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Thrombospondin-1 binds to polyhistidine with high affinity and specificity
V K Vanguri1, S Wang, S Godyna
1Department of Vascular Biology, Jerome H. Holland Laboratory, American Red Cross, Rockville, MD 20855, USA.
The Biochemical Journal
|April 6, 2000
Summary
Thrombospondin-1 (TSP1) specifically binds to polyhistidine stretches with high affinity. This interaction, mediated by TSP1's heparin-binding domain, suggests a molecular basis for TSP1's role in protein binding.
Area of Science:
- Biochemistry
- Molecular Biology
- Protein Interactions
Background:
- Thrombospondin-1 (TSP1) is a 450 kDa glycoprotein involved in cell growth, adhesion, and migration.
- TSP1 interacts with various cell surface receptors, growth factors, and extracellular matrix proteins.
- The precise molecular mechanisms underlying TSP1's diverse binding capabilities are not fully elucidated.
Purpose of the Study:
- To investigate the specific binding interactions of Thrombospondin-1 (TSP1).
- To determine if TSP1 binds to polyhistidine-containing proteins.
- To characterize the binding affinity and potential domains involved in TSP1-polyhistidine interactions.
Main Methods:
- Solid-phase binding assays using (125)I-labeled TSP1.
- Competition assays with histidine-containing peptides and other molecules (histidine, imidazole, heparin).
- Affinity chromatography utilizing a polyhistidine-peptide immobilized on agarose.
Main Results:
- TSP1 demonstrated specific binding to proteins containing polyhistidine stretches with an average dissociation constant of 5 nM.
- Binding was inhibited by histidine-containing peptides, with inhibition proportional to histidine number.
- Heparin acted as a potent inhibitor (K(i) = 50 nM), and a recombinant heparin-binding domain of TSP1 competed for polyhistidine binding.
- TSP1 from platelet releasates was identified as the major binding protein on a polyhistidine affinity column.
Conclusions:
- TSP1 possesses a high-affinity binding site for polyhistidine sequences.
- The heparin-binding domain of TSP1 is likely involved in the interaction with polyhistidine.
- This finding provides a molecular explanation for TSP1 binding to histidine-rich glycoprotein and suggests potential interactions with other polyhistidine proteins.