Chemical tools for studying O-GlcNAc

Thesis type
(Thesis) Ph.D.
Date created
O-GlcNAcylation is a post-translational modification that governs certain cellular pathways essential for eukaryotic cell functioning including transcription, translation, and protein transport. Its activity has been discovered to relate closely with neurodegenerative disorders including Parkinson and Alzheimer disease. O-GlcNAcylation is facilitated by two enzymes: UDP-N-acetyl-D-glucosamine:polypeptidyl-transferase (OGT) transfers O-GlcNAc onto serine and threonine residues; O-GlcNAcase (OGA) cleaves O-GlcNAc off to liberate a free protein substrate. In this dissertation, we focused on developing new chemical tools to better understand the effect of O-GlcNAcylation in a biological context. To monitor the activity of OGT in cell, we demonstrated the use of a fluorophore integrated O-GlcNAc intermediate which can be tolerated by UDP-N-acetylglucosamine pyrophosphorylase and turned over by OGT to monitor O-GlcNAcylation in cells without the need of an engineered cell line. Upon further development, we envision this probe can potentially open the door to directly screen large numbers of OGT ligands in a cellular level. On the other hand, the use of OGA inhibitor has proven to decrease tau accumulation in tau-overexpression mice; aggregation of hyperphosphorylated tau is a pathological hallmark of Alzheimer disease. Current inhibitors have been hindered by the challenge of high total polar surface area hence lowering their permeability across blood brain barrier. We further investigated the effect of the side chain with the use of click chemistry approach to increase its versatility which allow quick tethering towards improving the pharmacokinetic of the inhibitors. To better access these iminocyclitol inhibitors, we also further developed new chemistries along with the incorporation of α-chlorohydrin aldol-ring annulation approach to generate the polyhydroxy-pyrrolidine core structure. We also demonstrated effort towards synthesis of a non-saccharide OGA inhibitor which incorporate structural key elements of 1,2,-dideoxy-2'-ethylamino-α-D-glucopyranoso-[2,1-d-Δ2'-thiazoline] (Thiamet-G), 6-acetamido-6-deoxy-castanospermine (6-Ac-Cas) and O-(2-acetamido-2-deoxy-D-glucopyranosylidene)-amino-N-phenylcarbamate (PUGNAc). These inhibitors should prove useful, following further refinement, for the wider community.
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Supervisor or Senior Supervisor
Thesis advisor: Vocadlo, David
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