Destruction of cholera toxin receptor on HeLa cell membrane using microbial endoglycoceramidase.
The sensitivity of HeLa cells to cholera toxin decreased by Corynebacterium sp. endoglycoceramidase treatment. This endo-enzyme destroyed the cholera toxin receptor, ganglioside G(M1), on the cell surface membrane by liberating intact oligosaccharide from it, which was confirmed by the decrease of intracellular cAMP accumulation and the results of the analysis of released oligosaccharide with a combination of pyridylamination method and HPLC. Fluorescence microscopy using the immunofluorescence method revealed that the amount of cholera toxin attached to the cells decreased in endoglycoceramidase-treated cells. The enzyme acted on cellular glycosphingolipids without addition of any activator protein which is required by other similar enzymes. Corynebacterium endoglycoceramidase is a useful tool to elucidate the function of glycosphingolipids on the cell surface in situ.
Yamamoto
K
K
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
Nagano
T
T
Kumagai
H
H
Okamoto
Y
Y
Otani
S
S
D002462:Cell Membrane / Q000378:metabolism*; D002772:Cholera Toxin / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D003352:Corynebacterium / Q000201:enzymology; D000242:Cyclic AMP / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D005677:G(M1) Ganglioside / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D009844:Oligosaccharides / Q000378:metabolism; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism*
D009844:Oligosaccharides; D011956:Receptors, Cell Surface; C023552:choleragen receptor; D005677:G(M1) Ganglioside; D002772:Cholera Toxin; D000242:Cyclic AMP; D006026:Glycoside Hydrolases; C050427:endoglycoceramidase