An Alternative Pathway of Oleate β-Oxidation in Escherichia coli Involving the Hydrolysis of a Dead End Intermediate by a Thioesterase

Abstract

The degradation of 2-trans,5-cis-tetradecadienoylCoA, a metabolite of oleic acid, by the purified complex of fatty acid oxidation from Escherichia coli was studied to determine how much of the metabolite is converted to 3,5-cis-tetradecadienoyl-CoA and thereby diverted from the classical, isomerase-dependent pathway of oleate ␤-oxidation. Approximately 10% of the 2,5-intermediate was converted to the 3,5-isomer. When the latter compound was allowed to accumulate, it strongly inhibited the flux through the main pathway. Since ⌬ 3,5 ,⌬ 2,4 -dienoyl-CoA isomerase was not detected in E. coli cells grown on oleate, the 3,5-intermediate cannot be metabolized via the reductase-dependent pathway. However, it was hydrolyzed by a thioesterase, which was most active with 3,5-cis-tetradecadienoyl-CoA as substrate and which was induced by growth of E. coli on oleate. An analysis of fatty acids present in the medium after growth of E. coli on oleate revealed the presence of 3,5-tetradecadienoate, which was not detected after cells were grown on palmitate or glucose. Altogether, these data prompt the conclusion that oleate is mostly degraded via the classical, isomerase-dependent pathway in E. coli but that a small amount of 2-trans,5-cistetradecadienoyl-CoA is diverted from the pathway via conversion to 3,5-cis-tetradecadienoyl-CoA by ⌬ 3 ,⌬ 2 -enoyl-CoA isomerase. The 3,5-intermediate, which would strongly inhibit ␤-oxidation if allowed to accumulate, is hydrolyzed, and the resultant 3,5-tetradecadienoate is excreted into the growth medium. This study provides evidence for the novel function of a thioesterase in ␤-oxidation.

Structure, Function, and Mechanism of the Phenylacetate Pathway Hot Dog-fold Thioesterase PaaI

“…Moreover, it belongs to the acyl-CoA thioesterase subfamily and, in particular, to the same clade of acyl-CoA thioesterases as does the Arthrobacter 4-hydroxybenzoyl-coenzyme A thioesterase (4-HBA-CoA thioesterase) 3 (39,40). Acyl-CoA thioesterases generally function in the cell to release the carboxylate unit for degradation, export (39,(41)(42)(43)(44)(45), or regulation (43, 46 -48) and/or to release the CoA in thioester metabolites for participation in alternate metabolic pathways (26).…”

Dead End Metabolites - Defining the Known Unknowns of the E. coli Metabolic Network

“…coli K-12), revealed a deficiency in our link between two related pathways, oleate β-oxidation and fatty acid β-oxidation. This was remedied by the addition of the following two metabolic reactions [10]:…”

Oleate β-oxidation in yeast involves thioesterase but not Yor180c protein that is not a dienoyl-CoA isomerase

“…This minor route of β-oxidation is referred to as the reductase-dependent pathway (see Scheme 1) even though dienoyl-CoA isomerase is the enzyme that is unique to this process while both of the other auxiliary enzymes, 2,4-dienoyl-CoA reductase and enoyl-CoA isomerase, have essential functions in other pathways of β-oxidation. E. coli has adopted a different strategy that relies on the hydrolysis of 3,5- cis -tetradecadienoyl-CoA (VI) to yield 3,5-tetradecadienoic acid (X), which is released from the cell [6]. This minor pathway of oleate β-oxidation is referred to as the thioesterase-dependent pathway (see Scheme 1).…”