Post-translational events in proteoglycan synthesis: Kinetics of synthesis of chondroitin sulfate and oligosaccharides on the core protein

Research output: Contribution to journalArticle

Abstract

Chondrocytes isolated from the Swarm rat chondrosarcoma were incubated in culture with [1-3H]glucose for 30 min to 8 h. Labeled proteoglycans were isolated, treated with borohydride under alkaline conditions, and the three complex sugar structures purified: N- and O-linked oligosaccharides and chondroitin sulfate chains. The amount of incorporated radioactivity into each component sugar was analyzed by HPLC after enzyme digestion and hydrolysis. The kinetic data for labeling of each sugar over the time course of the experiment were fit to first-order rate equations and the half times (t 1 2) to linear labeling were calculated. The t 1 2 values were essentially the same, 5-8 min, for galactose in all three complex sugar structures and for chain glucuronic acid in chondroitin sulfate, while that for xylitol in chondroitin sulfate, 15.8 min, was significantly longer. Thus, oligosaccharide synthesis is concomitant with chondroitin sulfate chain synthesis; the addition of the chondroitin sulfate linkage galactose occurs at or nearly at the same time as chain elongation while the addition of linkage xylose residues to the core protein may precede chain synthesis by up to 8 min. Since the intracellular t 1 2 of the core protein precursor for these cells is 45 to 90 min, the data strongly suggest that the addition of xylose is not completed to any significant extent while the polypeptide is still nascent or shortly after its release into the rough endoplasmic reticulum. It is proposed that the addition of xylose to the core protein precursor is a late endoplasmic reticulum or early Golgi event. The analytical data were consistent with the presence of ester phosphate on about 80% of the xylose residues of the newly synthesized proteoglycan.

Details

Authors
  • Stefan Lohmander
  • Vincent C. Hascall
  • Masaki Yanagishita
  • Klaus E. Kuettner
  • James H. Kimura
Organisations
External organisations
  • Rush Presbyterian St. Luke's Medical Center
  • National Institutes of Health, United States
  • Lund University
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Cell and Molecular Biology
Original languageEnglish
Pages (from-to)211-227
Number of pages17
JournalArchives of Biochemistry and Biophysics
Volume250
Issue number1
Publication statusPublished - 1986
Publication categoryResearch
Peer-reviewedYes