Purification and characterization of beta-galactoside (alpha 2 leads to 6)sialyltransferase from rat liver and hepatomas.
Asialofetuin sialyltransferase from Triton X-100 extracts of rat liver was resolved by phosphocellulose chromatography into two fractions, designated I and II in order of elution. When previously treated with Arthrobacter ureafaciens neuraminidase, fraction I eluted at about the same position as II while no alteration occurred in II. Primary rat hepatomas contained only a single asialofetuin sialyltransferase, identical to fraction I in chromatographic behavior. Transferases I and II were purified to near homogeneity. Transferase II, as well as neuraminidase-treated I, could be sialylated auto-catalytically, indicating that the lack of sialic acid in II is not due to the lack of a sialic-acid-accepting site. Both enzymes formed an (alpha 2 leads to 6)sialylgalactoside linkage with asialo-glycoproteins of the glycosylamine-type and with lactose, and were indistinguishable immunologically. Nevertheless, the transferases exhibited different molecular weights of 37000 (I) and 43000 (II). When heated at 50 degrees C, transferase I lost half its original activity within 20 min while II was scarcely inactivated. Kinetically, transferase I showed three-times higher affinity than II for CMP-N-acetylneuraminic acid and for desialylated plasma membrane. Asialofetuin sialyltransferase was also purified from primary rat hepatoma. The purified enzyme was identical to transferase I in every respect examined. We conclude that hepatomas contain transferase I but lack transferase II.
D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D002845:Chromatography; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008114:Liver Neoplasms, Experimental / Q000201:enzymology*; D008297:Male; D008970:Molecular Weight; D009439:Neuraminidase; D051381:Rats; D012799:Sialyltransferases / Q000302:isolation & purification*; D014166:Transferases / Q000302:isolation & purification*; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
D014166:Transferases; D012799:Sialyltransferases; D009439:Neuraminidase; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase