Thrombospondin-4 binds specifically to both collagenous and non-collagenous extracellular matrix proteins via its C-terminal domains.
Full-length and truncated forms of rat thrombospondin-4 (TSP-4) were expressed recombinantly in a mammalian cell line and purified to homogeneity. Biochemical analysis revealed a limited proteolytic processing, which detaches the N-terminal heparin-binding domain from the rest of the molecule and confirmed the importance of the heptad-repeat domain for pentamerization. In electron microscopy the uncleaved TSP-4 was seen as a large central particle to which five smaller globules are attached by elongated linker regions. Binding of TSP-4 to collagens and to non-collagenous proteins could be detected in enzyme-linked immunosorbent assay-style ligand binding assays, by surface plasmon resonance spectroscopy, and in rotary shadowing electron microscopy. Although the binding of TSP-4 to solid-phase collagens was enhanced by Zn(2+), that to non-collagenous proteins was not. The interactions of TSP-4 with both classes of proteins are mediated by C-terminal domains of the TSP-4 subunits but do not require an oligomeric structure. Major binding sites for TSP-4 are located in or close to the N- and C-terminal telopeptides in collagen I, but additional sites are detected in more central regions of the molecule.
Narouz-Ott
L
L
Institute for Biochemistry, Medical Faculty, University of Cologne, Joseph-Stelzmann-Strasse 52, Cologne D-50931, Germany.
Maurer
P
P
Nitsche
D P
DP
Smyth
N
N
Paulsson
M
M
D000595:Amino Acid Sequence; D000818:Animals; D015374:Biosensing Techniques; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D003094:Collagen / Q000378:metabolism*; D018076:DNA, Complementary / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D006801:Humans; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D051381:Rats; D012867:Skin / Q000737:chemistry; D013329:Structure-Activity Relationship; D019699:Thrombospondins / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism
D015815:Cell Adhesion Molecules; D018076:DNA, Complementary; D016326:Extracellular Matrix Proteins; D019699:Thrombospondins; C101864:thrombospondin 4; D003094:Collagen; D015032:Zinc