OGT (gene)

UDP-N-acetylglucosamine—peptide N-acetylglucosaminyltransferase, also known as O-linked β-N-acetylglucosamine transferase and O-GlcNAc transferase, OGT is an is_associated_with::enzyme that in humans is encoded by the OGT is_associated_with::gene.

Function
O-linked N-acetylglucosamine (O-GlcNAc) is_associated_with::transferase (OGT) catalyzes the addition of a single N-acetylglucosamine in O-glycosidic linkage to is_associated_with::serine or is_associated_with::threonine residues of is_associated_with::intracellular is_associated_with::proteins. Since both is_associated_with::phosphorylation and O-GlcNAcylation compete for similar serine or threonine residues, the two processes may compete for sites, or they may alter the substrate specificity of nearby sites by steric or electrostatic effects. The protein contains nine or 14 tetratricopeptide repeats, depending on the splice variant, and a putative bipartite nuclear localization signal. Two alternatively spliced transcript variants encoding distinct isoforms (nucleocytoplasmic and mitochondrial) have been found for this gene. OGT glycosylates many proteins including: is_associated_with::Histone H2B, is_associated_with::AKT1, is_associated_with::PFKL, KMT2E/MLL5, is_associated_with::MAPT/TAU, is_associated_with::Host cell factor C1, and is_associated_with::SIN3A.

O-GlcNAc transferase is a part of a host of biological functions within the is_associated_with::human body. OGT is involved in the resistance of is_associated_with::insulin in is_associated_with::muscle cells and is_associated_with::adipocytes by inhibiting the Threonine 308 phosphorylation of AKT1, increasing the rate of is_associated_with::IRS1 phosphorylation (at Serine 307 and Serine 632/635), reducing insulin signaling, and glycosylating components of insulin signals. Additionally, O-GlcNAc transferase catalyzes intracellular is_associated_with::glycosylation of serine and threonine residues with the addition of N-acetylglucosamine. Studies show that OGT alleles are vital for is_associated_with::embryogenesis, and that OGT is necessary for intracellular glycosylation and is_associated_with::embryonic stem cell vitality. O-GlcNAc transferase also catalyzes the is_associated_with::posttranslational modification that modifies transcription factors and is_associated_with::RNA polymerase II, however the specific function of this modification is mostly unknown.

OGT cleaves Host Cell Factor C1, at one of the 6 repeat sequences. The TPR repeat domain of OGT binds to the carboxyl terminal portion of an HCF1 proteolytic repeat so that the cleavage region is in the glycosyltransferase active site above uridine-diphosphate-GlcNAc The large proportion of OGT complexed with HCF1 is necessary for HCF1 cleavage, and HCFC1 is required for OGT stabilization in the nucleus. HCF1 regulates OGT stability using a post-transcriptional mechanism, however the mechanism of the interaction with HCFC1 is still unknown.

Structure
The human OGT gene has 1046 is_associated_with::amino acid residues, and is a heterotrimer consisting of two 110 kDa subunits and one 78 kDa subunit. The 110 kDa subunit contains 13 is_associated_with::tetratricopeptide (TPR) repeats; the 13th repeat is truncated. These subunits are dimerized by TPR repeats 6 and 7. OGT is highly expressed in the is_associated_with::pancreas and also expressed in the is_associated_with::heart, is_associated_with::brain, is_associated_with::skeletal muscle, and the is_associated_with::placenta. There have been trace amounts found in the is_associated_with::lung and the is_associated_with::liver. The binding sites have been determined for the 110 kDa subunit. It has 3 binding sites at amino acid residues 849, 852, and 935. The probable active site is residue 508.

The is_associated_with::crystal structure of O-GlcNAc transferase has not been well studied, but the structure of a binary complex with UDP and a ternary complex with UDP and a is_associated_with::peptide substrate has been researched. The OGT-UDP complex contains three domains in its catalytic region, the amino (N)-terminal domain, the carboxy (C)-terminal domain, and the intervening domain (Int-D). The catalytic region is linked to TPR repeats by a translational helix (H3), which loops from the C-cat domain to the N-Cat domain along the upper surface of the catalytic region. The OGT-UDP-peptide complex has a larger space between the TPR domain and the catalytic region than the OGT-UDP complex. The CKII peptide, which contains three serine residues and a threonine residue, binds in this space. This structure supports an ordered sequential bi-bi mechanism that matches the fact that “at saturating peptide concentrations, a is_associated_with::competitive inhibition pattern was obtained for UDP with respect to UDP-GlcNAc.”

Mechanism of catalysis
The molecular mechanism of O-linked N-acetylglucosamine transferase has not been extensively studied either, since there is not a confirmed crystal structure of the enzyme. A proposed mechanism supported by product inhibition patterns by UDP at saturating peptide conditions proceeds with starting materials Uridine diphosphate N-acetylglucosamine, and a peptide chain with a reactive serine or threonine is_associated_with::hydroxyl group. The proposed reaction is an ordered sequential bi-bi mechanism.



The chemical reaction can be written as:

(1) UDP-N-acetyl-D-glucosamine + [protein]-L-serine → UDP + [protein]-3-O-(N-acetyl-D-glucosaminyl)-L-serine

(2) UDP-N-acetyl-D-glucosamine + [protein]-L-threonine → UDP + [protein]-3-O-(N-acetyl-D-glucosaminyl)-L-threonine

First, the hydroxyl group of serine is deprotonated by Histidine 498, a catalytic base in this proposed reaction. Lysine 842 is also present to stabilize the UDP moiety. The is_associated_with::oxygen ion then attacks the sugar-phosphate bond between the is_associated_with::glucosamine and UDP. This results in the splitting of UDP-N-acetylglucosamine into N-acetylglucosamine – Peptide and UDP. is_associated_with::Proton transfers take place at the is_associated_with::phosphate and Histidine 498. This mechanism is spurred by OGT gene containing O-linked N-acetylglucosamine transferase. Aside from proton transfers the reaction proceeds in one step, as shown in Figure 2. Figure 2 uses a lone serine residue as a representative of the peptide with a reactive hydroxyl group. Threonine could have also been used in the mechanism.

Regulation


O-GlcNAc transferase is part of a dynamic competition for a serine or threonine hydroxyl functional group in a peptide unit. Figure 3 shows an example of both reciprocal same-site occupancy and adjacent-site occupancy. For the same-site occupancy, OGT competes with is_associated_with::kinase to catalyze the glycosylation of the protein instead of phosphorylation. The adjacent-site occupancy example shows the naked protein catalyzed by OGT converted to a is_associated_with::glycoprotein, which can increase the turnover of proteins such as the tumor repressor p53.

The post-translational modification of proteins by O-GlcNAc is spurred by is_associated_with::glucose flux through the is_associated_with::hexosamine biosynthetic pathway. OGT catalyzes attachment of the O-GlcNAc group to serine and threonine, while O-GlcNAcase spurs is_associated_with::sugar removal.

This regulation is important for multiple cellular processes including transcription, is_associated_with::signal transduction, and proteasomal degradation. Also, there is competitive regulation between OGT and kinase for the protein to attach to a phosphate group or O-GlcNAc, which can alter the function of proteins in the body through downstream effects. OGT inhibits the activity of 6-phosophofructosekinase PFKL by mediating the glycosylation process. This then acts as a part of is_associated_with::glycolysis regulation. O-GlcNAc has been defined as a negative transcription regulator in response to steroid hormone signaling.

Studies show that O-GlcNAc transferase interacts directly with the Ten eleven translocation 2 (is_associated_with::TET2) enzyme, which converts is_associated_with::5-methylcytosine to is_associated_with::5-hydroxymethylcytosine and regulates gene transcription. Additionally, increasing levels of OGT for O-GlcNAcylation may have therapeutic effects for Alzheimer's disease patients. Brain glucose metabolism is impaired in Alzheimer's disease, and a study suggests that this leads to hyperphosphorylation of tau and degerenation of tau O-GlcNCAcylation. Replenishing tau O-GlcNacylation in the brain along with protein phosphatase could deter this process and improve brain glucose metabolism.