The C-terminal domain of cartilage matrix protein assembles into a triple-stranded alpha-helical coiled-coil structure.
Cartilage matrix protein (CMP) is a major component of different cartilages and consists of a disulfide-linked homotrimer. To test whether the C-terminal region forms a three-stranded alpha-helical coiled-coil, we synthesized a peptide, CMP-C36, corresponding to the last 36 residues of human CMP. Analytical ultracentrifugation revealed that CMP-C36 forms a homotrimer under physiological conditions. The sedimentation coefficient of 1.12 S is consistent with a rod-shaped molecule of 5.8 nm length, suggesting a lateral packing of three peptide chains. Depending on conditions, circular dichroism spectroscopy showed 75 to 96% alpha-helical content. The shapes of the spectra are characteristic for a coiled-coil structure. Thermal and guanidine-HCI-induced denaturation revealed a high degree of cooperativity and high stability. The concentration dependence of the melting temperature suggest a two-state transition. The trimer is stabilized by increasing the ionic strength above 130 mM salt, above which six ions are released upon unfolding. The peptide characteristics make it very likely that the C-terminal domain serves as the trimerization site of CMP. The two cysteine residues preceding this sequence region might stabilize the complex after assembly.
Beck
K
K
Shriners Hospital for Crippled Children, Research Unit, Portland OR 97201, USA.
Gambee
J E
JE
Bohan
C A
CA
Bächinger
H P
HP
D000595:Amino Acid Sequence; D002356:Cartilage; D064236:Cartilage Oligomeric Matrix Protein; D002942:Circular Dichroism; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics; D019791:Guanidine; D006146:Guanidines; D006801:Humans; D064235:Matrilin Proteins; D008969:Molecular Sequence Data; D015394:Molecular Structure; D008970:Molecular Weight; D009994:Osmolar Concentration; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013816:Thermodynamics
D064236:Cartilage Oligomeric Matrix Protein; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D006146:Guanidines; D064235:Matrilin Proteins; C577553:TSP5 protein, human; D019791:Guanidine