title: Regulation of a cytosolic and nuclear O-GlcNAc transferase. Role of the tetratricopeptide repeats.
abstract: The O-GlcNAc transferase (OGT) is a unique nuclear and cytosolic glycosyltransferase that contains multiple tetratricopeptide repeats. We have begun to characterize the mechanisms regulating OGT using a combination of deletion analysis and kinetic studies. Here we show that the p110 subunit of the enzyme forms both homo- and heterotrimers that appear to have different binding affinities for UDP-GlcNAc. The multimerization domain of OGT lies within the tetratricopeptide repeat domain and is not necessary for activity. Kinetic analyses of the full-length trimer and the truncated monomer forms of OGT suggest that both forms function through a random bi-bi kinetic mechanism. Both the monomer and trimer have similar specific activities and similar K(m) values for peptide substrates. However, they differ in their binding affinities for UDP-GlcNAc, indicating that subunit interactions affect enzyme activity. The findings that recombinant OGT has three distinct K(m) values for UDP-GlcNAc and that UDP-GlcNAc concentrations modulates the affinity of OGT for peptides suggest that OGT is exquisitely regulated by the levels of UDP-GlcNAc within the nucleus and cytoplasm.
authors.affiliation: Department of Biological Chemistry, Johns Hopkins University, Baltimore, Maryland 21205, USA.
authors.forename: L K
authors.initials: LK
authors.lastname: Kreppel
authors.forename: G W
authors.initials: GW
authors.lastname: Hart
mesh_terms: D000818:Animals; D020134:Catalytic Domain; D002467:Cell Nucleus / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D007700:Kinetics; D017351:N-Acetylglucosaminyltransferases / Q000378:metabolism*; D011487:Protein Conformation; D051381:Rats; D018411:Spodoptera; D013329:Structure-Activity Relationship; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000378:metabolism
chemical_list: D014537:Uridine Diphosphate N-Acetylglucosamine; D017351:N-Acetylglucosaminyltransferases; C063057:UDP-N-acetylglucosamine-peptide beta-N-acetylglucosaminyltransferase