Post-translational 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 oligomeric matrix protein (COMP) by matrix-assisted laser desorption time-of-flight mass spectrometry indicated a protonated molecular mass of 86949 +/- 149 Da, compared with 83547.0 Da calculated from the sequence. Treatment 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 neuraminidase. 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 with an oligosaccharide of mass 1847. 2 +/- 6.6 Da, and the other was unsubstituted 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 methods indicated 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 substitution at Asn-101 in the fetal form. Since COMP is distributed 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.
Zaia
J
J
Osiris Therapeutics Inc., Baltimore, Maryland 21231, USA.
Boynton
R E
RE
McIntosh
A
A
Marshak
D R
DR
Olsson
H
H
Heinegârd
D
D
Barry
F P
FP
D000595:Amino Acid Sequence; D000818:Animals; D002356:Cartilage; D064236:Cartilage Oligomeric Matrix Protein; D002417:Cattle; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins / Q000378:metabolism*; D006801:Humans; D013058:Mass Spectrometry; D064235:Matrilin Proteins; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational
D064236:Cartilage Oligomeric Matrix Protein; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D064235:Matrilin Proteins; C577553:TSP5 protein, human