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

Lohmander, Stefan LU ; Hascall, Vincent C.; Yanagishita, Masaki; Kuettner, Klaus E. and Kimura, James H. (1986) In Archives of Biochemistry and Biophysics 250(1). p.211-227
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... (More)

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. © 1986.

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organization
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type
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publication status
published
subject
in
Archives of Biochemistry and Biophysics
volume
250
issue
1
pages
17 pages
publisher
Academic Press
external identifiers
  • Scopus:0022491721
ISSN
0003-9861
DOI
10.1016/0003-9861(86)90719-8
language
English
LU publication?
yes
id
2fd19df4-f41e-4c31-8baa-2696995e3eb2
date added to LUP
2016-05-04 12:57:46
date last changed
2016-07-04 11:50:03
@misc{2fd19df4-f41e-4c31-8baa-2696995e3eb2,
  abstract     = {<p>Chondrocytes isolated from the Swarm rat chondrosarcoma were incubated in culture with [1-<sup>3</sup>H]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<sub> 1 2</sub>) to linear labeling were calculated. The t<sub> 1 2</sub> 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<sub> 1 2</sub> 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. © 1986.</p>},
  author       = {Lohmander, Stefan and Hascall, Vincent C. and Yanagishita, Masaki and Kuettner, Klaus E. and Kimura, James H.},
  issn         = {0003-9861},
  language     = {eng},
  number       = {1},
  pages        = {211--227},
  publisher    = {ARRAY(0xa2d8c88)},
  series       = {Archives of Biochemistry and Biophysics},
  title        = {Post-translational events in proteoglycan synthesis : Kinetics of synthesis of chondroitin sulfate and oligosaccharides on the core protein},
  url          = {http://dx.doi.org/10.1016/0003-9861(86)90719-8},
  volume       = {250},
  year         = {1986},
}