Augmenting the activity of Glyoxalase 1 to slow biological ageing

The objective of this project is to gain a deeper understanding of the regulation of glyoxalase (GLO1), an enzyme that plays a crucial role in safeguarding against dicarbonyl-induced damage. By doing so, we aim to develop innovative techniques that can enhance dicarbonyl elimination and impede biological aging. All living organisms rely on sugar metabolism to produce energy. The downside of this dependence is the generation of reactive glycolytic intermediates, particularly α,β-dicarbonyls such as methylglyoxal (MG), which are produced at a rate of around 120 μmol/kg daily in healthy adults. The production of dicarbonyls increases with heightened glycolytic flux, impairment of metabolic shunts that divert glycolytic intermediates, or anaerobic glycolysis (e.g. in hypoxia). Dicarbonyls readily penetrate cell membranes and react with and modify vulnerable targets on proteins, phospholipids, and nucleic acids, ultimately leading to the creation of Advanced Glycation End-products (AGEs) such as MG-H1 [Nδ-(5-hydro-5-methyl-4-imidazolon-2-yl) ornithine]. The accumulation of AGEs has been linked to almost all age-related diseases, including Alzheimer's disease, diabetes, heart disease, mental illness, renal failure, and cancer. Reactive dicarbonyls are the primary source of AGEs in humans, and MG modifies approximately 0.1–1% of lysine and arginine residues on proteins, 1% of nucleotides on DNA, and 0.1% of phospholipids. Smaller molecules such as nitric oxide and glutathione (GSH) are also affected by dicarbonyls, causing dysfunction in systems regulated by them. We have taken a structure/function approach to the study of glyoxalase 1 (GLO1), the main enzyme involved in the breakdown of methylglyoxal. GLO1 is highly conserved in eukaryotes, and we have performed a screen of the protein by modifying key amino acids that alter the activity of GLO1. We will use CRISPR to generate cell-lines expressing variants of GLO1 with augmented/no activity. Using two different methodologies, we will restore/measure GLO1 activity and track how long our interventions work in cell culture models. Informed by this work, we will move towards the goal of generating transgenic animals with increased longevity. This work will increase our understanding of the fundamental role of GLO1 in ageing and is a critical step in the development of novel therapeutics.