Abstract
Pyruvate kinase catalyses the last step in glycolysis and may contribute to regulation of aerobic glycolysis in cancer cells, where there is a more pro-oxidant state. This state also favours lipid peroxidation and the formation of electrophilic fragmentation products, including short-chain aldehydes e.g. 4-hydroxy-2-hexenal (HHE), malondialdehyde and acrolein. As little is known about the interaction of these molecules with pyruvate kinase, we investigated their reactions with and effects on the activity of the enzyme. Analysis by LC-MSMS showed unique modification profiles for each aldehyde, with only Lys 393 and 475 modified by all 3 compounds. Acrolein was the most potent inhibitor, and at incubation times of 2 hours or more, pathophysiological concentrations (2-10 μM) induced significant effects. Treatment of MCF-7 cells caused similar losses of activity to treatment in vitro. Cys326, Cys358, and Cys474 were found to be modified in acrolein-treated pyruvate kinase, and as they are conserved residues located at or near the allosteric or active sites of the pyruvate kinase isoforms, they may contribute to loss of activity. These results suggest that reactive short-chain aldehydes generated during inflammatory diseases or environmental exposure could be involved in metabolic changes observed in cancer cells through alteration of pyruvate kinase activity.