POSTTRANSLATIONAL MODIFICATIONS IN CARTILAGE OLIGOMERIC MATRIX PROTEIN - CHARACTERIZATION OF THE N-LINKED OLIGOSACCHARIDES BY MATRIX-ASSISTED LASER-DESORPTION IONIZATION TIME-OF-FLIGHT MASS-SPECTROMETRY

Analysis of the carboxymethylated subunit of human cartilage oligomeri c matrix protein (COMP) by matrix-assisted laser desorption time-of-fl ight mass spectrometry indicated a protonated molecular mass of 86949 +/- 149 Da, compared with 83547.0 Da calculated from the sequence, Tre atment with N-glycanase caused a reduction in mass of 3571 +/- 219 Da, but there was no loss of mass after treatment with O-glycanase or neu raminidase, Peptides containing two putative sites of N-glycosylation were purified and characterized, Analysis of the masses of these after N-glycanase treatment indicated that one was substituted at Asn-101 w ith an oligosaccharide of mass 1847.2 +/- 6.6 Da, and the other was un substituted at Asn-124, The remaining site of attachment, at Asn-721, was, therefore, also substituted with an oligosaccharide of mass 1724 +/- 226 Da. Analysis of the total monosaccharide content by chemical m ethods indicated that there were no additional oligosaccharide substit uents, The MALDI-TOF mass spectra of COMP from bovine fetal and adult cartilage were compared, indicating a more heterogeneous pattern of su bstitution at Asn-101 in the fetal form, Since COMP is distributed thr oughout the pericellular and territorial environments in developing ca rtilage but occupies the interterritorial zone in mature cartilage, th ese changes in glycosylation may allow for different intermolecular in teractions.