Structure and function of collagen type X
The primary structure of collagen type X, a hypertrophic cartilage-specific molecule, has been studied at the protein level. The helical domain of collagen type X was biochemically characterized in detail using traditional automated sequencing and novel, mass spectrometry-based, sequencing techniques; namely, peptide ladder sequencing and skimmer collision-induced dissociation electrospray mass spectrometry. Methods were developed for and adapted to the analysis of collagen glycosides, the isolation of peptides containing lysyl oxidase-mediated crosslinks from growth plate cyanogen bromide peptides, and the analysis of reducible lysine-derived crosslinks. Collagen type X was shown to contain relatively high levels of hydroxyproline and hydroxylysine. One residue of 3-hydroxyproline was discovered, located at the N-terminal end of the helix. The hydroxylysine residues appeared to be almost fully glycosylated with glucosyl-galactosyl-hydroxylysine, imparting increased hydrophilicity to the helical part of the molecule. The glycosides were concentrated within the C-terminal half of the helix. Both intermediate crosslinks, dihydroxylysinonorleucine and hydroxylysinonorleucine, were present in collagen type X in a 2:1 ratio. One in five triple-helical molecules became crosslinked by their C-terminal cyanogen bromide peptide with the most likely crosslinking partner being collagen type II within heterotypic cartilage fibrils. However, crosslinking partner(s) of collagen type X were not fully characterized. The function of collagen type X is not fully understood. There is substantial evidence that collagen type X is essential for preparing the matrix for calcification and resorption by diffusing into the interterritorial matrix and altering its properties. We hypothesize that, besides the rod-like shape and small size of the molecule, extensive glycosylation of the collagen type X helical domain is an important prerequisite for rapid diffusion through the extracellular matrix.