Influence of chlorate on proteoglycan biosynthesis by cultured human fibroblasts.
The influence of chlorate, an inhibitor of sulfate adenylyltransferase, on biosynthesis and secretion of proteoglycans was investigated in cultured human skin fibroblasts. At up to 10 mM concentrations, chlorate caused a reduction of [35S]sulfate incorporation into small chondroitin sulfate/dermatan sulfate proteoglycan by up to 96%. Incorporation of [3H]leucine and [3H] glucosamine was only slightly affected. No influence was seen on the polymerization degree of the polysaccharide chain as judged by gel filtration, and on the kinetics of secretion of the proteoglycan. Concomitant with reduced sulfation, however, was an increased sensitivity toward chondroitin AC lyase which suggests a diminished epimerization of D-glucuronic acid to L-iduronic acid residues. Agarose gel electrophoresis revealed that all polysaccharide chains of control chondroitin sulfate/dermatan sulfate proteoglycan exhibited a similar sulfation degree. Chlorate treatment led to the formation of polysaccharide chains of widely varying degree of sulfation, but fully sulfated chains were synthesized even in the presence of 3 mM chlorate, and sulfate-free chondroitin was not detected. Studying the effects of chlorate treatment on the synthesis of other proteoglycan types it was found that, in cell-associated galactosaminoglycans, 6-sulfation of N-acetylgalactosamine residues was less affected than was 4-sulfation. In case of heparan sulfate the synthesis of sulfamate groups was less impaired than sulfate ester formation. Nitrous acid degradation at pH 4.1 indicated the presence of unsubstituted amino groups. Chlorate treatment may be considered as a means for the production of proteoglycans with defined structural alterations.
Department of Physiological Chemistry and Pathobiochemistry, University of Münster, Federal Republic of Germany.
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Greve
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Cully
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Blumberg
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Kresse
D000328:Adult; D002478:Cells, Cultured; D002648:Child; D002704:Chlorates / Q000494:pharmacology*; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D005944:Glucosamine / Q000378:metabolism; D006801:Humans; D007930:Leucine / Q000378:metabolism; D011509:Proteoglycans / Q000096:biosynthesis*; D013430:Sulfate Adenylyltransferase / Q000037:antagonists & inhibitors; D013431:Sulfates / Q000378:metabolism
D002704:Chlorates; D011509:Proteoglycans; D013431:Sulfates; D013430:Sulfate Adenylyltransferase; D007930:Leucine; D005944:Glucosamine