Deglycosylation of chondroitin sulfate proteoglycan by hydrogen fluoride in pyridine.
The original deglycosylation procedure using HF/pyridine has been modified for maximal removal of carbohydrate from chondroitin sulfate proteoglycan, with minimal alteration of the core protein. Gas-liquid chromatography analysis after treatment for various times showed that 95% of xylose and mannose and 70-85% of other sugars were removed within 30 min, indicating that almost all chondroitin sulfate chains and about 80% of N- and O-linked oligosaccharides were removed. In contrast to the loss of carbohydrate, no change in amino acid composition or loss of immunoreactivity occurred. Longer treatment of up to 16 h resulted in little additional removal of carbohydrate, but did cause a significant decrease in solubility and recovery of the deglycosylated product. Optimal removal of xylose residues after about 1 h was also shown by maximal acceptor activity of the product in a xylosyltransferase assay. Rapid removal of the HF reagent by vacuum evacuation and ion-exchange chromatography, coupled with the reduced time of treatment allowed recovery of an intact, homogenous protein core that is amenable to structural and sequence studies.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002241:Carbohydrates / Q000032:analysis*; D055598:Chemical Phenomena; D002621:Chemistry; D011508:Chondroitin Sulfate Proteoglycans / Q000032:analysis*; D002813:Chondrosarcoma / Q000032:analysis; D002849:Chromatography, Gas; D006858:Hydrofluoric Acid; D010430:Pentosyltransferases; D011509:Proteoglycans / Q000032:analysis*; D011725:Pyridines; D011863:Radioimmunoassay; D051381:Rats; D000097608:UDP Xylose-Protein Xylosyltransferase
D000596:Amino Acids; D002241:Carbohydrates; D011508:Chondroitin Sulfate Proteoglycans; D006858:Hydrofluoric Acid; D010430:Pentosyltransferases; D011509:Proteoglycans; D011725:Pyridines; D000097608:UDP Xylose-Protein Xylosyltransferase