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Characterization of human thrombospondin-4
J Lawler1, K McHenry, M Duquette
1Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts 02115.
The Journal of Biological Chemistry
|February 10, 1995
Summary
Human thrombospondin-4 (TSP-4) is a pentameric extracellular protein. TSP-4 binds calcium and heparin, and its structure is calcium-dependent.
Area of Science:
- Biochemistry
- Molecular Biology
- Extracellular Matrix Proteins
Background:
- Thrombospondins (TSPs) are extracellular calcium-binding proteins regulating cell functions.
- Human TSP-4 characterization is essential for understanding its biological roles.
Purpose of the Study:
- To sequence and characterize full-length human thrombospondin-4 (TSP-4).
- To investigate the structural and functional properties of recombinant human TSP-4.
Main Methods:
- Full-length human TSP-4 cDNA sequencing.
- Recombinant protein expression in NIH3T3 fibroblasts and C2C12 myoblasts.
- Purification using heparin-Sepharose and antibody affinity chromatography.
- Electron microscopy and limited tryptic digestion for structural analysis.
Main Results:
- Human TSP-4 contains an RGD cell-binding sequence, unlike Xenopus TSP-4.
- Expressed TSP-4 shows reduced (140,000 Da) and apparent (550,000 Da) molecular weights.
- Electron microscopy reveals TSP-4 is a pentameric protein with globular domains.
- TSP-4 exhibits calcium-dependent structural changes and resistance to tryptic digestion in the presence of calcium.
Conclusions:
- Human TSP-4 is a pentameric protein that binds heparin and calcium.
- Calcium binding influences TSP-4 structure and stability.
- The RGD sequence suggests a role in cell adhesion and migration.