A disulfide-bonded dimer of the Golgi beta-galactoside alpha2,6-sialyltransferase is catalytically inactive yet still retains the ability to bind galactose.
The alpha2,6-sialyltransferase is a terminal glycosyltransferase localized in the trans Golgi and trans Golgi network. Here we show that 30% of the total rat liver Golgi alpha2,6-sialyltransferase forms a disulfide-bonded 100-kDa species that can be converted to the 50-kDa monomer form of the enzyme upon reduction. Limited proteolysis of both enzyme forms demonstrates that the 100-kDa species is a disulfide-bonded homodimer of the alpha2,6-sialyltransferase. The alpha2,6-sialyltransferase disulfide-bonded dimer is found in bovine liver Golgi membranes and in Golgi membranes prepared and solubilized in the presence of 100 mM iodoacetamide, suggesting that it is not unique to rat liver or formed aberrantly upon membrane lysis. The dimer form of the enzyme possesses no significant catalytic activity and has a much lower affinity for CDP-hexanolamine-agarose compared with the monomer form. In contrast, both the alpha2,6-sialyltransferase monomer and the disulfide-bonded dimer bind strongly to galactose and galactose-terminated substrates. These results suggest that the alpha2,6-sialyltransferase disulfide-bonded dimer lacks catalytic activity due to a weak affinity for its sugar nucleotide donor, CMP-NeuAc, and that this catalytically inactive form of the enzyme may act as a galactose-specific lectin in the Golgi.
Ma
J
J
Department of Biochemistry, University of Illinois College of Medicine, Chicago, 60612, USA.
Colley
K J
KJ
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002499:Centrifugation, Density Gradient; D002846:Chromatography, Affinity; D004220:Disulfides; D004591:Electrophoresis, Polyacrylamide Gel; D018870:Endoplasmic Reticulum, Rough / Q000201:enzymology; D018871:Endoplasmic Reticulum, Smooth / Q000201:enzymology; D005690:Galactose / Q000378:metabolism*; D006056:Golgi Apparatus / Q000201:enzymology*; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D008970:Molecular Weight; D010449:Peptide Mapping; D051381:Rats; D012697:Serine Endopeptidases; D012799:Sialyltransferases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
D004220:Disulfides; D046911:Macromolecular Substances; D012799:Sialyltransferases; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D005690:Galactose; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase