PURIFICATION AND CHARACTERIZATION OF BETA-GALACTOSIDE (ALPHA-2-]6)SIALYLTRANSFERASE FROM RAT-LIVER AND HEPATOMAS

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  • 1 January 1982
    • journal article
    • research article
    • Vol. 126  (2) , 253-261
Abstract
Asialofetuin sialyltransferase from Triton X-100 extracts of rat liver was resolved by phosphocellulose chromatography into 2 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 .fwdarw. 6)sialylgalactoside linkage with asialo-glycoproteins of the glycosylamine-type and with lactose, and were indistinguishable immunologically. Nevertheless, the transferases exhibited different MW of 37,000 (I) and 43,000 (II). When heated at 50.degree. C, transferase I lost half its original activity within 20 min, while II was scarcely inactivated. Kinetically, transferase I showed 3-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. Hepatomas apparently contain transferase I, but lack transferase II.