title: Isolation and functional characterization of the heavy and light chains of human tissue-type plasminogen activator.
abstract: Two-chain tissue-type plasminogen activator (t-PA), which consists of a heavy chain (Mr congruent to 38,000) and a light chain (Mr congruent to 31,000) connected by a disulfide bridge, was reduced with 2-mercaptoethanol and then air-reoxidized at a low protein concentration and carboxamidomethylated. The two chains were separated by means of zinc chelate-agarose, which was found to bind the light chain selectively. The light chain was fully active on the tripeptide substrate H-D-isoleucyl-L-prolyl-L-arginine p-nitroanilide (S-2288) and partially active on plasminogen. The plasminogen activator activity of the light chain was, in contrast to that of two-chain t-PA, not stimulated by fibrin or fibrinogen fragments. Fibrin-agarose chromatography of radiolabeled chains showed that only the heavy chain bound to fibrin. These results indicate that the active site-containing light chain in t-PA needs the heavy chain for fibrin stimulation of its plasminogen activator activity.
authors.forename: D C
authors.initials: DC
authors.lastname: Rijken
authors.forename: E
authors.initials: E
authors.lastname: Groeneveld
mesh_terms: D000596:Amino Acids / Q000032:analysis; D001665:Binding Sites; D002850:Chromatography, Gel; D003488:Cyanogen Bromide / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D005340:Fibrinogen / Q000378:metabolism; D005342:Fibrinolysis; D006801:Humans; D007460:Iodoacetamide / Q000494:pharmacology; D008623:Mercaptoethanol / Q000494:pharmacology; D013053:Spectrophotometry; D010959:Tissue Plasminogen Activator / Q000032:analysis* / Q000302:isolation & purification
chemical_list: D000596:Amino Acids; D008623:Mercaptoethanol; D005340:Fibrinogen; D010959:Tissue Plasminogen Activator; D003488:Cyanogen Bromide; D007460:Iodoacetamide