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Thromb Haemost 1989; 61(02): 208-210
DOI: 10.1055/s-0038-1646560
Original Article
Schattauer GmbH Stuttgart

A Rapid and Simple Method for the Separation of Four Molecular Forms of Human Plasminogen

W Nieuwenhuizen
The Gaubius Institute TNO, Leiden, The Netherlands
,
D W Traas
The Gaubius Institute TNO, Leiden, The Netherlands
› Author Affiliations
Further Information

Publication History

Received 21 September 1988

Accepted after revision 01 December 1988

Publication Date:
30 June 2018 (online)

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Summary

At least four molecular forms of plasminogen are known. Two of those forms have glutamic acid at their amino-terminal end, and are designated as glu-plasminogen. The other two have lysine, methionine and/or valine as amino-terminal amino acid and are collectively designated as lys-plasminogen. Two subforms (I and II) each of glu- and lys-plasminogen exist. The I-forms are glycosylated at asn-288 and thr-345, whereas the II-forms are only glycosylated at thr-345. In a previous publication (Thromb Haemostas 1984; 52: 347-349) we have described the separation of the I- and II-forms of plasminogen in lysine-Sepharose in phosphate buffers. Now we have combined those findings with the differential affinity of glu- and lys-plasminogen for aminohexyl-Sepharose through their aminohexyl-sites, recently described by Christensen (Biochem J 1984; 223: 431-421). Acid/urea electrophoresis, end-group determination and carbohydrate analysis show that the combination of affinity chromatography on lysine-Sepharose in phosphate buffers, and on aminohexyl-Sepharose provides an efficient procedure to separate the four molecular forms of plasminogen.

Keywords

Plasminogen - Affinity chromatography - Lysine-Sepharose - Aminohexyl-Sepharose
 

  • References

  • 1 Castellino FJ. Biochemistry of human plasminogen. Semin Thromb Haemostas 1984; 10: 18-23
    Article in Thieme ConnectPubMedGoogle Scholar
  • 2 Sottrup-Jensen L, Claeys H, Zajdal M, Petersen TE, Magnusson S. The primary structure of human plasminogen: Isolation of two lysinebinding fragments and one “mini” plasminogen (MW. 38,000) by elastase-catalysed-specific limited proteolysis. Prog Chem Fibrinolysis Thrombolysis 1978; 3: 191-209
    PubMedGoogle Scholar
  • 3 Lerch PG, Rickli EE, Lergier W, Gillesen D. Localization of individual lysine-binding regions in human plasminogen and investigations on their complex-forming properties. Eur J Biochem 1980; 107: 7-13
    CrossrefPubMedGoogle Scholar
  • 4 Markus G, De Pasquale JL, Wissler FC. Quantitative determination of the binding of epsilon-aminocaproic acid to native plasminogen. J Biol Chem 1978; 253: 727-732
    PubMedGoogle Scholar
  • 5 Hochschwender SM, Laursen RA. The lysine binding sites of human plasminogen. Evidence for a critical tryptophan in the binding site of kringle 4. J Biol Chem 1981; 257: 11172-11176
    PubMedGoogle Scholar
  • 6 Trexler M, Váli Z, Patthy L. Structure of the ω-amino carboxylic acidbinding sites of human plasminogen, Arginine 70 and aspartic acid 56 are essential for binding of ligand by kringle 4. J Biol Chem 1982; 257: 7401-7406
    PubMedGoogle Scholar
  • 7 Christensen U. The AH-site of plasminogen and two C-terminal fragments. A weak lysine-binding site preferring ligands not carrying a free carboxylate function Biochem J 1984; 223: 413-421
    CrossrefPubMedGoogle Scholar
  • 8 Castellino FJ, Powel JR. Human plasminogen. In: Methods in Enzymology. Lorand L. ed Academic Press; New York: 1981. 80 365-378
    Google Scholar
  • 9 Wallén P, Wiman B. Characterization of human plasminogen. II. Separation and partial characterization of different molecular forms of human plasminogen. Biochem Biophys Acta 1972; 257: 122-134
    CrossrefPubMedGoogle Scholar
  • 10 Traas DW, Hoegee-de Nobel B, Nieuwenhuizen W. Factors influencing the separation of glu-plasminogen affinity forms I and II by affinity chromatography. Thromb Haemostas 1984; 52: 347-349
    PubMedGoogle Scholar
  • 11 Robbins KC, Summaria L. Plasminogen and plasmin. Methods Enzymol 1976; 45 B 257-273
    PubMedGoogle Scholar
  • 12 Deutsch DG, Mertz ET. Plasminogen: purification from human plasma by affinity chromatography. Science 1970; 170: 1095-1096
    CrossrefPubMedGoogle Scholar
  • 13 Lucas MA, Straight DL, Fretto LJ, McKees PA. The effects of fibrinogen and its cleavage products on the kinetics of plasminogen activation by urokinase and subsequent plasmin activity. J Biol Chem 1983; 258: 12171-12177
    PubMedGoogle Scholar
  • 14 Gray WR. End-group analysis using dansyl chloride. Methods Enzymol 1972; 25 B 121-138
    PubMedGoogle Scholar
  • 15 Kamerling JP, Vliegenthart JF G. Sialic acids. Chemistry metabolism and function Cell Biol Monogr 1982; 10: 95-125
    PubMedGoogle Scholar
  • 16 Kamerling JP, Gerwig GJ, Vliegenthart JF G, Clamp JR. Characterization by Gas-Liquid chromatography-mass spectrometry and proton-magnetic-resonance spectroscopy of pertrimethylsilylmethyl glycosides obtained in the methanolysis of glycoproteins and glycopeptides. Biochem J 1975; 151: 491-495
    CrossrefPubMedGoogle Scholar
  • 17 Hayes ML, Castellino FJ. Carbohydrate of the human plasminogen variants. I. Carbohydrate composition, glycopeptide isolation, and characterization. J Biol Chem 1979; 254: 8768-8771
    PubMedGoogle Scholar