Ganglioside glycosyltransferase assay using ion-exchange chromatography.
A rapid method for determination of ganglioside glycosyltransferase activity has been developed employing ion-exchange chromatography. Using 13-day chick brain as a source of uridine diphospho-N-acetyl-D-galactosamine: ganglioside GM3 N-acetylgalactosaminyltransferase (ganglioside GM2 synthetase), we were able to accurately measure transfer of N-[3H]acetylgalactosamine (GalNAc) from UDP-[3H]GalNAc to GM3 by application of the reaction mixture to small columns of DEAE-Sepharose and elution of radiolabeled GM2 reaction product with 10 mM potassium acetate in methanol. This method proved to be as reliable and sensitive as previously published assays but requires less time and fewer manipulations.
Walton
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Schnaar
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D000818:Animals; D001921:Brain / Q000201:enzymology; D002642:Chick Embryo; D002852:Chromatography, Ion Exchange; D005700:Galactosyltransferases / Q000032:analysis*; D008861:Microsomes / Q000201:enzymology; D017350:N-Acetylgalactosaminyltransferases; D000097763:Polypeptide N-acetylgalactosaminyltransferase
D005700:Galactosyltransferases; D017350:N-Acetylgalactosaminyltransferases; D000097763:Polypeptide N-acetylgalactosaminyltransferase; C023220:(N-acetylneuraminyl)-galactosylglucosylceramide N-acetylgalactosaminyltransferase