Download PDF
Thromb Haemost 1963; 10(02): 390-399
DOI: 10.1055/s-0038-1654792
Originalarbeiten — Original Article — Travaux Originaux
Schattauer GmbH

Proteolytic Digestion of Purified Bovine Prothrombin: Formation of Active Fragments[*]

Garson H. Tishkoff
1   Department of Medicine, UCLA Medical Center, Los Angeles, Cal
› Author Affiliations
Further Information

Publication History

Publication Date:
22 June 2018 (online)

Also available at
    Permissions and Reprints

Summary

Purified bovine prothrombin was subjected to proteolysis by a number of highly purified proteinases. Under specified conditions, papain and trypsin degraded prothrombin with the formation of protein fragments. These fragments were still capable of being biologically activated to thrombin by tissue thromboplastin, Factor V and calcium. Extensive enzymatic degradation of the protein moiety was achieved as evidenced by large quantities of peptide and carbohydrate rendered soluble in trichloracetic acid. Digestion of prothrombin with Nagarse, Pronase and plasmin at an identical enzyme-substrate ratio resulted in almost total inactivation of prothrombin although the extent of degradation was similar to that observed with papain. Chymotrypsin resulted in complete inactivation of clotting activities. It was also established that digested prothrombin was associated with the non-dialysable fraction. These and related observations suggest that prothrombin activity may be associated with a polypeptide fragment contained within the parent glycoprotein structure.

* This study supported by a grant, H-06021, from the National Institutes of Health.


 

  • References

  • 1 Alexander B, Pechet L, Kliman A. Proteolysis, fibrinolysis and coagulation. Circulation 26: 596 1962;
    CrossrefPubMedSearch in Google Scholar
  • 2 Bachmann F, Duckert F, Koller F. The Stuart-Prower factor assay and its clinical significance. Thrombos. Diathes. haemorrh. (Stuttg) 02: 24 1958;
    PubMedSearch in Google Scholar
  • 3 Eagle H. Recent advances of the blood coagulation problem. Medicine 16: 95 1937;
    CrossrefPubMedSearch in Google Scholar
  • 4 Esnouf M. P, Williams W. J. Purification, properties and activation of factor X. Thrombos. Diathes. haemorrh. (Stuttg.) (Suppl. 01) 07: 213 1962;
    PubMedSearch in Google Scholar
  • 5 Goldstein R, LeBolloc’h A, Alexander B, Zonderman E. Preparation and properties of prothrombin. J. biol. Chem. 234: 2857 1959;
    PubMedSearch in Google Scholar
  • 6 Harmison C. R, Seegers W. H. Some physicochemical properties of bovine autoprothrombin II. J. biol. Chem. 237: 3074 1962;
    PubMedSearch in Google Scholar
  • 7 Hewitt B. R. Spectrophotometric determination of protein in alkaline solution. Nature 182: 246 1958;
    PubMedSearch in Google Scholar
  • 8 Lamy F, Waugh D. F. Transformation of prothrombin and thrombin. Physiol. Rev. 34: 722 1954;
    CrossrefPubMedSearch in Google Scholar
  • 9 Lanchantin G. F, Friedman J. A, DeGroll J, Mehl J. W. Preparation of human plasma prothrombin and some of its sedimentation properties. J. biol. Chem. 238: 238 1963;
    PubMedSearch in Google Scholar
  • 10 Lorand L, Alkjaersig N, Seegers W. H. Carbohydrate and nitrogen distribution during the activation of purified prothrombin in sodium citrate solution. Arch. Biochem. 45: 312 1953;
    CrossrefPubMedSearch in Google Scholar
  • 11 Mammen E. F, Thomas W. R, Seegers W. H. Activation of purified prothrombin to autoprothrombin I or autoprothrombin II (platelet cofactor II or autoprothrombin II-A). Thrombos. Diathes. haemorrh. (Stuttg) 05: 218 1960;
    PubMedSearch in Google Scholar
  • 12 Moore S, Stein W. H. A modified ninhydrin reagent for the photometric determination of amino acids and related compounds. J. biol. Chem. 211: 907 1954;
    PubMedSearch in Google Scholar
  • 13 Rapaport S. I, Schiffman S, Patch M. J, Ames S. B. The importance of activation of antihemophilic globulin and proaccelerin by traces of thrombin in the generation of intrinsic prothrombinase activity. Blood 21: 221 1963;
    PubMedSearch in Google Scholar
  • 14 Ronwin E. Enzymatic properties of bovine plasmin preparations; evidence for similarity to but non-identity with trypsin. Canad. J. Biochem. 34: 1169 1956;
    PubMedSearch in Google Scholar
  • 15 Schwick G, Schultze H. E. Immunchemische Untersuchungen mit Prothrombin und Thrombin. Clin. chim. Acta. 04: 26 1959;
    CrossrefPubMedSearch in Google Scholar
  • 16 Seegers W. H. Prothrombin and fibrinolysin. Science 104: 461 1946;
    CrossrefPubMedSearch in Google Scholar
  • 17 Streuli F, Bang N. U, Luchini B. W. Prothrombin-thrombin conversion caused by proteolytic enzymes. Fed. Proc. 22: 560 1963;
    PubMedSearch in Google Scholar
  • 18 Svennerholm L. The determination of hexosamines with special reference to nervous tissue. Acta Soc. Med. upsalien. 61: 287 1957;
    PubMedSearch in Google Scholar
  • 19 Tishkoff G. H, Pechet L, Alexander B. Some biochemical and electrophoretic studies on purified prothrombin, factor VII (proconvertin) and factor X (Stuart). Blood 15: 778 1960;
    PubMedSearch in Google Scholar
  • 20 Ware A. G, Seegers W. H. Two-stage procedure for the quantitative determination of prothrombin concentration. Amer. J. clin. Path. 19: 471 1949;
    PubMedSearch in Google Scholar
  • 21 Warren L. The thiobarbituric acid assay of sialic acids. J. biol. Chem. 234: 1971 1959;
    PubMedSearch in Google Scholar
  • 22 Weimer H. E, Moshin J. R. Serum glycoprotein concentration in experimental tuberculosis of guinea pigs. Amer. Rev. Tuberc. 68: 594 1952;
    PubMedSearch in Google Scholar