Identification and characterization of the immunoprecipitating medium- and low-density tryptic fragments of bovine nasal cartilage proteoglycan.
The fragments responsible for the immunodiffusion reactivity of middle- and low-density fractions of trypsin-digested bovine nasal cartilage proteoglycan have been identified and obtained in relatively homogeneous fractions. Glycosaminoglycan-bearing tryptic fragments were isolated from 4 M guanidinium chloride extracts of cartilage by ion-exchange chromatography and fractionated by dissociative equilibrium density gradient ultracentrifugation at a starting density of 1.50. Fragments in the middle fractions of the density gradient were digested with chondroitinase ABC and subfractionated by Sepharose 6B column chromatography. Middle-density subfractions contained fragments which were chemically and immunologically identical to those in high-density fragment subfractions of similar elution from Sepharose 6B. The middle-density subfractions contained two additional immunoprecipitating fragments. One, with alanine as N-terminal amino acid, was isolated by virtue of its retention by a column of concanavalin A-Sepharose 4B and its resistance to digestion with keratanase; the second was concentrated in a subfraction whose elution from concanavalin A-Sepharose 4B was retarded. The gradient fraction of lowest density contained fragments with the properties of the major tryptic fragments of the hyaluronic acid-binding segment of the proteoglycan monomer and the link proteins. These were recovered as a complex in the void volume upon Sepharose gel chromatography in saline-buffer and were resolved into relatively homogeneous fractions by column chromotography on CL-Sepharose 6B in 4 M guanidinium chloride. In all, tryptic digests of cartilage proteoglycan contain at least seven different immunoprecipitating fragments, some of which may not have been correctly identified previously.
Keiser
H D
HD
Keyserman
F Z
FZ
D000818:Animals; D002356:Cartilage / Q000032:analysis*; D002417:Cattle; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005779:Immunodiffusion; D009666:Nose; D010446:Peptide Fragments / Q000032:analysis*; D011509:Proteoglycans / Q000032:analysis*; D014357:Trypsin / Q000378:metabolism*
D010446:Peptide Fragments; D011509:Proteoglycans; D014357:Trypsin