Biosynthesis of chondroitin sulfate. Organization of sulfation.
The potential relationship of an intact membrane organization for the synthesis of chondroitin and chondroitin 4-sulfate was examined after modification of a mouse mast cell microsomal system with the nonionic detergent, Triton X-100. The results indicated that Triton X-100 had no effect on the rate of polymerization but had a slight effect on the size of glycosaminoglycan chains. An "all or nothing" pattern of sulfation of newly formed chondroitin was obtained in both the presence and the absence of Triton X-100, and this pattern did not change whether sulfation was initiated concurrent with or subsequent to polymerization. Sulfation of exogenous [14C]chondroitin and exogenous proteo[3H]chondroitin by the microsomal system required Triton X-100 but still produced an all or nothing pattern rather than a random sulfation pattern. When a 100,000 x g supernatant fraction was utilized for sulfation of [14C]chondroitin or proteo[3H]chondroitin, Triton X-100 was not needed, and a partial sulfation pattern was obtained. However, it was similar to the all or nothing pattern in that it still produced two populations, with some chains nonsulfated and others approximately 50% sulfated. When chondroitin hexasaccharide was used with 3'-phosphoadenylylphospho[35S]sulfate, multiple GalNAc residues of the individual hexasaccharides were found to be sulfated. This was relatively independent of Triton X-100 or the concentration of the hexasaccharide acceptors. With soluble enzyme, sulfation of multiple GalNAc residues on the individual hexasaccharide molecules was even greater, so that trisulfated products were found. These results suggest that efficient sulfation of chondroitin is related to enzyme-substrate interaction more than to membrane organization.
Sugumaran
G
G
Connective Tissue Research Laboratory, Veterans Administration Outpatient Clinic, Boston, Massachusetts.
Silbert
J E
JE
D000818:Animals; D002460:Cell Line; D002807:Chondroitin / Q000031:analogs & derivatives*; D011508:Chondroitin Sulfate Proteoglycans / Q000378:metabolism*; D002809:Chondroitin Sulfates / Q000096:biosynthesis*; D002848:Chromatography, DEAE-Cellulose; D066298:In Vitro Techniques; D007700:Kinetics; D008407:Mast Cells / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D011509:Proteoglycans / Q000378:metabolism*; D015238:Sulfotransferases / Q000378:metabolism*
D011508:Chondroitin Sulfate Proteoglycans; D011509:Proteoglycans; D011092:Polyethylene Glycols; D017830:Octoxynol; D002807:Chondroitin; D002809:Chondroitin Sulfates; D015238:Sulfotransferases