Overexpression of different isoforms of glucosaminyl N-deacetylase/N-sulfotransferase results in distinct heparan sulfate N-sulfation patterns.
Functional interactions of heparan sulfate (HS) with selected proteins depend on distinct saccharide sequences which are generated during biosynthesis of the polysaccharide. Glucosaminyl N-deacetylase/N-sulfotransferases (NDSTs) catalyze both the N-deacetylation and N-sulfation reactions that initiate the modification of the (GlcNAc-GlcA)(n) polysaccharide backbone. The N-acetyl/N-sulfate exchange is restricted to certain regions of the polysaccharide chains, and only these can be further modified by glucuronyl C5-epimerization and O-sulfation at various positions. To investigate whether NDST isoforms influenced differently the structure of HS, murine NDST-1 was overexpressed in human kidney 293 cells, and the structure of the HS produced was compared to HS from NDST-2 overexpressing cells [Cheung, W. F., Eriksson, I., Kusche-Gullberg M., Lindahl, U., and Kjellén, L. (1996) Biochemistry 35, 5250-5256]. The level of N-sulfation increased from 40% in control cells to 60% and 80%, respectively, in NDST-1 and NDST-2 transfected cells. Interestingly, the increase in N-sulfation was accompanied by an increased chain length, while no effect on IdoA content or O-sulfation was seen. The most extended N-sulfated domains were found in HS synthesized by NDST-2 transfected cells. Since both the N-deacetylase and the N-sulfotransferase activities were lower in these cells than in the NDST-1 overexpressing cells, we conclude that, in addition to the level of enzyme expression, the NDST isoform also is important in determining the N-sulfation pattern in HS.
Pikas
D S
DS
Department of Medical Biochemistry and Microbiology, University of Uppsala, Uppsala, Sweden.
Eriksson
I
I
Kjellén
L
L
D000107:Acetylation; D000581:Amidohydrolases / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D002850:Chromatography, Gel; D004187:Disaccharides / Q000032:analysis / Q000737:chemistry; D020723:Glucuronic Acid / Q000032:analysis; D006497:Heparan Sulfate / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007067:Iduronic Acid / Q000032:analysis; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008970:Molecular Weight; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013431:Sulfates / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D015238:Sulfotransferases / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D000581:Amidohydrolases; D004187:Disaccharides; D020723:Glucuronic Acid; D006497:Heparan Sulfate; D007067:Iduronic Acid; D007527:Isoenzymes; D011134:Polysaccharides; D012333:RNA, Messenger; D013431:Sulfates; D015238:Sulfotransferases; C113066:NDST2 protein, human; C491855:Ndst2 protein, mouse; C021350:heparitin sulfotransferase