Characterization of a chondroitin sulfate proteoglycan synthesized by monkey arterial smooth muscle cells in vitro.
A monoclonal antibody against arterial smooth muscle cell chondroitin sulfate proteoglycan has been developed. Incubation of [35S]-methionine labeled proteoglycans with MAb 941 quantitatively immunoprecipitated all the chondroitin sulfate proteoglycan (CSPG) synthesized by these cells. Digestion of the immunoprecipitate with chondroitin AC lyase revealed one major protein band (Mr 420,000) and two minor bands (Mr 509,000 and 390,000) on SDS-PAGE that are composed of very similar peptides when analyzed by limited peptide digestion by S. aureus V8 protease. Additional studies demonstrated that this monoclonal antibody recognized an epitope on the chondroitin sulfate chains. However, only a minor subpopulation (5-12%) of the alkaline-borohydride released glycosaminoglycan chains was immunoprecipitated and this subset of chains was slightly larger than the non-immunoprecipitated chains. High pressure liquid chromatography analysis of the disaccharides generated from the immunoprecipitated glycosaminoglycan chains demonstrated that these chains were enriched in chondroitin-6-sulfate relative to chondroitin-4-sulfate (2:1) while that of the non-immunoprecipitated chains had a ratio of 1:1. These studies indicate that at least two distinct pools of chondroitin sulfate chains are present on all the chondroitin sulfate proteoglycan synthesized by arterial smooth muscle cells: a major population (89-95%) containing 6-sulfate and 4-sulfate in relatively equal proportion and a minor population (5-12%) which is hydrodynamically larger with a 6-sulfate to 4-sulfate ratio of 2:1.
Yeo
T K
TK
Department of Pathology, School of Medicine, University of Washington, Seattle 98195.
Macfarlane
S
S
Wight
T N
TN
D000818:Animals; D000911:Antibodies, Monoclonal; D002808:Chondroitin Lyases / Q000378:metabolism; D011508:Chondroitin Sulfate Proteoglycans / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D000939:Epitopes / Q000276:immunology; D007163:Immunosorbent Techniques; D008254:Macaca nemestrina; D008970:Molecular Weight; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism
D000911:Antibodies, Monoclonal; D011508:Chondroitin Sulfate Proteoglycans; D000939:Epitopes; D010446:Peptide Fragments; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D002808:Chondroitin Lyases