Fig 1.
Flavin oxidation can be an intracellular source of both O2- and H2O2.
Molecular oxygen can abstract a single electron from dihydroflavin, forming a caged pair of O2- and flavosemiquinone. If O2- dissociates (left pathway), a second oxygen molecule will accept the remaining electron, yielding a second molecule of O2-. Cellular superoxide dismutase converts these molecules to H2O2. Alternatively, after a spin inversion (right pathway), O2- can combine with flavosemiquinone to form a peroxy adduct, which upon hydrolysis releases H2O2.
Table 1.
Bacterial strains, and oligonucleotide primers.
Fig 2.
Overview of the pathway of long-chain fatty acid degradation.
FadE (acyl-CoA dehydrogenase) is marked with an asterisk. In this figure it transfers two electrons to the quinone pool via Electron Transfer Flavoprotein (ETF). However, it is possible that ETF is absent from E. coli and FadE is a membrane protein that reduces quinones directly.
Fig 3.
FadE-dependent H2O2 production by whole cells.
Mutants lacking the H2O2-scavenging enzymes catalase and peroxidase (HPX-) were suspended in medium containing dodecanoic acid, and H2O2 release was monitored. The fadR mutant fully induces the fatty-acid degradation pathway, including FadE.
Fig 4.
Non-scavenging mutants grow poorly when catabolizing dodecanoic acid.
(A). At time zero cells were shifted to medium containing glycerol and dodecanoic acid. (B) Even after adaptation to the medium, the fadR mutant exhibited slower growth.
Fig 5.
Beta-oxidation does not generate enough H2O2 to induce the OxyR regulon.
Scavenging-proficient cells were cultured with glycerol and/or dodecanoic acid as a carbon source, and the expression of an OxyR-controlled katG-lacZ fusion was monitored. Expression in a catalase/peroxidase-deficient mutant (HPX-) is shown for comparison.
Fig 6.
Dihydrolipoic acid (DHLA) does not degrade H2O2.
Scavenging by dithiothreitol (DTT) is shown for comparison (B).
Fig 7.
FadE structure and possible route of H2O2 production.
(A) Structure predicted by AlphaFold. (B) Predicted oxidation mechanism. Proton abstraction by a glutamate residue triggers transfer of a hydride anion to the adjacent FAD. The subsequent oxidation of FADH2 would proceed by either of the two routes depicted in Fig 1.