Characterization of a pseudoachondroplasia-associated mutation (His587–>Arg) in the C-terminal, collagen-binding domain of cartilage oligomeric matrix protein (COMP)

B11 BAUMANN/SENGLE Spitznagel, L., Nitsche, D.P., Paulsson, M., Maurer, P., and Zaucke, F., Biochem J. 2004 Jan 15;377(Pt 2):479-87.


We have introduced a pseudoachondroplasia-associated mutation (His(587)–>Arg) into the C-terminal collagen-binding domain of COMP (cartilage oligomeric matrix protein) and recombinantly expressed the full-length protein as well as truncated fragments in HEK-293 cells. CD spectroscopy revealed only subtle differences in the overall secondary structure of full-length proteins. Interestingly, the mutant COMP did not aggregate in the presence of calcium, as does the wild-type protein. The binding site for collagens was recently mapped to amino acids 579-595 and it was assumed that the His(587)–>Arg mutation influences collagen binding. However full-length mutant COMP bound to collagens I, II and IX, and the binding was not significantly different from that of wild-type COMP. Also a COMP His(587)–>Arg fragment encompassing the calcium-binding repeats and the C-terminal collagen-binding domain bound collagens equally well as the corresponding wild-type protein. The recombinant fragments encompassing the C-terminal domain alone showed multiple bands following SDS/PAGE, although their theoretical molecular masses could be verified by MS. A temperature-induced conformational change was observed in CD spectroscopy, and negative-staining electron microscopy demonstrated that both wild-type and mutant proteins formed defined elongated aggregates after heating to 60 degrees C. Our results suggest that the His(587)–>Arg mutation is not itself deleterious to the structure and collagen-binding of COMP.



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