Human O-GlcNAc transferase

O-linked β-N-acetylglucosamine transferase (O-GlcNAc transferase) is an essential mammalian enzyme that acts as a nutrient sensor, coupling metabolic status to the regulation of a wide variety of cellular signaling pathways.^[1] OGT catalyses the transfer of N-acetylglucosamine from UDP-N-acetylglucosamine (UDP-GlcNAc) to serines and threonines of cytoplasmic, nuclear and mitochondrial proteins, including numerous transcription factors, tumour suppressors, kinases, phospahateses and histone-modifying proteins.^[2] Two crystal structures of human OGT are reported here as a ternary complex with UDP and a binary complex with UDP and a peptide substrate.

O-GlcNAc transferase Function

The major mechanism for nutrient sensing in eukaryotes involves OGT. OGT senses cellular glucose levels via UDP-GlcNAc concentration, and responds by O-GlcNAcylating a broad range of nuclear anf cytoplasmic proteins.^[3] Insulin-like signaling pathways and transcriptional activators that regulate glucose levels by controlling gluconeogenisis include proteins that are O-GlcNAcylated by OGT.^[4] Numerous O-GlcNAcylation sites are also phosphorylation sites. OGT is suggested to play a major role in modulating cellular kinase signaling cascades.^[5] Widespread transcriptional regulations also involve OGT.^[6]

OGT Modifications

O-GlcNAc modification has been described for a large and still increasing number of proteins, many of which are key modulators of cellular signalling. O-GlcNAc modifications are catalysed by a OGT, and are removed by the antagonistic enzyme β-N-acetylglucosaminidase (O-GlcNAcase). The general scheme of O-linked N-acetylglucosamine modification suggests that N-acetylglucosamine is added to serine/threonine (Ser/Thr) residues of target proteins by the enzyme OGT, using UDP-GlcNAc as substrate. The N-acetylglucosamine group is removed by the antagonistic activity of O-GlcNAcase.^[7]

OGT Features of Interest

The OGT protein possess two ligands SO4 and uridine-5-diphosphate (UDP). OGT is the only known member of the GT-B superfamily of glycosyltransferases to glycosylate polypeptides, and it contains a long uncharacterized intervening sequence (~120 amino acids) in the middle of the catalytic region. Studies suggest that OGT contains a phosphatidylinositol (3,4,5)-trisphosphate (PIP₃)binding domain. The most unusual feature of OGT is the intervening domain between the catalytic lobes, which is only found in metazoans. This polypeptide adopts a topologically novel fold with a seven-stranded beta sheet core stabilized by flanking alpha helices. There are two long unstructured loops for which electron density is missing.^[8]

OGT Structure

OGT is comprised of two distinct regions: a multidomain catalytic region, which has no available structure and an N-terminal region consisting of a series of tetratricopeptide repeat(TPR) units.^[9] The N terminus of OGT is unusual, consisting of 2.5-13.5 tetratricopeptide repeats (TPRs) depending on alternative splicing.^[10] The N-terminal domain of tetratricopeptide (TPR) mediates the recognition of a broad range of target proteins. Components of the nuclear pore complex are major OGT targets, as OGT depletion by RNA interference (RNAi) results in the loss of GlcNAc modification at the nuclear envelope.

N Terminus

Superhelical TPR domain of OGT, structural similarities to importin alpha (PDB entry 1w3b).

The crystal structure of the homodimeric TPR domain of human OGT, which contains 11.5 TPR repeats gives insight into the mechanism of target recognition. The repeats form an elongated superhilix. The concave surface of the superhelix is lined by absolutely conserved asparagines, in a manner reminiscent of the peptide-binding site of importin alpha. Based on this structural similarity, it is proposed that OGT uses an analogous molecular mechanism to recognize its targets.^[11]

OGT Mediated Disease

Faulty regulation of O-GlcNAc modifications has been suggested to be involved in neurodegenerative diseases, diabetes mellitus and cancer. Biochemical details of these processes are still unclear.^[12] Proteins modified by O-GlcNAc have been directly shown to have a role in the pathology of human diseases. For instance, the Ser/The kinase AKT,PI₍₃₎K,insulin receptor substrate 1, glycogen synthase and endothelial nitric oxide synthase,all of which are enzymes that have a crucial role in insulin signalling,are reversibly modified by OGT. A recent study showed that recruitment of OGT to the plasma membrane specifically prevents the phosphorylation of AKT and possibly other proteins, thereby terminating insulin signalling.^[13] This adds evidence to the view that increasing the level of O-GlcNAc modifications correlates with the development of insulin resistance, which is a characteristic of type II diabetes.^[14][15] Some indications suggest that O-GlcNAc modifications have a role in Alzheimer disease. Higher levels of O-GlcNAc can be detected in the brain tissue, and several proteins involved in neuronal signaling are modified with O-GlcNAc. Among them are the β-amyloid precursor protein, clathrin-assembly proteins and neurofilaments. In the brains of patients with Alzheimer disease, hyperphosphorylated Tau protein was modified by O-GlcNAc to a lesser extent than in healthy individuals.^[16] Studies have shown that some oncogenes and tumour suppressors are targets of O-glycosylation, including the SV40 T antigen and c-MYC.^[17] Tumour cells have an altered glucose metabolism that is expected to produce changes in O-GlcNAc levels and to affect different signaling pathways.