Biopolym. Cell. 2015; 31(5):345-350.
http://dx.doi.org/10.7124/bc.0008F6
Structure and Function of Biopolymers
Plasmin enzymatic activity in the presence of actin
1Yusova E. I.
  1. Palladin Institute of Biochemistry, NAS of Ukraine
    9, Leontovycha Str., Kyiv, Ukraine, 01601

Abstract

Aim. To study the changes in the plasmin activity towards substrates with high and low molecular mass in the presence of actin. Methods. The proteins used for this investigation were obtained by affinity chromatography and gel-filtration. The plasmin enzymatic activity was determined by a turbidimetric assay and a chromogenic substrate-based assay. The enzyme linked immunosorbent assay and biotin-avidin-phosphatase system were used to study the interaction of plasminogen and its fragments with actin. Results. It was shown that G-actin causes 1.5-fold decrease in the rate of polymeric fibrin hydrolysis by plasmin and Glu-plasminogen activated by the tissue plasminogen activator. However, actin did not impede plasmin autolysis and had no influence on its amidase activity. We have studied an interaction of biotinylated Glu-plasminogen and its fragments (kringle 1-3, kringle 4 and mini-plasminogen) with immobilized G-actin. Glu-plasminogen and kringle 4 had a high affinity towards actin (C50 is 113 and 117 nM correspondingly). Mini-plasminogen and kringe 4 did not bind to actin. A similar affinity of Glu-plasminogen and kringle 1-3 towards actin proves the involvement of the kringle 1-3 lysine-binding sites of the native plasminogen form in the actin interaction. Conclusions. Actin can modulate plasmin specificity towards high molecular mass substrates through its interaction with lysine-binding sites of the enzyme kringle domains. Actin inhibition of the fibrinolytic activity of plasmin is due to its competition with fibrin for thelysine binding sites of plasminogen/plasmin.
Keywords: Plasmin, actin, plasminogen fragments, enzymatic activity of plasmin.
Full text: (PDF, in English)

References

[1] Syrovets T, Lunov O, Simmet T. Plasmin as a proinflammatory cell activator. J Leukoc Biol. 2012;92(3):509-19.
[2] Marcum JA, Kline DL. Species specificity of streptokinase. Comp Biochem Physiol B. 1983;75(3):389-94.
[3] Wohl RC, Sinio L, Summaria L, Robbins KC. Comparative activation kinetics of mammalian plasminogens. Biochim Biophys Acta. 1983;745(1):20-31.
[4] Castellino FJ, Sodetz JM, Brockway WJ, Siefring GE Jr. Streptokinase. Methods Enzymol. 1976;45:244-57.
[5] Cederholm-Williams SA, De Cock F, Lijnen HR, Collen D. Kinetics of the reactions between streptokinase, plasmin and alpha 2-antiplasmin. Eur J Biochem. 1979;100(1):125-32.
[6] Wang H, Schultz R, Hong J, Cundiff DL, Jiang K, Soff GA. Cell surface-dependent generation of angiostatin4.5. Cancer Res. 2004;64(1):162-8.
[7] Mejean C, Roustan C, Benyamin Y. Anti-actin antibodies. Detection and quantitation of total and skeletal muscle actin in human plasma using a competitive ELISA. J Immunol Methods. 1987;99(1):129-35.
[8] Deutsch DG, Mertz ET. Plasminogen: purification from human plasma by affinity chromatography. Science. 1970;170(3962):1095-6.
[9] Sottrup-Jensen L, Claeys H, Zajdel M, Petersen TE, Magnusson S. The primary structure of human plasminogen: isolation of two lysine-binding fragment and one “mini-plasminogen” (M.W. 38000) by elastase catalyzed specific limited proteolysis In: Progress in chemical fibrinolysis and thrombolysis. Eds VF Davidson, Rowan RH, Samama MM, Desnoyers DC. Raven Press, New-York. 1977;3:191–9,
[10] Pozdnjakova TM, Musjalkovskaja AA, Ugarova TP, Protvin DD, Kotsjuruba VN. On the properties of fibrin monomer prepared from fibrin clot with acetic acid. Thromb Res. 1979;16(1-2):283-8.
[11] Bouvier SA, Beretta-Piccoli M, Giacometti N. Light scattering measurements of polymerization and depolimerization of fibrin: a tool for studing coagulation and fibrinolysis. Progress in chemical fibrinolysis and thrombolysis. Eds. Davidson JF et al. N.Y. Raven Press 1975;1:281–88.
[12] Gitlin G, Bayer EA, Wilchek M. Studies on the biotin-binding site of avidin. Lysine residues involved in the active site. Biochem J. 1987;242(3):923-6.
[13] Violand BN, Castellino FJ. Mechanism of the urokinase-catalyzed activation of human plasminogen. J Biol Chem. 1976;251(13):3906-12.
[14] Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970;227(5259):680-5.
[15] Robbins KC, Summaria L, Hsieh B, Shah RJ. The peptide chains of human plasmin. Mechanism of activation of human plasminogen to plasmin. J Biol Chem. 1967;242(10):2333-42.
[16] Hoylaerts M, Rijken DC, Lijnen HR, Collen D. Kinetics of the activation of plasminogen by human tissue plasminogen activator. Role of fibrin. J Biol Chem. 1982;257(6):2912-9.
[17] Stack S, Gonzalez-Gronow M, Pizzo SV. Regulation of plasminogen activation by components of the extracellular matrix. Biochemistry. 1990;29(20):4966-70.
[18] Zhernosekov DD, Iusova EI, Grinenko TV. [Role of plasminogen/plasmin in functional activity of blood cells]. Ukr Biokhim Zh (1999). 2012;84(4):5-19.
[19] Tykhomyrov AA. Interaction of actin with plasminogen. plasmin system: mechanisms and physiological role. Biopolym Cell. 2012; 28(6):413–23.
[20] Lind SE, Smith CJ. Actin accelerates plasmin generation by tissue plasminogen activator. J Biol Chem. 1991;266(26):17673-8.