Document Detail


Immunodetection of alpha 1-3 fucosyltransferase (FucT-V).
MedLine Citation:
PMID:  8738418     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The fucosyltransferases constitute a family of glycosyltransferases incorporating fucose residues into glycoprotein or glycolipid glycans. They afford one of the possible termination steps of glycoconjugate biosynthesis creating the sialyl Lewisx or sialyl Lewisa determinant, which play an important role in cell-cell interaction. While cDNA, chromosomal localization and kinetic properties of a number of fucosyltransferases are known, immunocytochemical localization and trafficking studies have been delayed because of the lack of specific antibodies due to the pronounced homology of alpha 1, 3 fucolsyltransferases III, V and VI. Here we report development and characterization of monospecific polyclonal antibodies to alpha 1-3 fucosyltransferase V (FucT-V) and their application for immunodetection in transfected cells. Antisera against FucT-V were raised in two different ways: first by producing a fusion protein beta-galactosidase-FucT-V in Escherichia coli, and by synthesizing a peptide stretch specific for FucT-V. Polyclonal antisera were raised against each of both antigens and characterized by enzyme-linked immunosorbent assay, neutralization of activity, immunoblotting, immunofluorescence and immunoprecipitation of metabolically labeled COS cells, transiently transfected with cDNA encoding FucT-V. Both antibodies recognized only FucT-V. No cross-reactivity to FucT-III or FucT-VI was observed. FucT-V was localized mainly to the Golgi apparatus by colocalization with beta 1, 4-galactosyltransferase, and to the cell surface of COS, CHO and HeLa cells. Expression of FucT-V in COS cells revealed three enzyme forms of 58, 53 and 50 kDa, respectively. These size differences arose by post-translational modifications, as shown by pulse-chase experiments. Our results indicate that alpha 1-3 fucosyltransferase is a Golgi-associated enzyme and suggest its possible occurrence on the cell surface.
Authors:
L Borsig; R Kleene; A Dinter; E G Berger
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  European journal of cell biology     Volume:  70     ISSN:  0171-9335     ISO Abbreviation:  Eur. J. Cell Biol.     Publication Date:  1996 May 
Date Detail:
Created Date:  1996-10-10     Completed Date:  1996-10-10     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  7906240     Medline TA:  Eur J Cell Biol     Country:  GERMANY    
Other Details:
Languages:  eng     Pagination:  42-53     Citation Subset:  IM    
Affiliation:
Institute of Physiology, University of Zürich, Switzerland.
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MeSH Terms
Descriptor/Qualifier:
Animals
Antibody Specificity
CHO Cells
Cell Line
Cricetinae
Enzyme-Linked Immunosorbent Assay
Fucosyltransferases / analysis*,  immunology
Hela Cells
Humans
Immunohistochemistry
Precipitin Tests
Chemical
Reg. No./Substance:
EC 2.4.1.-/Fucosyltransferases; EC 2.4.1.152/galactoside 3-fucosyltransferase

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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