The mitochondrial pyruvate dehydrogenase complex (PDC) plays a crucial role in glucose oxidation.

PDC is inhibited by exposure to singlet oxygen (1O2).

Peptide mass mapping indicates modification at the lipoyllysines in the PDC subunits.

Oxidation is detected at 42 Met, Trp, His and Tyr residues on all 4 subunits at 20–50 % levels.

Inhibition is modulated by both pre- and post-oxidation treatment with lipoic acid and lipoamide.

The multi-subunit pyruvate dehydrogenase complex (PDC) plays a crucial role in glucose oxidation as it determines whether pyruvate is used for mitochondrial oxidative phosphorylation or is converted to lactate for aerobic glycolysis. PDC contains multiple lipoic acid groups, covalently attached at lysine residues to give lipoyllysine, which are responsible for acyl group transfer and critical to complex activity. We have recently reported that both free lipoic acid, and lipoyllysine in alpha-keto glutarate dehydrogenase, are highly susceptible to singlet oxygen (1O2)-induced oxidation. We therefore hypothesized that PDC activity and structure would be influenced by 1O2 (generated using Rose Bengal and light) via modification of the lipoyllysines and other residues. PDC activity was decreased by photooxidation, with this being dependent on light exposure, O2, the presence of Rose Bengal, and D2O consistent with 1O2-mediated reactions. These changes were modulated by pre-illumination addition of free lipoic acid and lipoamide. Activity loss occurred concurrently with lipoyllysine and sidechain modification (determined by mass spectrometry) and protein aggregation (detected by SDS-PAGE). Peptide mass mapping provided evidence for modification at 42 residues (Met, Trp, His and Tyr; with modification extents of 20–50 %) and each of the lipoyllysine sites (6–20 % modification). Structure modelling indicated the modifications occur across all 4 subunit types, and occur in functional domains or at multimer interfaces, consistent with damage at multiple sites contributing to the overall loss of activity. These data indicate that PDC activity and structure are susceptible to 1O2-induced damage with potential effects on cellular pathways of glucose metabolism.

Pyruvate dehydrogenase complex

singlet oxygen

Photooxidation

Protein oxidation

Crosslinking

Lipoic acid

Lipoyllysine

Glucose metabolism

Tricarboxylic acid cycle

Mitochondria

dihydrolipoyl transacetylase (also known as E2)

dihydrolipoamide dehydrogenase (also known as E3)

dihydrolipoamide dehydrogenase-binding protein (also known as E3BP)

alpha-ketoglutarate dehydrogenase complex

liquid chromatography-mass spectrometry

the lowest energy (first) excited singlet state of molecular oxygen

commonly referred to as singlet oxygen

pyruvate dehydrogenase complex

pyruvate dehydrogenase α chain (also known as E1α)

pyruvate dehydrogenase β chain (also known as E1β)

peripheral subunit binding domain

© 2025 The Authors. Published by Elsevier Inc.