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1. Heparan sulphates from normal 3T3 fibroblasts are association-prone as indicated by their affinity for agarose gels substituted with cognate heparan sulphate species. Heparan sulphates from SV40-transformed or polyoma-virus-transformed cells have no affinity for the same gels. 2. Heparan sulphates from the medium, the pericellular and intracellular pools of normal, SV40-transformed and polyoma-transformed 3T3 cells were separated into four subfractions (HS1-HS4) by ion-exchange chromatography. In general, HS1-HS3 were found in cell-derived heparan sulphates, whereas HS3-HS4 were present in the medium. The heparan sulphates from transformed cells were more heterogeneous and of lower charge density than those from the normal counterpart. 3. Degradations via periodate oxidation/alkaline elimination yielded the oligomers glucosamine-(hexuronate-glucosamine)(n)-R with n=1-5 and a large proportion of N-sulphate groups. There was a large contribution of fragments n=4-5 from heparan sulphates of normal cells. These fragments were less common in low-sulphated heparan sulphates of transformed cells. In the case of medium-drived heparan sulphates all species had a low content of fragments n=4-5. 4. The size distribution of (glucuronate-N-acetylglucosamine)(n) regions was assessed after deaminative cleavage. It was broad and ranged from n=1-10 for all heparan sulphate species. In the case of medium-derived heparan sulphates there were distinct differences between normal and transformed cells. In the latter chains the N-acetyl-rich segments were both shorter and longer than in the normal case. The shape of the disaccharide peak was consistent with a lower content of O-sulphate in the heparan sulphates from transformed cells. 5. It was concluded that heparan sulphates from medium or transformed cells exhibit the greatest structural deviation from the normal case. The finding of lower proportions of extended, iduronate/glucuronate-bearing, N-sulphate-rich segments in heparan sulphates of transformed cells was particularly interesting in view of the fact that these elements have been associated with ability to self-interact.

The mechanisms by which phosphate regulates the activity of alkaline phosphatase (orthophosphoric monoester phosphohydrolase, EC 3.1.3.1) in rat kidney were investigated. Measurements of incorporation of [(14)C]leucine into kidney alkaline phosphatase in rats fed on complete or phosphate-free diet provide evidence of a twofold increase in the rate of synthesis of the enzyme in diet-treated animals. Cycloheximide experiments indicated that control and diet-adapted enzyme decreases in activity according to first-order kinetics with a calculated half-life of 10.3 and 6.5h after complete and phosphate-free diet administration respectively. Basal and diet-adapted enzymes exhibit similar K(m) values for several phosphomonoesters and an identical degree of inhibition is produced by cysteine. In addition, the enzyme from both sources is the same with regard to heat inactivation at 45, 56 or 64 degrees C, to the profile of elution from Sephadex and to electrophoretic properties on polyacrylamide gel. A failure of rat kidney alkaline phosphatase to respond to cortisol (hydrocortisone) was also observed.