Cytokeratin 13 contains O-glycosidically linked N-acetylglucosamine residues |
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Authors: | I A King E F Hounsell |
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Institution: | Dermatology Research Group, Medical Research Council Clinical Research Center, Harrow, United Kingdom. |
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Abstract: | The glycosylation of human cytokeratins was investigated in cultured human keratinocytes and A431 cells by metabolic labeling with 3H]glucosamine. In the presence of tunicamycin, keratinocytes incorporated 3H]glucosamine into a vitamin A-regulated acidic 53-kDa component of the cytoskeleton which was identified as cytokeratin 13 by one- and two-dimensional immunoblotting with specific monoclonal antibodies. This cytoskeletal component was also labeled with 3H]glucosamine in A431 cells but not in KB cells, which do not express cytokeratin 13. Its labeling was resistant to tunicamycin, suggesting that 3H]glucosamine had not been incorporated into N-linked oligosaccharides. Acid hydrolysis followed by paper and ion-exchange chromatography showed that the radioactivity in electrophoretically purified cytokeratin 13 was still present as glucosamine. Radioactivity was completely removed by treatment with beta-N-acetylglucosaminidase, suggesting that it was present in terminal N-acetylglucosamine residues. The labeled carbohydrate was released by alkaline borohydride treatment and was bound by a phenylboronic acid column, indicating an O-glycosidic linkage. On Bio-Gel P-2 columns, the beta-eliminated carbohydrate co-eluted with authentic N-acetylglucosaminitol. The results indicate that cytokeratin 13 contains single residues of N-acetylglucosamine O-glycosidically linked to the polypeptide chain. |
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