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Thromb Haemost 1977; 38(02): 0475-0485
DOI: 10.1055/s-0038-1651853
Original Article
Schattauer GmbH

Isolation of Antithrombin III from Normal and α1-Antitrypsin-Deficient Human Plasma

Anna D. Borsodi   *
1   Department of Biological Chemistry, Washington University School of Medicine, St. Louis, MO 63110, U. S. A.
,
Ralph A. Bradshaw
1   Department of Biological Chemistry, Washington University School of Medicine, St. Louis, MO 63110, U. S. A.
› Author Affiliations
Further Information

Publication History

Received 16 December 1976

Accepted 10 April 1977

Publication Date:
04 July 2018 (online)

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Summary

The plasma of individuals, hetero- or homozygous for α1-antitrypsin deficiency, contains greatly decreased amounts of antithrombin activity as assayed against factor Xa. However, heparin stimulation of the residual antithrombin activity is observed, which is comparable to that of normal plasma. Antithrombins isolated from both normal and α1-antitrypsin deficient plasma by a simplified procedure are indistinguishable in both properties and yields. The microheterogeneity observed on isoelectric focusing of both preparations can be eliminated by treatment with neuraminidase. Neither purified human antithrombin nor α1-antitrypsin, when assayed against bovine trypsin, is stimulated by heparin. These results clearly establish the unique natures of antithrombin and α1-antitrypsin and show that about 75% of the antithrombin activity measured in normal plasma is due to α1-antitrypsin. Estimates of anti thrombin III activity in normal plasma by assays dependent on enzymatic activity can probably be obtained only in the presence of heparin.

* Present address: Institute of Biochemistry, Biological Research Center, Hungarian Academy of Sciences, H-6071 Szeged, Hungary.


 

  • References

  • 1 Abildgaard U. 1967; a Purification of Two Progressive Antithrombins of Human Plasma. Scandinavian Journal of Clinical and Laboratory Investigations 19: 190.
    CrossrefPubMedGoogle Scholar
  • 2 Abildgaard U. 1967; b Inhibition of the Thrombin-fibrinogen Reaction by Antithrombin III, Studied by N-terminal Analysis. Scandinavian Journal of Clinical and Laboratory Investigations 20: 207.
    CrossrefPubMedGoogle Scholar
  • 3 Abildgaard U. 1968; Highly Purified Antithrombin III with Heparin Cofactor Activity Prepared by Disc Electrophoresis. Scandinavian Journal of Clinical and Laboratory Investigation 21: 89.
    CrossrefPubMedGoogle Scholar
  • 4 Abildgaard U, Egeberg O. 1968; Thrombin Inhibitory Activity of Fractions Obtained by Gel Filtration of Antithrombin III Deficient Plasma. Scandinavian Journal of Haematology 5: 155.
    PubMedGoogle Scholar
  • 5 Biggs R, Denson K. W. E, Akman N, Borrett R, Hadden M. 1970; Antithrombin III, Antifactor Xa and Heparin. British Journal of Haematology 19: 283.
    CrossrefPubMedGoogle Scholar
  • 6 Borsodi A. D, Bradshaw R. A, Rughani I. K, Bruce R. M. 1975. Structural and Functional Relationships of Human Antithrombin III and α1-Antitrypsin. In: Bradshaw R. A, Wessler S. (eds.) Heparin: Structure, Function and Clinical Implications. Plenum Publishing Co.; N.Y.: 249-253.
    Google Scholar
  • 7 Borsodi A, Narasimhan T. R. 1977 Microheterogeneity of Human Antithrombin. Biochimica et Biophysica Acta. in press.
    PubMedGoogle Scholar
  • 8 Crawford I. P. 1973; Purification and Properties of Normal Human α1-Antitrypsin. Archives of Biochemistry and Biophysics 156: 215.
    CrossrefPubMedGoogle Scholar
  • 9 Danishefsky F, Tzeng F, Ahrens M, Klein S. 1976; Synthesis of Heparin-Sepharoses and Their Binding with Thrombin and Antithrombin-Heparin Cofactor. Thrombosis Research 8: 131.
    CrossrefPubMedGoogle Scholar
  • 10 Davis B. J. 1964; Disc Electrophoresis – II. Method and Application to Human Serum Proteins. Annals of New York Academy of Sciences 121: 404.
    PubMedGoogle Scholar
  • 11 Desnuelle P, Neurath H, Ottensen M. (eds.) 1969. Structure-Function Relationships of Proteolytic Enzymes. Proceedings of the International Symposium Copenhagen. Academic Press; N. Y.:
    Google Scholar
  • 12 Duda E. 1973; A New, Sensitive Method for the Determination of Streptomycin. Analytical Biochemistry 51: 651.
    CrossrefPubMedGoogle Scholar
  • 13 Egeberg O. 1965; a Inherited Antithrombin Deficiency Causing Thrombophilia. Thrombosis et Diathesis Haemorrhagica 13: 516.
    PubMedGoogle Scholar
  • 14 Egeberg O. 1965; b On the Natural Blood Coagulation Inhibitor System. Investigations of Inhibitor Factors Based on Antithrombin Deficient Blood. Thrombosis et Diathesis Haermorrhagica 14: 473.
    PubMedGoogle Scholar
  • 15 Eriksson S. 1965; Studies on α1-Antitrypsin Deficiency. Acta Medica Scandinavica 177 Suppl. 432: 5.
    PubMedGoogle Scholar
  • 16 Frénoy J.-P, Bourrillon R. 1974; Studies on the Structure of Human α1-Macroglobulin. IV. Analysis of the Microheterogeneity by Isoelectric Focussing. Biochimica et Biophysica Acta 371: 168.
    CrossrefPubMedGoogle Scholar
  • 17 Gans H, Tan B. H. 1967; α1-Anti trypsin, An Inhibitor for Thrombin and Plasmin. Clinica Chimica Acta 17: 111.
    CrossrefPubMedGoogle Scholar
  • 18 Kueppers F, Bearn A. G. 1966; A Possible Experimental Approach to the Association of Hereditary α1-Antitrypsin Deficiency and Pulmonary Emphysema. Proceedings of the Society for Experimental Biology and Medicine 121: 1207.
    PubMedGoogle Scholar
  • 19 Kurachi K, Schmer G, HermOdson M. A, Teller D, Davie E. W. 1976; Characterization of Human, Bovine, and Horse Antithrombin III. Biochemistry 15: 368.
    CrossrefPubMedGoogle Scholar
  • 20 Laemmli U. K. 1970; Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4. Nature 227: 680.
    CrossrefPubMedGoogle Scholar
  • 21 Lanchantin G. F, Plesset M. L, Friedmann J. A, Hart D. W. 1966; Dissociation of Esterolytic and Clotting Activities of Thrombin by Trypsin-binding Macroglobulin. Proceedings of the Society for Experimental Biology and Medicine 121: 444.
    PubMedGoogle Scholar
  • 22 Laurell C.-B, Eriksson S. 1963; The Electrophoretic α1-Globulin Pattern of Serum in α1-Antitrypsin Deficiency. Scandinavian Journal of Clinical and Laboratory Investigation 15: 132.
    CrossrefPubMedGoogle Scholar
  • 23 Laurell C.-B, Eriksson S. 1965; The Serum α1-Antitrypsin in Families with Hypo-α1-Antitrypsinemia. Clínica Chimica Acta 11: 395.
    PubMedGoogle Scholar
  • 24 Monkhouse F. C. 1970; Preparation and Assay of Plasma Antithrombin. Methods in Enzymology XIX: 915.
    PubMedGoogle Scholar
  • 25 Porter P, Porter M. C, Shanberge J. N. 1967; Heparin Cofactor and Plasma Antithrombin in Relation to the Mechanism of Inactivation of Thrombin by Heparin. Clinica Chimica Acta 77: 189.
    PubMedGoogle Scholar
  • 26 Rimon A, Shamash Y, Shapiro B. 1966; The Plasmin Inhibitor of Human Plasma. IV. Its Action on Plasmin, Trypsin, Chymotrypsin and Thrombin. Journal of Biological Chemistry 241: 5102.
    PubMedGoogle Scholar
  • 27 Rosenberg R. D, Damus P. S. 1973; The Purification and Mechanism of Action of Human Antithrombin-Heparin Cofactor. Journal of Biological Chemistry 248: 6490.
    PubMedGoogle Scholar
  • 28 Schultze H. E, Heide K, Haupt H. 1962; Alpha1-antitrypsin and Human Serum. Klinische Wochenschrift 40: 427.
    CrossrefPubMedGoogle Scholar
  • 29 Seegers W. H, Marciniak E. 1962; Inhibition of Autoprothrombin C Activity with Plasma. Nature 193: 1188.
    PubMedGoogle Scholar
  • 30 Szezeklik A, Teresiak T, KoDaczkowska B. 1971; Niedobor Alfa1-antytrypsyny I Antytrombiny III W Przypadku Rozedmy Pluc. Polskie Archiwum Medycyny Wewnetrznejwi 47: 407.
    PubMedGoogle Scholar
  • 31 Tan B. H, Gans H. 1969; Alpha1-Antitrypsin. III. Inhibition of Trypsin, Thrombin, Plasmin and Leucoproteinases. Pharmaceutisch Weekblad 104: 717.
    PubMedGoogle Scholar
  • 32 Wessler S. 1975. Clinical Implications of Heparin. In: Bradshaw R. A, Wessler S. (Eds.) Heparin: Structure, Function, and Clinical Implications. Plenum Publishing Co.; N. Y.: 309-322.
    Google Scholar
  • 33 Yin E. T, Wessler S, Stoll P. J. 1971; a Rabbit Plasma Inhibitor of The Activated Species of Blood Coagulation Factor X. Purification and Some Properties. Journal of Biological Chemistry 246: 3694.
    PubMedGoogle Scholar
  • 34 Yin E. T, Wessler S, Stoll P. J. 1971; b Identity of Plasma-activated Factor X Inhibitor with Antithrombin III and Heparin Cofactor. Journal of Biological Chemistry 246: 3712.
    PubMedGoogle Scholar
  • 35 Yin E. T, Wessler S, Butler J. V. 1973; Plasma Heparin: A Unique, Practical, Submicrogram-sensitive Assay. Journal of Clinical and Laboratory Investigation 81: 298.
    PubMedGoogle Scholar
  • 36 Zacharius R. M, Zell T. E, Morrison J. H, Woodlock J. J. 1969; Glycoprotein Staining Following Electrophoresis on Acrylamide Gels. Analytical Biochemistry 30: 148.
    CrossrefPubMedGoogle Scholar