism*; D008024:Ligands; D008239:Lysine / Q000378:metabolism; D010449:Peptide Mapping; D010455:Peptides / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D011485:Protein Binding; D000979:alpha-2-Antiplasmin / Q000378:metabolism*
D008024:Ligands; D010455:Peptides; D000979:alpha-2-Antiplasmin; D005337:Fibrin; D001120:Arginine; D010744:Phosphoric Monoester Hydrolases; D005341:Fibrinolysin; D008239:Lysine