Coupling of N-deacetylation and N-sulfation in a Chinese hamster ovary cell mutant defective in heparan sulfate N-sulfotransferase.
The coordination of N-deacetylation and N-sulfation of heparan sulfate was examined in wild-type Chinese hamster ovary cells and mutant pgsE-606. This mutant expresses about 3-fold less N-sulfotransferase activity, which causes the proportion of N-sulfated GlcN residues in heparan sulfate to decline from 39 to 21% of total GlcN (Bame, K.J., and Esko, J.D. (1989) J. Biol. Chem. 264, 8059-8065). In this report, we show that microsomes from pgsE-606 cells have about twice the N-deacetylase activity found in microsomes from wild-type cells. However, N-deacetylation in vivo was actually depressed since heparan sulfate preparations from the mutant contained very few unsubstituted GlcN residues and 2-fold less N-sulfated GlcN residues. Treatment of mutant cells with chlorate, a general inhibitor of sulfation, depressed adenosine 3'-phosphate-5'-phosphosulfate pools more than 10-fold and further reduced the extent of N-sulfation from 21% to less than 6% of total GlcN. Unsubstituted GlcN residues accumulated under these conditions to the extent that N-sulfated residues declined. Thus, N-deacetylation remained depressed in the mutant in the presence of chlorate. These findings show that N-deacetylation is regulated in vivo and support the idea that the activity of N-deacetylase may be linked to N-sulfotransferase.
Bame
K J
KJ
Department of Biochemistry, School of Medicine, University of Alabama, Birmingham 35294.
Reddy
R V
RV
Esko
J D
JD
D000107:Acetylation; D000581:Amidohydrolases / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002852:Chromatography, Ion Exchange; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D006025:Glycosaminoglycans / Q000378:metabolism; D006497:Heparan Sulfate / Q000378:metabolism*; D009154:Mutation; D010053:Ovary / Q000166:cytology / Q000201:enzymology; D015238:Sulfotransferases; D013466:Sulfurtransferases / Q000378:metabolism*
D000581:Amidohydrolases; D006025:Glycosaminoglycans; D006497:Heparan Sulfate; D015238:Sulfotransferases; D013466:Sulfurtransferases; C021350:heparitin sulfotransferase