Mapping the heparin-binding site of the BMP antagonist gremlin by site-directed mutagenesis based on predictive modelling

2015 ◽  
Vol 470 (1) ◽  
pp. 53-64 ◽  
Author(s):  
Arnold Junior Tatsinkam ◽  
Barbara Mulloy ◽  
Christopher C. Rider
Keyword(s):  
Binding Site ◽  
Kidney Development ◽  
Predictive Modelling ◽  
Targeted Mutagenesis ◽  
Site Directed Mutagenesis ◽  
Sulfated Polysaccharides ◽  
Heparin Binding ◽  
Heparin Binding Site ◽  
Foetal Kidney ◽  
Bmp Antagonist

Gremlin, a secreted protein critical in foetal kidney development, is known to bind to the sulfated polysaccharides, heparin and heparan sulfate (HS). In the present study, we employed targeted mutagenesis to map its heparin-binding site. We also demonstrate that gremlin forms non-covalent dimers.

Localization of the Heparin-Binding Site of Glia-Derived Nexin/Protease Nexin-1 by Site-Directed Mutagenesis

Biochemistry ◽  
1994 ◽  
Vol 33 (24) ◽  
pp. 7731-7735 ◽  
Author(s):  
Stuart R. Stone ◽  
Marianne L. Brown-Luedi ◽  
Giorgio Rovelli ◽  
Angelo Guidolin ◽  
Elaine McGlynn ◽  
...  
Keyword(s):  
Binding Site ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Heparin Binding ◽  
Heparin Binding Site ◽  
Protease Nexin

scholarly journals Mapping the heparin-binding site of the osteoinductive protein NELL1 by site-directed mutagenesis

FEBS Letters ◽  
2015 ◽  
Vol 589 (24PartB) ◽  
pp. 4026-4032 ◽  
Author(s):  
Kaneyoshi Takahashi ◽  
Arisa Imai ◽  
Masumi Iijima ◽  
Nobuo Yoshimoto ◽  
Andrés D. Maturana ◽  
...  
Keyword(s):  
Binding Site ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Heparin Binding ◽  
Heparin Binding Site

Antithrombin-mediated Inhibition of Factor Vlla-Tissue Factor Complex by the Synthetic Pentasaccharide Representing the Heparin Binding Site to Antithrombin

1996 ◽  
Vol 76 (01) ◽  
pp. 005-008 ◽  
Author(s):  
Jean Claude Lormeau ◽  
Jean Pascal Herault ◽  
Jean Marc Herbert
Keyword(s):  
Binding Site ◽  
Tissue Factor ◽  
Residual Activity ◽  
Coagulation Factors ◽  
Heparin Binding ◽  
Experimental Conditions ◽  
First Order ◽  
Heparin Binding Site ◽  
Coagulant Activity

SummaryWe examined the effect of the synthetic pentasaccharide representing the minimal binding site of heparin to antithrombin on the antithrombin-mediated inactivation of factor Vila bound to tissue factor. This effect was compared to the effect of unfractionated heparin. Using purified recombinant human coagulation factors and either a clotting or an amidolytic assay for the determination of the residual activity of factor Vila, we showed that the pentasaccharide was an efficient antithrombin-dependent inhibitor of the coagulant activity of tissue factor-factor Vila complex. In our experimental conditions, assuming a mean MW of 14,000 for heparin, the molar pseudo-first order rate constants for ATIII-mediated FVIIa inhibition by ATIII-binding heparin and by the synthetic pentasaccharide were found to be similar with respective values of 104,000 ± 10,500 min-1 and 112,000 ± 12,000 min-1 (mean ± s.e.m., n = 3)

scholarly journals Localization of the major heparin-binding site in fibronectin.

1991 ◽  
Vol 266 (12) ◽  
pp. 7812-7818 ◽  
Author(s):  
F J Barkalow ◽  
J E Schwarzbauer
Keyword(s):  
Binding Site ◽  
Heparin Binding ◽  
Heparin Binding Site

Overlap of IGF- and heparin-binding sites in rat IGF-binding protein-5

2000 ◽  
Vol 24 (1) ◽  
pp. 43-51 ◽  
Author(s):  
H Song ◽  
J Beattie ◽  
IW Campbell ◽  
GJ Allan
Keyword(s):  
Binding Site ◽  
Conformational Changes ◽  
Binding Protein ◽  
Binding Activity ◽  
Binding Domain ◽  
Site Directed Mutagenesis ◽  
Heparin Binding ◽  
Igf I ◽  
Heparin Binding Domain ◽  
Igf Binding

Using site-directed mutagenesis, we have undertaken a study of a potential IGF-binding site in the C-terminal domain of rat IGFBP-5, lying close to or within a previously described heparin-binding domain (residues 201-218) in this protein. After analysis of binding activity using three different methods - ligand blotting, solution phase equilibrium binding and biosensor measurement of real-time on- and off-rates - we report that the mutation of two highly conserved residues within this region (glycine 203 and glutamine 209) reduces the affinity of the binding protein for both IGF-I and IGF-II, while having no effect on heparin binding. In addition, we confirm that mutation of basic residues within the heparin-binding domain (R201L, K202E, K206Q and R214A) results in a protein that has attenuated heparin binding but shows only a small reduction in affinity for IGF-I and -II. Previous findings have described the reduction in affinity of IGFBP-5 for IGFs that occurs after complexation of the binding protein with heparin or other components of the extracellular matrix (ECM) and have postulated that such an interaction may result in conformational changes in protein structure, affecting subsequent IGF interaction. Our data suggesting potential overlap of heparin- and IGF-binding domains argue for a more direct effect of ECM modulation of the affinity of IGFBP-5 for ligand by partial occlusion of the IGF-binding site after interaction with ECM.

Structural Analysis of the Heparin-Binding Site of the NC1 Domain of Collagen XIV by CD and NMR†,‡

Biochemistry ◽  
1999 ◽  
Vol 38 (20) ◽  
pp. 6479-6488 ◽  
Author(s):  
Roland Montserret ◽  
Elisabeth Aubert-Foucher ◽  
Michael J. McLeish ◽  
Joanna M. Hill ◽  
Damien Ficheux ◽  
...  
Keyword(s):  
Structural Analysis ◽  
Binding Site ◽  
Heparin Binding ◽  
Heparin Binding Site ◽  
Nc1 Domain

Pulmonary arterial thrombosis in a neonate with homozygous deficiency of antithrombin III: successful outcome following pulmonary thrombectomy and infusions of antithrombin III concentrate

2000 ◽  
Vol 10 (3) ◽  
pp. 275-278 ◽  
Author(s):  
K. Roman ◽  
E. Rosenthal ◽  
R. Razavi
Keyword(s):  
Binding Site ◽  
Arterial Thrombosis ◽  
Antithrombin Iii ◽  
Pulmonary Trunk ◽  
Successful Outcome ◽  
Heparin Binding ◽  
The Third ◽  
Heparin Binding Site ◽  
Surgical Thrombectomy ◽  
Pulmonary Arterial

AbstractWe report a newborn male who presented with severe central cyanosis on the third day of life. Partial thrombotic obstruction of the pulmonary trunk secondary to Antithrombin III (homozygous defect of heparin binding site) deficiency was subsequently diagnosed. Surgical thrombectomy, and infusions of Antithrombin III concentrate, led to a successful outcome. We postulate that intrauterine thrombosis occurred to give this unusual presentation.

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