Identification and characterization of protein glycosylation using specific endo- and exoglycosidases.
Protein Expression in Mammalian Cells
Methods in molecular biology, September 2011
Paula Magnelli, Alicia Bielik, Ellen Guthrie, Magnelli, Paula, Bielik, Alicia, Guthrie, Ellen
James L Hartley
Enzymatic deglycosylation followed by SDS-PAGE is a valuable method to detect glycan modifications on protein samples. Specific glycosidases were used to remove sugars from glycoproteins in a controlled fashion leaving the protein core intact; the resulting change in molecular weight could be detected as shifts in gel mobility. Alternatively, glycan-sensitive reagents were used to visualize the intensity of glycoprotein bands before and after enzyme treatment. The ease of use of these techniques, which require only basic laboratory instrumentation and reagents, makes them the methodology of choice for initial glycobiology studies. These protocols are also well suited to screen for optimal expression conditions, since multiple glycoprotein samples can be processed at once.
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