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Substrate specificity provides insights into the sugar donor recognition mechanism of O-GlcNAc transferase (OGT). 2013

Xiaofeng Ma, and Pi Liu, and Hui Yan, and Hong Sun, and Xiaoyan Liu, and Feng Zhou, and Lei Li, and Yi Chen, and Musleh M Muthana, and Xi Chen, and Peng G Wang, and Lianwen Zhang
College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin, China.

O-Linked β-N-acetylglucosaminyl transferase (OGT) plays an important role in the glycosylation of proteins, which is involved in various cellular events. In human, three isoforms of OGT (short OGT [sOGT]; mitochondrial OGT [mOGT]; and nucleocytoplasmic OGT [ncOGT]) share the same catalytic domain, implying that they might adopt a similar catalytic mechanism, including sugar donor recognition. In this work, the sugar-nucleotide tolerance of sOGT was investigated. Among a series of uridine 5'-diphosphate-N-acetylglucosamine (UDP-GlcNAc) analogs tested using the casein kinase II (CKII) peptide as the sugar acceptor, four compounds could be used by sOGT, including UDP-6-deoxy-GlcNAc, UDP-GlcNPr, UDP-6-deoxy-GalNAc and UDP-4-deoxy-GlcNAc. Determined values of Km showed that the substitution of the N-acyl group, deoxy modification of C6/C4-OH or epimerization of C4-OH of the GlcNAc in UDP-GlcNAc decreased its affinity to sOGT. A molecular docking study combined with site-directed mutagenesis indicated that the backbone carbonyl oxygen of Leu653 and the hydroxyl group of Thr560 in sOGT contributed to the recognition of the sugar moiety via hydrogen bonds. The close vicinity between Met501 and the N-acyl group of GlcNPr, as well as the hydrophobic environment near Met501, were responsible for the selective binding of UDP-GlcNPr. These findings illustrate the interaction of OGT and sugar nucleotide donor, providing insights into the OGT catalytic mechanism.

UI MeSH Term Description Entries
D007700 Kinetics The rate dynamics in chemical or physical systems.
D011485 Protein Binding The process in which substances, either endogenous or exogenous, bind to proteins, peptides, enzymes, protein precursors, or allied compounds. Specific protein-binding measures are often used as assays in diagnostic assessments. Plasma Protein Binding Capacity,Binding, Protein
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D006860 Hydrogen Bonding A low-energy attractive force between hydrogen and another element. It plays a major role in determining the properties of water, proteins, and other compounds. Hydrogen Bonds,Bond, Hydrogen,Hydrogen Bond
D000117 Acetylglucosamine The N-acetyl derivative of glucosamine. Acetyl Glucosamine,N-Acetyl Glucosamine,N-Acetyl-beta-D-Glucosamine,N-Acetylglucosamine,beta-N-Acetylglucosamine,2-Acetamido-2-Deoxy-D-Glucose,2-Acetamido-2-Deoxyglucose,N-Acetyl-D-Glucosamine,2 Acetamido 2 Deoxy D Glucose,2 Acetamido 2 Deoxyglucose,Glucosamine, Acetyl,Glucosamine, N-Acetyl,N Acetyl D Glucosamine,N Acetyl Glucosamine,N Acetyl beta D Glucosamine,N Acetylglucosamine,beta N Acetylglucosamine
D013379 Substrate Specificity A characteristic feature of enzyme activity in relation to the kind of substrate on which the enzyme or catalytic molecule reacts. Specificities, Substrate,Specificity, Substrate,Substrate Specificities
D016297 Mutagenesis, Site-Directed Genetically engineered MUTAGENESIS at a specific site in the DNA molecule that introduces a base substitution, or an insertion or deletion. Mutagenesis, Oligonucleotide-Directed,Mutagenesis, Site-Specific,Oligonucleotide-Directed Mutagenesis,Site-Directed Mutagenesis,Site-Specific Mutagenesis,Mutageneses, Oligonucleotide-Directed,Mutageneses, Site-Directed,Mutageneses, Site-Specific,Mutagenesis, Oligonucleotide Directed,Mutagenesis, Site Directed,Mutagenesis, Site Specific,Oligonucleotide Directed Mutagenesis,Oligonucleotide-Directed Mutageneses,Site Directed Mutagenesis,Site Specific Mutagenesis,Site-Directed Mutageneses,Site-Specific Mutageneses
D017351 N-Acetylglucosaminyltransferases Enzymes that catalyze the transfer of N-acetylglucosamine from a nucleoside diphosphate N-acetylglucosamine to an acceptor molecule which is frequently another carbohydrate. EC 2.4.1.-. N-Acetylglucosamine Transferases,N Acetylglucosamine Transferases,N Acetylglucosaminyltransferases,Transferases, N-Acetylglucosamine
D062105 Molecular Docking Simulation A computer simulation technique that is used to model the interaction between two molecules. Typically the docking simulation measures the interactions of a small molecule or ligand with a part of a larger molecule such as a protein. Molecular Docking,Molecular Docking Simulations,Molecular Docking Analysis,Analysis, Molecular Docking,Docking Analysis, Molecular,Docking Simulation, Molecular,Docking, Molecular,Molecular Docking Analyses,Molecular Dockings,Simulation, Molecular Docking
D019943 Amino Acid Substitution The naturally occurring or experimentally induced replacement of one or more AMINO ACIDS in a protein with another. If a functionally equivalent amino acid is substituted, the protein may retain wild-type activity. Substitution may also diminish, enhance, or eliminate protein function. Experimentally induced substitution is often used to study enzyme activities and binding site properties. Amino Acid Substitutions,Substitution, Amino Acid,Substitutions, Amino Acid

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