Post-translational Modifications in Cartilage Oligomeric Matrix Protein

Abstract

Analysis of the carboxymethylated subunit of human cartilage oligomeric matrix protein (COMP) by matrix- assisted laser desorption time-of-flight mass spectrome- try indicated a protonated molecular mass of 86949 ? 149 Da, compared with 83547.0 Da calculated from the sequence. Treatment with N-glycanase caused a reduc- tion in mass of 3571 ? 219 Da, but there was no loss of mass after treatment with O-glycanase or neuramini- dase. Peptides containing two putative sites of N-glyco- sylation were purified and characterized. Analysis of the masses of these after N-glycanase treatment indi- cated that one was substituted at Asn-101 with an oligo- saccharide of mass 1847.2 ? 6.6 Da, and the other was unsubstituted at Asn-124. The remaining site of attach- ment, at Asn-721, was, therefore, also substituted with an oligosaccharide of mass 1724?226 Da. Analysis of the total monosaccharide content by chemical methods in- dicated that there were no additional oligosaccharide substituents. The MALDI-TOF mass spectra of COMP from bovine fetal and adult cartilage were compared, indicating a more heterogeneous pattern of substitu- tion at Asn-101 in the fetal form. Since COMP is dis- tributed throughout the pericellular and territorial environments in developing cartilage but occupies the interterritorial zone in mature cartilage, these changes in glycosylation may allow for different intermolecular interactions. Cartilage