Thrombospondin-1 binds to polyhistidine with high affinity and specificity

2000 ◽  
Vol 347 (2) ◽  
pp. 469-473 ◽  
Author(s):  
Vijay K. VANGURI ◽  
Shuxia WANG ◽  
Svetlana GODYNA ◽  
Sripriya RANGANATHAN ◽  
Gene LIAU
Keyword(s):  
Solid Phase ◽  
Imidazole Ring ◽  
Binding Domain ◽  
Heparin Binding ◽  
Surface Receptors ◽  
High Affinity ◽  
Thrombospondin 1 ◽  
Heparin Binding Domain ◽  
Solid Phase Binding ◽  
And Migration

Thrombospondin-1 (TSP1) is a secreted trimeric glycoprotein of 450 kDa with demonstrated effects on cell growth, adhesion and migration. Its complex biological activity is attributed to its ability to bind to cell-surface receptors, growth factors and extracellular-matrix proteins. In this study, we used a 125I solid-phase binding assay to demonstrate that TSP1 binds specifically to proteins containing polyhistidine stretches. Based on studies with three different six-histidine-containing recombinant proteins, we derived an average dissociation constant of 5 nM. The binding of 125I-labelled TSP1 to these proteins was inhibited by peptides containing histidine residues, with the degree of competition being a function of the number of histidines within the peptide. Binding was not inhibited by excess histidine or imidazole, indicating that the imidazole ring is not sufficient for recognition by TSP1. Heparin was a potent inhibitor of binding with a Ki of 50 nM, suggesting that the heparin-binding domain of TSP1 may be involved in this interaction. This was confirmed by the ability of a recombinant heparin-binding domain of TSP1 to directly compete for TSP1 binding to polyhistidine-containing proteins. Affinity chromatography with a polyhistidine-containing peptide immobilized on agarose revealed that TSP1 in platelet releasates is the major polypeptide retained on the six-histidine-peptide column. We conclude that TSP1 contains a high-affinity binding site for polyhistidine and this is likely to be the molecular basis for the observed binding of TSP1 to histidine-rich glycoprotein. The possibility that other polyhistidine-containing proteins also interact with TSP1 warrants further study.

scholarly journals Cellular Internalization and Degradation of Thrombospondin-1 Is Mediated by the Amino-terminal Heparin Binding Domain (HBD)

1997 ◽  
Vol 272 (10) ◽  
pp. 6784-6791 ◽  
Author(s):  
Irina Mikhailenko ◽  
Dmitry Krylov ◽  
Kelley McTigue Argraves ◽  
David D. Roberts ◽  
Gene Liau ◽  
...  
Keyword(s):  
Binding Domain ◽  
Cellular Internalization ◽  
Heparin Binding ◽  
Amino Terminal ◽  
Thrombospondin 1 ◽  
Heparin Binding Domain

Apoptotic cell thrombospondin-1 and heparin-binding domain lead to dendritic-cell phagocytic and tolerizing states

Blood ◽  
2006 ◽  
Vol 108 (10) ◽  
pp. 3580-3589 ◽  
Author(s):  
Alon Krispin ◽  
Yaniv Bledi ◽  
Mizhir Atallah ◽  
Uriel Trahtemberg ◽  
Inna Verbovetski ◽  
...  
Keyword(s):  
Dendritic Cell ◽  
Apoptotic Cell ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Binding Domain ◽  
Apoptotic Cells ◽  
Heparin Binding ◽  
Proteomic Approach ◽  
Thrombospondin 1 ◽  
Heparin Binding Domain

Abstract Apoptotic cells were shown to induce dendritic cell immune tolerance. We applied a proteomic approach to identify molecules that are secreted from apoptotic monocytes, and thus may mediate engulfment and immune suppression. Supernatants of monocytes undergoing apoptosis were collected and compared using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and differentially expressed proteins were identified using tandem mass spectrometry. Thrombospondin-1 (TSP-1) and its cleaved 26-kDa heparin-binding domain (HBD) were identified. We show that TSP-1 is expressed upon induction of monocyte apoptosis in a caspase-dependent pattern and the HBD is cleaved by chymotrypsin-like serine protease. We further show that CD29, CD36, CD47, CD51, and CD91 simultaneously participate in engulfment induction and generation of an immature dendritic cell (iDC) tolerogenic and phagocytic state. We conclude that apoptotic cell TSP-1, and notably its HBD, creates a signalosome in iDCs to improve engulfment and to tolerate engulfed material prior to the interaction with apoptotic cells.

scholarly journals Structural domains of heparan sulphate for specific recognition of the C-terminal heparin-binding domain of human plasma fibronectin (HEPII)

1996 ◽  
Vol 317 (3) ◽  
pp. 871-877 ◽  
Author(s):  
Andrew WALKER ◽  
John T. GALLAGHER
Keyword(s):  
Human Plasma ◽  
Soft Tissues ◽  
Chondroitin Sulphate ◽  
Wide Spectrum ◽  
Heparan Sulphate ◽  
Binding Domain ◽  
Heparin Binding ◽  
Plasma Fibronectin ◽  
Heparin Binding Domain ◽  
And Migration

Heparan sulphate (HS) is an abundant polysaccharide component of the pericellular domain and is found in most soft tissues and all adherent cells in culture. It interacts with a wide spectrum of proteins including polypeptide growth factors and glycoproteins of the extracellular matrix. These interactions might influence fundamental cellular activities such as adhesion, growth and migration. HS might therefore represent a highly adaptive mechanism by which cells respond to their environment. The present study shows that the interaction between fibroblast HS, metabolically labelled with [3H]glucosamine, and the C-terminal heparin-binding domain of human plasma fibronectin (HEPII), is determined by distinct regions of the polysaccharide chain. By using a very sensitive affinity-chromatography method and specific polysaccharide scission it was shown that the HEPII-binding regions of HS reside within sulphated domains that are resistant to degradation by heparinase III. In addition, optimal binding was achieved with specific heparinase III-resistant fragments of 14–16 monosaccharides in length. The affinity of HS for HEPII was significantly decreased when the polysaccharide was cleaved with heparinase I. Chondroitin sulphate and dermatan sulphate were poor competitive inhibitors of [3H]HS binding to HEPII whereas unlabelled HS and heparin gave a strong inhibitory activity, with heparin being the most potent inhibitor. These findings suggest that the interaction between HEPII and HS is specific and requires extended sequences of seven to eight N-sulphated disaccharides in which a proportion of the iduronate residues are sulphated at C-2. The results have important implications for the functions of HS in cell adhesion and migration.

Plasmin degradation of insulin-like growth factor-binding protein-5 (IGFBP-5): regulation by IGFBP-5-(201—218)

1997 ◽  
Vol 273 (5) ◽  
pp. E996-E1004 ◽  
Author(s):  
Phil G. Campbell ◽  
Dennis L. Andress
Keyword(s):  
Growth Factor ◽  
Solid Phase ◽  
Binding Protein ◽  
Competitive Inhibitor ◽  
Binding Domain ◽  
Tissue Type ◽  
Insulin Like Growth Factor ◽  
Heparin Binding ◽  
Factor Binding ◽  
Heparin Binding Domain

Using the major bone insulin-like growth factor-binding protein (IGFBP) IGFBP-5, we took a mechanistic approach in evaluating the role of the heparin-binding domain of IGFBP-5 in regulating plasmin (Pm) proteolysis of IGFBP-5. Using synthetic IGFBP-5 peptide fragments, we determined that the heparin-binding domain, IGFBP-5-(208—218), inhibits Pm proteolysis of intact IGFBP-5. The mechanism of action of IGFBP-5-(201—218) was by inhibiting Pm binding to substrate IGFBP-5. IGFBP-5-(201—218) action was independent of site of proteolysis, fluid, or solid phase interaction. In addition, IGFBP-5-(201—218) was found to inhibit plasminogen (Pg) activation to Pm. IGFBP-5-(201—218) did not directly inhibit the activity of Pm, urokinase Pg activator (PA), or tissue-type PA but acted as a competitive inhibitor of Pg activation by PA, which is in contrast to the stimulating effect of heparin on Pg activation. These data indicate that the heparin-binding domain contains the serine protease (Pg-to-Pm) binding site region of IGFBP-5, and that this region, which is presumed to represent a Pm-induced proteolytic product of IGFBP-5, is capable of regulating Pm action.

A recombinant NH2-terminal heparin-binding domain of the adhesive glycoprotein, thrombospondin-1, promotes endothelial tube formation and cell survival: a possible role for syndecan-4 proteoglycan

2002 ◽  
Vol 21 (4) ◽  
pp. 311-324 ◽  
Author(s):  
Marianna A Ferrari do Outeiro-Bernstein ◽  
Sara Santana Nunes ◽  
Ana Carolina Magalhães Andrade ◽  
Tercia Rodrigues Alves ◽  
Chantal Legrand ◽  
...  
Keyword(s):  
Cell Survival ◽  
Tube Formation ◽  
Binding Domain ◽  
Heparin Binding ◽  
Endothelial Tube Formation ◽  
Thrombospondin 1 ◽  
Heparin Binding Domain

scholarly journals Expression and analysis of COOH-terminal deletions of the human thrombospondin molecule.

1989 ◽  
Vol 109 (2) ◽  
pp. 843-852 ◽  
Author(s):  
E V Prochownik ◽  
K O'Rourke ◽  
V M Dixit
Keyword(s):  
Extracellular Matrix ◽  
Amino Acid ◽  
Solid Phase ◽  
Binding Domain ◽  
Heparin Binding ◽  
Disulphide Bridge ◽  
Cell Growth And Migration ◽  
African Green Monkey Kidney ◽  
Amino Terminal ◽  
Heparin Binding Domain

Thrombospondin (TSP) is a homotrimeric extracellular glycoprotein with a subunit molecular mass of 140 kD. The subunits have a modular or domain-like structure and are held together by interchain disulphide bonds. A number of domains have been identified including those for the binding of collagen, fibrinogen, and heparin. Due to the trimeric form of the TSP molecule, the various domains are trivalent in nature and this contributes to the ability of TSP to mediate cell-substrate interactions. Indeed, TSP has recently been shown not only to promote cell adhesion but also to be intimately involved in cell growth and migration. The adhesive function of TSP is attributable to the "solid-phase" or matrix-bound form of the molecule. There is some evidence that the heparin-binding domain mediates incorporation of soluble TSP into the insoluble matrix form. The heparin-binding domain of TSP is a compact globular amino-terminal moiety that contains two clusters of basic amino acids and a single intrachain disulphide bond. To delineate the role of the heparin-binding domain in matrix assembly and to define further the precise region of interchain disulphide bonding that results in trimer formation, we have expressed deleted forms of the cDNA encoding TSP in SV-40-transformed. African green monkey kidney cells. The proteins synthesized from the various deleted TSP cDNAs were examined for (a) secretion into the culture medium and incorporation into the extracellular matrix; (b) binding to heparin-Sepharose; (c) immunoprecipitability by a conformation-specific monoclonal antibody; and (d) ability to form trimers. This analysis allowed us to draw the following conclusions. (a) A 218 amino acid NH2-terminal protein that preserves the intrachain disulphide bridge of the heparin-binding domain is capable of binding to heparin-Sepharose and incorporating into the extracellular matrix. (b) A shorter 164 amino acid NH2-terminal peptide that does not contain the intrachain disulphide bridge of the heparin-binding domain is neither able to bind to heparin-Sepharose nor able to incorporate into the extracellular matrix. (c) The region of interchain disulphide bridging necessary for trimer assembly resides within a cluster of seven cysteine residues immediately adjacent to the heparin-binding domain.

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