Strategies on Designing Thiamine Analogues for Inhibition of Thiamine Diphosphate (ThDP)-dependent Enzymes: Systematic Investigation through Scaffold Switching and C2-Functionalisation

20 February 2023, Version 2
This content is a preprint and has not undergone peer review at the time of posting.


Thiamine diphosphate (ThDP), the bioactive form of vitamin B1, is an essential coenzyme needed for processes of cellular metabolism in all organisms. ThDP-dependent enzymes all require ThDP as a coenzyme for catalytic activity, although individual enzymes vary significantly in substrate preferences and biochemical reactions. A popular way to study the role of these enzymes through chemical inhibition is to use thiamine/ThDP analogues, which typically feature a neutral aromatic ring in place of the positive thiazolium ring of ThDP. While ThDP analogues have aided work in understanding the structural and mechanistic aspects of the enzyme family, at least two key questions regarding the ligand design strategy remain unresolved: 1) among the reported aromatic rings, which is the best? and 2) how can we achieve selectivity towards a given ThDP-dependent enzyme? In this work, we synthesise derivatives of these analogues covering all central aromatic rings used in the past decade and make a head-to-head comparison of all the compounds as inhibitors of several ThDP-dependent enzymes. Thus, we establish the relationship between the nature of the central ring and the inhibitory profile of these ThDP-competitive enzyme inhibitors. We also demonstrate that introducing a C2-substituent onto the central ring to explore the unique substrate-binding pocket can improve selectivity.


enzyme inhibition
thiamine diphosphate

Supplementary materials

Methods, Results and NMR spectra
Synthetic Schemes S1–8 Enzyme Inhibition Assays – Methods and Results (Figures S1–4, S7–11 and S13) Determination of Exit Vector Geometry – Methods and Results (Figure S5) Computational Docking – Methods and Results (Figures S6 and S12) Crystallography Data General Synthesis Methods Synthetic Procedures and NMR spectra Supplementary References


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