Low Molecular Weight Trypsin-Plasmin Inhibitors Isolated from Papain Treated Urinary Trypsin Inhibitor^[*]

Summary

Papain treatment of human urinary trypsin inhibitor (UTI₆₇; mol. wt. 43,000 by SDS-polyacrylamide gel electrophoresis, specific activity 1,897 U/mg protein) produced four new protease inhibitors, which were highly purified by gel chromatography on Sephadex G-100 and isoelectric focusing. The purified inhibitors (UTI₂₆, UTI₉-I, UTI₉-II, and UTI₉-III) were shown to be homogeneous by polyacrylamide disc gel electrophoresis, and had apparent molecular weights of 26,000, 9,000, 9,000, and 9,800, respectively, by sodium dodecyl sulfate gel electrophoresis. During enzymatic degradation of UTI₆₇, the amino acid compositions changed to more basic, and the isoelectric point increased from pH 2.0 (UTI₆₇) to pHs 4.4, 5.2, 6.6, and 8.3 (UTI₂₆, UTI₉-I, UTI₉-II, and UTI₉-III), respectively. Both the parent and degraded inhibitors had anti-plasmin activity as well as antitrypsin and anti-chymotrypsin activities. Much higher anti-plasmin/anti-trypsin and anti-plasmin/anti-chymotrypsin activities were observed in the degraded inhibitors than in the parent UTI₆₇. They competitively inhibited human plasmin with Ki values of 1.13 X 10_-7 - 2.12 X 10_-6 M (H-D-Val-Leu-Lys-pNA substrate). The reactions were very fast and the active site of the inhibitors to plasmin was thought to be different from that to trypsin or chymotrypsin.

Key words

Urinary trypsin inhibitor - Enzymatic modification of inhibitor - Plasmin inhibitor

^* A preliminary report of this work was presented at the XVIIth Congress of the International Society of Hematology, in Paris, July 23-29, 1978. Abstract p. 631.

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