Regulation of Serum Glycosaminoglycan Sulfotransferase Activities

Inhibition by Sulfated Glycosaminoglycans and Activation by Polyamines and Basic Peptides Including a Polylysine-Containing Segment of the c-Ki-ras 2 Protein

Abstract

The regulatory mechanisms for the glycosaminoglycan sulfotransferases in fetal calf serum were investigated. The enzymes examined were those which transfer sulfate from 3'-phosphoadenosine 5'-phosphosulfate to 1) position 6 of the internal N-acetylgalactosamine units of chondroitin, 2) position 6 of galactose units of keratan sulfate, and 3) position 2 (an amino group) of glucosamine units of heparan sulfate. The former two enzymes were activated by spermidine, spermine, protamine, and poly L-lysine. All the enzymes were strongly inhibited by heparin and dextran sulfate, whereas only the chondroitin 6-O-sulfotransferase was inhibited by sulfated galactosaminoglycans. The inhibition of this enzyme by the sulfated glycosaminoglycans was abolished by polylysine, indicating that the activation by polylysine is partly due to the neutralization of endogenous acidic inhibitors, including sulfated glycosaminoglycans. Affinity chromatographic studies demonstrated that heparin specifically binds to the three enzymes, which have anionic isoelectric points, and that chondroitin 6-sulfate, spermine, and polylysine bind to the former two enzymes under physiological conditions. Thus, the activation by spermine and polylysine as well as the inhibition by sulfated glycosaminoglycans also appears to occur through their binding to the enzymes. Studies with synthetic lysine oligomers and an affinity-purified (approximately 700-fold) fraction containing the former two enzymes indicated that the pentamer is the minimum unit required for the activation. A synthetic peptide, containing six consecutive lysines at the carboxy terminus of the human c-Ki-ras 2 protein, also regulated the two enzyme activities at micromolar concentrations. The possible physiological implications of the oberved effects of these regulatory substances on the glycosaminoglycan sulfotransferases are discussed in relation to glycosaminoglycan synthesis during the proliferation, differentiation, and transformation of cells. The possibility of sulfated glycosaminoglycans being enzyme regulators is also discussed.