Kinetic studies on the effect of heparin and fibrin on plasminogen activators

Fears, Robin

doi:10.1042/bj2490077

Cited by 51 publications

(35 citation statements)

References 33 publications

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“…Heparin and heparin-like glycosaminoglycans are reported to enhance the activation of Glu-Plg by t-PA in vitro [4][5][6][7][8][9]. Activation of Glu-Plg by t-PA was reported [32] to be enhanced in a synergistic manner when fucoidan was added in conjunction with 6-AH.…”

Section: Discussionmentioning

confidence: 98%

“…Heparin and heparin-like compounds are also reported to enhance the in-vitro activation of plasminogen by tissue plasminogen activator (t-PA) and by urokinase [4][5][6][7][8][9]. Fibrin monomer, fibrin polymer, and its mimic, CNBr-digested fibrinogen are also reported to enhance the activation of glutamic type plasminogen (Glu-Plg) by t-PA [9][10][11][12][13][14][15]. Fucoidan, a sulfated poly(L-fucopyranose) from brown marine algae Fucus vesiculosus was reported to show anticoagulant activity in vitro and in vivo [16,17].…”

Section: Introductionmentioning

confidence: 99%

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The effect of fucoidan, heparin and cyanogen bromide-fibrinogen on the activation of human glutamic-plasminogen by tissue plasminogen activator

Bell

Duhon²,

Doctor³

2003

Blood Coagulation & Fibrinolysis

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Earlier studies on the stimulatory effect of fucoidan, heparin, and cyanogen bromide (CNBr)-fibrinogen digest on the in-vitro activation of glutamic type plasminogen by tissue plasminogen activator, which were performed using subphysiologic ionic strengths of buffers, gave inconsistent results because of the variation in the ionic strengths of the buffers used. Studies were therefore conducted on the effect of these cofactors using 0.05 mol/l Tris buffer containing a physiologic concentration of sodium chloride. The double reciprocal plots of the activation of glutamic type plasminogen by tissue plasminogen activator in the presence of fucoidan and 6-aminohexanoic acid (6-AH) or heparin and 6-AH showed a four- to six-fold increase in K(cat), while the K(m) remained unchanged. On the other hand, there was greater than six-fold lowering of K(m) from 0.213 to 0.035 micromol/l in the presence of CNBr-fibrinogen, while K(cat) was only slightly increased. The ratios of the initial rate of plasmin generation in the presence or absence of the cofactors were plotted against the inverse of the volume fraction of glutamic type plasminogen or of tissue plasminogen activator after serial dilution. The results suggested that the enhancements by fucoidan and 6-AH or CNBr-fibrinogen were due to their interactions directed towards glutamic type plasminogen, while for heparin and 6-AH, the interaction was directed towards tissue plasminogen activator. Circular dichroism studies in the near ultraviolet range (250-308 nm) showed that 6-AH enhanced the circular dichroism spectra of glutamic type plasminogen around certain chromophores, while fucoidan and heparin had no effect, suggesting that the enhancement by the cofactors may be related to the favorable conformational changes of glutamic type plasminogen by 6-AH.

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Section: Discussionmentioning

confidence: 98%

Section: Introductionmentioning

confidence: 99%

The effect of fucoidan, heparin and cyanogen bromide-fibrinogen on the activation of human glutamic-plasminogen by tissue plasminogen activator

Bell

Duhon²,

Doctor³

2003

Blood Coagulation & Fibrinolysis

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“…The resulting organic extracts were subjected to successive chromatographic fractionation using silica gel, Sephadex LH‐20, and silica C 18 columns, giving 24 mg of purified staplabin. The CNBr fragment of fibrinogen (CNBr‐Fbg) was prepared as described [24, 25]. Radioiodination of Glu‐Plg, Lys‐Plg and ovalbumin was performed using the chloramine‐T method [26].…”

Section: Methodsmentioning

confidence: 99%

Enhancement of fibrin binding and activation of plasminogen by staplabin through induction of a conformational change in plasminogen

Takayasu¹,

Hasumi²,

Shinohara³

et al. 1997

FEBS Letters

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Staplabin (0.3-0.6 mM), a fungal triprenyl phenol, enhanced 3-6-fold the plasminogen activator-catalyzed activation of Glu-plasminogen and Lys-plasminogen as well as their binding to fibrin. Staplabin was not stimulatory to the amidolytic activity of plasmin and plasminogen activators. Even in the presence of e-aminocaproic acid (EACA) and fibrinogen fragments, allosteric effectors for Glu-plasminogen, staplabin increased the activation of both forms of plasminogen. In sizeexclusion chromatography of Glu-plasminogen and Lys-plasminogen, the molecular elution time, which varies as the conformation of a protein changes, was shortened by staplabin. These results suggest that staplabin causes plasminogens to be more susceptible to activation and fibrin binding by inducing a conformational change that is, at least in part, different from that induced by EACA and fibrinogen fragments.© 1997 Federation of European Biochemical Societies.

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“…By using purified components of the fibrinolytic system, heparin has been shown to enhance the activity of tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (6,7,24). Whereas commercial heparin binds tightly to both t-PA and Lysplasminogen, low-molecular-weight heparin does not bind t-PA and binds Lys-plasminogen only weakly (25).…”

mentioning

confidence: 99%

Fractionation of heparin by chromatography on a tissue plasminogen activator-Sepharose column.

Andrade‐Gordon

Strickland

1990

Proc. Natl. Acad. Sci. U.S.A.

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Heparin stimulates the activity of tissue plasminogen activator (t-PA) and binds to t-PA. To study this interaction, a complex between t-PA and N-acetylated heparin was formed and then linked to Sepharose. This procedure selectively links the t-PA to the column because the acetylated heparin has no free amino groups. The procedure also protects the heparin-binding site(s) on the enzyme during coupling to the matrix. The t-PA column separates heparin into two fractions, one with low affinity for t-PA and one with high affinity. Both fractions of heparin effectively accelerate inactivation of thrombin by antithrombin m. However, the fractions differ in their ability to stimulate t-PA: the low-affinity heparin has no effect on the activity of t-PA, whereas the high-affinity heparin enhances this activity. These heparin fractions will be useful in characterizing the biochemical basis and physiological consequences of the heparin-t-PA interaction.Heparin can bind many proteins of biological interest, including antithrombin III (AT-IIl), clotting factors (1), growth factors (2, 3), apolipoproteins (4), fibronectin (5), and plasminogen activators (6)(7)(8). The interaction between heparin and AT-III has been extensively studied. The anticoagulant potency of heparin, due primarily to its binding to AT-III (9, 10), depends on the molecular size of the preparation (11)(12)(13)(14) and a particular polysaccharide sequence (15)(16)(17). This work was aided by fractionation of heparin on columns of immobilized AT-Ill and chemically analyzing the fractions that bound most tightly (15,16,(18)(19)(20)(21)(22).Heparin has been reported to enhance fibrinolysis in vitro and in vivo (23). By using purified components of the fibrinolytic system, heparin has been shown to enhance the activity of tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (6,7,24). Whereas commercial heparin binds tightly to both t-PA and Lysplasminogen, low-molecular-weight heparin does not bind t-PA and binds Lys-plasminogen only weakly (25). These low-molecular-weight heparins do not stimulate plasmin formation by t-PA but still retain anticoagulant properties, such as binding to AT-III and acceleration of the inactivation of thrombin. Thus, size of the mucopolysaccharide appears important for its interaction with the fibrinolytic system.To study heparin-t-PA interaction, we sought a way to fractionate heparin on columns of immobilized t-PA. This approach was successful only when t-PA was protected by N-acetylated heparin during the coupling of t-PA to the matrix, similar to the protocol used for purification of AT-III-binding heparins (18). The chromatography has separated heparin into two fractions, one with high affinity for t-PA and the other with low affinity for t-PA. The anticoagulant activities of both fractions are similar. These two fractions should be useful in exploring the biochemical basis and clinical consequences of the heparin-t-PA interaction. EXPERIMENTAL PROCEDURES MaterialsHeparin (calcium salt) wa...

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Kinetic studies on the effect of heparin and fibrin on plasminogen activators

Cited by 51 publications

References 33 publications

The effect of fucoidan, heparin and cyanogen bromide-fibrinogen on the activation of human glutamic-plasminogen by tissue plasminogen activator

The effect of fucoidan, heparin and cyanogen bromide-fibrinogen on the activation of human glutamic-plasminogen by tissue plasminogen activator

Enhancement of fibrin binding and activation of plasminogen by staplabin through induction of a conformational change in plasminogen

Fractionation of heparin by chromatography on a tissue plasminogen activator-Sepharose column.

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