Acid-stable trypsin-plasmin inhibitors formed enzymatically from plasma precursor protein.
Enzymatic formation of acid-stable trypsin-plasmin inhibitors (ASTPIs) in human plasma with several proteinases, particularly SH-proteinases, was demonstrated. The maximal activity obtained with bromelain was 40 U/ml plasma, which corresponded to about a 10-fold increase as compared to the untreated control plasma (4.2 U/ml). Gel filtration revealed at least two ASTPI activity peaks of molecular weight 16,000 (main peak) and 8000 (minor peak). The main ASTPI was further purified by trypsin-Sepharose affinity chromatography, isoelectric focusing and gel filtration on Sephadex G-75 superfine. The purified inhibitor was found to be identical to the active fragment of plasma ASTPI or urinary trypsin inhibitor (UTI) formed by bromelain treatment. It had an isoelectric point (pI) of 3.7, a molecular weight of 16,000 by SDS-polyacrylamide gel electrophoresis and was a glycine- and glutamic acid-rich protein lacking histidine. The NH2-terminal amino acid sequence was H2N-(Lys)-Glu-Asp-Ser-X-Gln-Leu-Gly-Tyr-Ser-Ala-Gly-Pro-X-Met-Gly-Met-Th r-X-Arg - Tyr-Phe-Tyr-... COOH, which was homologous to the Lys22-Met36 part (or Glu23-Met36 part; 30% of the total) of the plasma ASTPI or UTI molecule (molecular weight 70,000-80,000 by gel filtration). The purified ASTPI displayed the same antigenicity as UTI and exerted strong inhibitory effects on trypsin, chymotrypsin and plasmin amidolysis, but had a much lesser effect on plasmin fibrinolysis. It also strongly inhibited non-plasmic fibrinolysis with human leukocyte proteinase and earthworm proteinase.
Sumi
H
H
Department of Physiology, Miyazaki Medical College, Okayama, Japan.
Yoshida
E
E
Hamada
H
H
Mihara
H
H
D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005341:Fibrinolysin / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D005342:Fibrinolysis; D006801:Humans; D006863:Hydrogen-Ion Concentration; D005779:Immunodiffusion; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000378:metabolism; D011498:Protein Precursors / Q000378:metabolism; D014361:Trypsin Inhibitors / Q000302:isolation & purification*
D001798:Blood Proteins; D011498:Protein Precursors; D014361:Trypsin Inhibitors; D010447:Peptide Hydrolases; D005341:Fibrinolysin