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Glycobiology Advance Access published online on December 17, 2002
Glycobiology, doi:10.1093/glycob/cwg028
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Submitted on August 5, 2002
Revised on November 2, 2002
Accepted on November 6, 2002

© 2002 Oxford University Press

ORIGINAL ARTICLES

The contribution of tandem repeat number to the O-Glycosylation of mucins

Howard Scott Silverman 1, Mark Sutton-Smith 2, Kimberley McDermott 3, Paul Heal 1, Shih-Hsing Leir 1, Howard R Morris 2, Michael A Hollingsworth 3, Anne Dell 2, Ann Harris 1*

1 Paediatric Molecular Genetics, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, OX3 9DS, UK
2 Department of Biological Sciences, Imperial College of Science, Technology and Medicine, South Kensington, London SW7 2AZ, UK
3 Eppley Institute, University of Nebraska Medical Center, Omaha NE, 68198-6805, USA

* To whom correspondence should be addressed. E-mail: aharris{at}molbiol.ox.ac.uk.

Abstract

The serine and threonine-rich tandem repeat (TR) units that comprise the characteristic feature of mucin glycoproteins are often polymorphic, with substantial genetic variation in TR number. The precise effect of TR number on O-glycosylation is not fully understood, though the TR number of several mucins may be associated with apparent susceptibility to certain human diseases. To evaluate the contribution of TR number to O-glycosylation we generated a series of chimeric mucins carrying increasing numbers of TR units from the MUC5B mucin in the context of an epitope-tagged MUC1 mucin backbone. These mucins were expressed in Caco2 colon carcinoma cell clones and purified by immunoprecipitation. O-glycosylation was investigated by western blotting with antibodies to known carbohydrate structures and by Fast Atom Bombardment - Mass Spectrometry. Additional carbohydrate epitopes were detected with antibodies on chimeric mucins with a higher TR number in comparison to those with fewer TRs. Using mass spectrometry higher molecular weight glycans were detected more frequently on the mucins with extended TRs compared to those with fewer TRs. However no novel carbohydrate structures were seen, suggesting TR number does not affect the specificity of O-glycosylation.


Key words: Mucin tandem repeat, O-glycosylation/MUC5B.
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