Identification of chondroitin sulfate E in human lung mast cells.
Human lung mast cells (HLMC) enriched up to 99% purity by counter current elutriation and density gradient centrifugation were labeled with 35S-sulfate to determine cell-associated proteoglycans. The 35S-labeled proteoglycans were extracted by the addition of detergent and 4 M guanidine-HCl, and separated from unincorporated precursor by Sephadex G-50 chromatography. 35S-Proteoglycans chromatographed over Sepharose 4B with a Kav of 0.48. 35S-Glycosaminoglycans separated from the parent 35S-proteoglycans by beta-elimination and chromatographed over Sepharose 4B with a Kav of 0.63. Characterization of 35S-proteoglycans by chondroitin ABC lyase treatment revealed approximately 36% of the proteoglycan to be composed of chondroitin sulfates. Analysis by HPLC of component disaccharides liberated by chondroitin ABC lyase using an amino-cyano-substituted silica column indicated that the chondroitin sulfates consisted of the monosulfated A disaccharide (GlcUA----GaINAc4SO4) (75%) and the over-sulfated E disaccharide (GlcUA----GaINAc4,6-diSO4) (25%). Nitrous acid/heparinase-susceptible heparin proteoglycans accounted for approximately 62% of the total 35S-proteoglycans present in the HLMC. Proteoglycans remaining after exposure of the original proteoglycan extract to either heparinase or chondroitin ABC lyase were of similar size, suggesting that the majority of heparin and chondroitin sulfate glycosaminoglycans were on separate protein cores. Proteoglycans extracted from HLMC were protease insensitive. Hence, in addition to heparin proteoglycans, HLMC synthesize a hitherto unrecognized quantity of chondroitin sulfate E proteoglycans.
Thompson
H L
HL
Mast Cell Physiology Section, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892.
Schulman
E S
ES
Metcalfe
D D
DD
D002808:Chondroitin Lyases; D019763:Heparin Lyase; D006801:Humans; D008168:Lung / Q000166:cytology*; D008407:Mast Cells / Q000032:analysis*; D011133:Polysaccharide-Lyases; D011509:Proteoglycans / Q000032:analysis
D011509:Proteoglycans; D002808:Chondroitin Lyases; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase