2-dimensional electrophoresis: detection of glycosylation and influence on spot pattern

• Published in: Methods in molecular biology (Clifton, N.J.) Vol. 446; p. 199
• Main Authors: Löster, Klemens, Kannicht, Christoph
• Format: Journal Article
• Language: English
• Published: United States 2008
• Subjects: Blotting, Western; Electrophoresis, Gel, Two-Dimensional - methods; Glycosylation
• ISSN: 1064-3745
• DOI: 10.1007/978-1-60327-084-7_14

The detailed characterization of complex protein mixtures as in samples from biological sources cannot be sufficiently performed by separation of polypeptides according to their molecular weight as is done by conventional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (1DE). For analysis of such samples, 2-dimensional gel electrophoresis (2DE) is the preferable methodological approach because it combines separation of polypeptides according to isoelectric properties and molecular weight as well. The resulting pattern of protein spots does not only provide information on composition of samples because of the complexity of a mixture of polypeptides. It delivers also a picture on the microheterogenity of polypeptides caused by post-translational modifications. These might be of natural or artificial type and occur during biosynthetic processing of a polypeptide or within industrial scale production. The presented method describes an experimental approach to investigate the influence of glycosylation in general and sialylation exclusively on spot pattern of proteins separated by 2DE.