Variations in the chondroitin sulfate-protein linkage region of aggrecans from bovine nasal and human articular cartilages

Fang Cheng, Dick Heinegård, Lars Åke Fransson, Michael Bayliss, Julie Bielicki, John Hopwood, Keiichi Yoshida

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Aggrecan-derived chondroitin sulfate (CS) chains, released by β- elimination, were derivatized with p-aminobenzoic acid or p-aminophenol; radioiodinated; and subjected to graded or complete degradations by chondroitin ABC lyase to generate linkage region fragments of the basic structure ΔGlyUA-GalNAc-GlcUA-Gal-Gal-Xyl-R (where ΔGlyUA represents 4,5- unsaturated glycuronic acid, and R is the adduct), by chondroitin AC lyase to generate the shorter fragment ΔGlyUA-Gal-Gal-Xyl-R, or by chondroitin C lyase to generate the same fragment when it was linked to a 6-O-sulfated or unsulfated GalNAc at the nonreducing end. Fragments were separated by size using gel chromatography, by charge using ion-exchange chromatography, and by size/charge using electrophoresis and then characterized by stepwise degradations from the nonreducing end by using mercuric acetate to remove all terminal ΔGlyUA, by bacterial glucuronidase to remove the same residue when linked to unsulfated or 6-O-sulfated GalNAc/Gal, by mammalian 4-sulfatase to remove sulfate from terminal GalNAc 4-O-sulfate, by chondro-4-sulfatase to remove 4-O-sulfate from other GalNAc/Gal residues, and by β-galactosidase to remove terminal Gal. Results with CS from bovine nasal cartilage aggrecan show that, in nearly all chains, Xyl and probably also the first Gal are unsubstituted, whereas the second Gal is 4-O-sulfated in one CS chain out of five. The first disaccharide repeat is sulfated at C-4 of GalNAc in one chain out of three and unsulfated in the other two. A sulfated first disaccharide is always joined to an unsulfated GlcUA-Gal-Gal sequence. In contrast, CS from human articular cartilage usually has a sulfated first disaccharide repeat. In CS from young human cartilage, sulfate groups are mostly at C-4 of GalNAc in the major part of the chain, but st C-6 in the nonreducing distal portion. In CS from old cartilage, sulfation at C-6 of GalNAc is a major feature from the nonreducing end down to approximately positions 4 and 5 from the linkage region, where GalNAc 4-O-sulfate is common.

Original languageEnglish
Pages (from-to)28572-28580
Number of pages9
JournalJournal of Biological Chemistry
Issue number45
Publication statusPublished or Issued - 1996

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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