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Specific binding of urinary-type plasminogen activator (u-PA) to vitronectin and its role in mediating u-PA-dependent adhesion of U937 cells.

Publication ,  Journal Article
Moser, TL; Enghild, JJ; Pizzo, SV; Stack, MS
Published in: Biochem J
May 1, 1995

The present paper described interactions of urinary-type plasminogen activator (u-PA) with isolated protein components of the extracellular matrix (ECM) using kinetic and ligand-blotting analyses, as well as adhesion studies with u-PA-saturated U937 monocytic cells. Kinetic analyses showed that fibronectin and laminin were moderately effective at decreasing activation of plasminogen by u-PA (3-4-fold decrease in kcat/Km), while activation was stimulated slightly by collagen types I and IV (2-4-fold increase in kcat/Km). Ligand-blotting experiments using intact immobilized ECM proteins demonstrated that u-PA binds predominantly to vitronectin. This was supported by ELISA studies, which showed concentration dependent, saturable, reversible binding of u-PA to vitronectin (Kd,app. of 97 nM). Limited proteolysis of vitronectin followed by ligand-blotting analysis demonstrated u-PA binding to a specific vitronectin fragment (M(r) 49,000), and binding was shown to occur through the N-terminal fragment of u-PA. N-terminal sequence analysis indicated that this binding fragment of vitronectin originates with Thr-122 and comprises the hemopexin domain, including the heparin-binding region of the vitronectin molecule. Plasminogen activator inhibitor type I did not compete with u-PA for binding to vitronectin, suggesting both molecules may co-localize on vitronectin. In contrast, binding of u-PA to vitronectin was significantly inhibited by plasminogen, suggesting these molecules share a common binding site on vitronectin. In addition to in vitro studies, experiments were performed to assess the contribution of direct binding of u-PA to vitronectin on the adhesive behaviour of U937 cells. Binding of u-PA-saturated U937 cells to vitronectin was inhibited 66% by excess vitronectin, suggesting that direct binding of u-PA to vitronectin is the mechanism by which u-PA-dependent adhesion of U937 cells to vitronectin is mediated.

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Published In

Biochem J

DOI

ISSN

0264-6021

Publication Date

May 1, 1995

Volume

307 ( Pt 3)

Issue

Pt 3

Start / End Page

867 / 873

Location

England

Related Subject Headings

  • Vitronectin
  • Urokinase-Type Plasminogen Activator
  • Tumor Cells, Cultured
  • Sensitivity and Specificity
  • Receptors, Urokinase Plasminogen Activator
  • Receptors, Cell Surface
  • Protein Binding
  • Plasminogen Activator Inhibitor 1
  • Peptide Fragments
  • Molecular Sequence Data
 

Citation

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Moser, T. L., Enghild, J. J., Pizzo, S. V., & Stack, M. S. (1995). Specific binding of urinary-type plasminogen activator (u-PA) to vitronectin and its role in mediating u-PA-dependent adhesion of U937 cells. Biochem J, 307 ( Pt 3)(Pt 3), 867–873. https://doi.org/10.1042/bj3070867
Moser, T. L., J. J. Enghild, S. V. Pizzo, and M. S. Stack. “Specific binding of urinary-type plasminogen activator (u-PA) to vitronectin and its role in mediating u-PA-dependent adhesion of U937 cells.Biochem J 307 ( Pt 3), no. Pt 3 (May 1, 1995): 867–73. https://doi.org/10.1042/bj3070867.
Moser, T. L., et al. “Specific binding of urinary-type plasminogen activator (u-PA) to vitronectin and its role in mediating u-PA-dependent adhesion of U937 cells.Biochem J, vol. 307 ( Pt 3), no. Pt 3, May 1995, pp. 867–73. Pubmed, doi:10.1042/bj3070867.
Journal cover image

Published In

Biochem J

DOI

ISSN

0264-6021

Publication Date

May 1, 1995

Volume

307 ( Pt 3)

Issue

Pt 3

Start / End Page

867 / 873

Location

England

Related Subject Headings

  • Vitronectin
  • Urokinase-Type Plasminogen Activator
  • Tumor Cells, Cultured
  • Sensitivity and Specificity
  • Receptors, Urokinase Plasminogen Activator
  • Receptors, Cell Surface
  • Protein Binding
  • Plasminogen Activator Inhibitor 1
  • Peptide Fragments
  • Molecular Sequence Data