Partial purification and characterization of 3'-phosphoadenylylsulfate:keratan sulfate sulfotransferases.
Two 3'-phosphoadenylylsulfate:keratan sulfate sulfotransferases were purified 600-fold and 340-fold, respectively, from isolated bovine cornea cells. Sulfotransferase I exhibited an apparent Mr = 220,000, whereas an Mr = 140,000 was calculated for sulfotransferase II. The final preparations were both devoid of chondroitin sulfate sulfotransferase activity. The position of sulfation was determined by proton nuclear magnetic resonance spectroscopy. Sixty per cent of the sulfate ester groups formed by sulfotransferase I were linked to the C-6 atom of galactosyl residues, the other ones to the C-6 atom of N-acetylglucosamine. Sulfotransferase II showed a different specificity: 23% of the newly formed sulfate ester groups were on galactosyl and 77% on N-acetylglucosaminyl residues. Both sulfotransferase preparations acted in a cooperative manner. In the presence of both sulfotransferases, the incorporation of [35S]sulfate into keratan sulfate was up to 75% higher than could be expected from the sum of individual activities. From the specific radioactivities of the oligosaccharides produced by digestion with endo-beta-galactosidase, it was also concluded that both enzyme species reacted best with keratan sulfate segments exhibiting a relatively high degree of sulfation.
D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003315:Cornea / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D007632:Keratan Sulfate / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008970:Molecular Weight; D010724:Phosphoadenosine Phosphosulfate / Q000378:metabolism; D051381:Rats; D013431:Sulfates / Q000378:metabolism; D015238:Sulfotransferases; D013466:Sulfurtransferases / Q000302:isolation & purification*; D000097811:Carbohydrate Sulfotransferases
D007632:Keratan Sulfate; D010724:Phosphoadenosine Phosphosulfate; D013431:Sulfates; D015238:Sulfotransferases; D013466:Sulfurtransferases; D000097811:Carbohydrate Sulfotransferases