An Antiproteolytic and Antifibrinolytic Preparation Obtained from the Potato by Means of Chromatography on SE-Sephadex C-50

K Worowski

doi:10.1055/s-0038-1647731

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1974; 32(02/03): 617-632
DOI: 10.1055/s-0038-1647731

Original Article

Schattauer GmbH

An Antiproteolytic and Antifibrinolytic Preparation Obtained from the Potato by Means of Chromatography on SE-Sephadex C-50

K Worowski

¹Department of Biochemistry, Institute Physiology and Biochemistry, Medical School, 15-952 Białystoik 8, Poland

› Author Affiliations

Abstract

Summary

A crude preparation of a proteolytic enzyme inhibitor from potatoes was purified by means of column chromatography with SE-Sephadex C-50 using a stepwise gradient of concentration and pH when eluting proteins from the column. The inhibitor preparation thus obtained is stable for a 28 day period of storage and resistant to the action of various physical and chemical factors harmful to the spatial structure of proteins. The potato protease inhibitor markedly inhibits the activity of trypsin, chymotrypsin, plasmin and alkaline proteases from Aspergillus flavus. The inhibition of pronase and a mixture of acid cellular proteases by this inhibitor is considerably less. Pepsin, cathepsin D, and collagenase are not affected by this inhibitor. The potato inhibitor has a marked inhibitory effect on plasmin fibrinolytic activity both in purified systems and in the plasma euglobulin fraction. It also protects the fibrinogen from the action of this enzyme. It does not, however, inhibit plasminogen activation under the influence of either streptokinase or urokinase.

Full Text

References

References
1 Barret A. J, Dingle J. T. (Eds.) 1971. Tissue proteinases. North-Holland Publishing Co; Amsterdam, London.:
2 Bauer C. A, Löfqwist B. 1973; Studies of the heterogenity of streptomyces griseus protease. Isolation and characterization of an alkaline serine protease, from commercial pronase-P derived from Streptomyces griseus K 1. Acta chimica Scandinavica 27: 3147.
3 Belitz H. D, Kaiser K. P, Santaritjs K. 1971; Trypsin and a-chymotrypsin inhibitors from potatoes : isolation and some properties. Biochemical and Biophysical Research Communications 42: 420.
4 Brendel W, Haberland G. L. (Eds.) 1972. New Aspects of Trasylol Therapy. Report on an International Symposium, Wiesbaden, 15-16 november 1971. F.K. Schattauer Verlag; Stuttgart, New York.:
5 Crafoord C. (Ed.) 1967. The Clinical Significance of Proteinase Inhibitors. Symposium held at Falsterbohus, 1966. Acta Chirurgica Scandinavica. 378 (Suppl.).
6 Farbiszewski R, Worowski K, Rzeczycki W. 1972; The application of a modified Sakaguchi’s method for quantitative estimation of protein-bound arginine. Chem. Analyt 17: 133.
7 Fenney R. E, Means G. E, Bigler J. C. 1969; Inhibition of human trypsin, plasmin and thrombin by naturally occurring inhibitors of proteolytic enzymes. Journal of Biological Chemistry 244: 1957.
8 Folin O, Ciocalteu V. 1927; Tyrosine and tryptophan determination in proteins. Journal of Biological Chemistry 73: 627.
9 Fritz H, Tschesche H. (Eds.) 1971. International Research Conference on Proteinase Inhibitors, Munich, 1970. W. de Gruyter; Berlin, New York.:
10 Green N. M. (1953a): Competition among trypsin inhibitors. Journal of Biological Chemistry 205: 535.
11 Green N. M, Work E. (1953b): Pancreatic trypsin inhibitor. Biochemical Journal 54: 347.
12 Hochstrasser K, Werle E, Siegelmattn R, Schwarz S. 1969; Isolierung und Charakterisierung einiger polyvalenter Proteaseinhibitoren aus Solanum tuberosum. Zeitschrift für physiologische Chemie 350: 897.
13 Iodice A. A, Leong Y, Weinstock I. M. 1966; Separation of cathepsins A and D of scelet muscle. Archives of Biochemistry and Biophysics 117: 477.
14 Iwasaki T, Kiyohara T, Yoshikawa M. 1971; Purification and partial charakterization of two different types of proteinase inhibitors (inhibitors Il-a and Il-b) from potato. Journal of Biochemistry 70: 817.
15 Iwasaki T, Kiyohara T, Yoshikawa M. 1973; Identification of the reactive site of potato proteinase inhibitor Il-a. Journal of Biochemistry 73: 1039.
16 Janicki J, Warchalewski J, Skupin J, Kowalczyk J. 1970; Trypsin inhibitors of plant origin (in Polish). Posçtpy Biochemii 16: 101.
17 Kalckar H. M. 1947; Differential spectrophotometry of purine compounds by means of specific enzymes. III. Studies of the enzymes of purine metabolism. Journal of Biological Chemistry 167: 461.
18 Keil B, Sormova Z. (Eds.) 1959. Laboratorni technika biochemie. Nakl. CSAV, Praha. 747.
19 Kekwick R. A, Mackay M. E, Nance M. H, Record B. R. 1955; The purification of human fibrinogen. Biochemical Journal 60: 671.
20 Kowalski E, Kopec M, Niewiarowski S. 1959; An evaluation of the euglobulin method for the determination of fibrinolysis. Journal of Clinical Pathology 12: 215.
21 Latallo Z. S. 1966; Conversion of fibrinogen into fibrin and influence of fibrinogen degradation products on this process (in Polish). Postçpy Higieny i Medycyny Doswiadczalnej 20: 657.
22 Lineaweaver H, Burk D. 1934; The determination of enzyme dissociation constants. Journal of the American Chemistry Society 56: 658.
23 Mansfeld Y, Rybäk M, Horakova Z, Hladovec J. 1960; Die antitryptische* antifibrinolytische und antiphlogistische Aktivität natürlicher Proteasen-inhibitoren. Zeitschrift für physiologische Chemie 318: 6.
24 Melville J. C, Ryan C. A. 1972; Chymotrypsin inhibitor I from potatoes. Journal of Biological Chemistry 247: 3445.
25 Mikes ü, Worowski K, Turkova J. 1973; Polymorphism of alkaline protease from Aspergillus flavus. Collection Czechoslov. Chem. Commun 38: 3339.
26 Narahashi Y, Shibuya K, Yanagita M. 1968; Studies on proteolytic enzymes (Pronase) of Streptomyces griseus K-l. Journal of Biochemistry 64: 427.
27 Page R. C, Benedit E. P. 1968; Molecular diseases of connective and vascular tissues. III. The aldehyde content of normal and Lathyritic soluble collagen. Laboratory Investigations 18: 124.
28 Prockop D. W, TJndenfriend S. 1960; A specific method for the analysis of hydroxyproline in tissues and urine. Analytical Biochemistry 01: 228.
29 Rabek Y, Mansfeld Y. 1963; Gel filtration of protease inhibitors from potatoes. Experientia 19: 151.
30 Reisfeld R. A, Lewis A. F, Williams D. E. 1962; Disk electrophoresis of basic proteins and peptides on polyacrylamide gels. Nature 195: 281.
31 Reisner W, Brunnen R. 1971; Zur Kinetik von Serinproteasen aus Mikroorganismen. Zeitschrift für Physiologische Chemie 352: 1377.
32 Sawaryn A. 1969; Naturally occuring inhibitors of proteolytic enzymes (in Polish). Postçpy Higieny i Medycyny Doswiadczalnej 23: 787.
33 Uhlig H, Kleine R. 1974; Natürlich vorkommende Protein-proteinase-Inhibitoren aus tierischen und pflanzlichen Organismen. Die Pharmazie 29: 81.
34 Yogel R, Trautschold I, Werle E. 1968. Natural Proteinase. Inhibitors. Academic Press; New York, London.:
35 Werle E, Zickgraf-Rüdel G. r. 1972; Natural proteinase inhibitors distribution, specificity, mode of action and physiological significance. Zeitschrift für Klinische Chemie und Klinische Biochemie 10: 139.
36 Worowski K. 1974; Antiproteolytic and haemoagglutination activity of various varieties of potato (Solanum tuberosum) (in Polish). Bromat. Chem. Toksykol 07: 223.
37 Worowski K. A simple method of purification of cathepsin D. in preparation.
38 Worowski K, Bankowski E, Szmitkowski M. 1970; Studies on red cell agglutinating and antitrypsin activities of proteins from the potato. Bulletin of the Polon. Academy of Sciences, serie Science of Biology 18: 739.
39 Worowski K, Rzeczycki W, Mariak R. T. (1972/73): The inhibition of proteolytic enzymes by protamine in dependence on the type of substrate. Enzyme 14: 286.
40 Worowski K, Szmitkowski M. 1972; Isolation of proteolytic enzymes inhibitor from potato (in Polish). Roczniki Technol. Chem. Zywn 22: 261.