Dataset Information

Assessing the Thiamine Diphosphate Dependent Pyruvate Dehydrogenase E1 Subunit for Carboligation Reactions with Aliphatic Ketoacids.

ABSTRACT: The synthetic properties of the Thiamine diphosphate (ThDP)-dependent pyruvate dehydrogenase E1 subunit from Escherichia coli (EcPDH E1) was assessed for carboligation reactions with aliphatic ketoacids. Due to its role in metabolism, EcPDH E1 was previously characterised with respect to its biochemical properties, but it was never applied for synthetic purposes. Here, we show that EcPDH E1 is a promising biocatalyst for the production of chiral ?-hydroxyketones. WT EcPDH E1 shows a 180-250-fold higher catalytic efficiency towards 2-oxobutyrate or pyruvate, respectively, in comparison to engineered transketolase variants from Geobacillus stearothermophilus (TK_GST). Its broad active site cleft allows for the efficient conversion of both (R)- and (S)-configured ?-hydroxyaldehydes, next to linear and branched aliphatic aldehydes as acceptor substrates under kinetically controlled conditions. The alternate, thermodynamically controlled self-reaction of aliphatic aldehydes was shown to be limited to low levels of conversion, which we propose to be due to their large hydration constants. Additionally, the thermodynamically controlled approach was demonstrated to suffer from a loss of stereoselectivity, which makes it unfeasible for aliphatic substrates.

SUBMITTER: Marsden SR

PROVIDER: S-EPMC7696235 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Assessing the Thiamine Diphosphate Dependent Pyruvate Dehydrogenase E1 Subunit for Carboligation Reactions with Aliphatic Ketoacids.

Marsden Stefan R SR McMillan Duncan G G DGG Hanefeld Ulf U

International journal of molecular sciences 20201116 22

The synthetic properties of the Thiamine diphosphate (ThDP)-dependent pyruvate dehydrogenase E1 subunit from Escherichia coli (EcPDH E1) was assessed for carboligation reactions with aliphatic ketoacids. Due to its role in metabolism, EcPDH E1 was previously characterised with respect to its biochemical properties, but it was never applied for synthetic purposes. Here, we show that EcPDH E1 is a promising biocatalyst for the production of chiral α-hydroxyketones. WT < ...[more]

PMID: 33207817

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Project description:BackgroundDuring and following myocardial ischemia, glucose oxidation rates are low and fatty acids dominate as a source of oxidative metabolism. This metabolic phenotype is associated with contractile dysfunction during reperfusion. To determine the mechanism of this reliance on fatty acid oxidation as a source of ATP generation, a functional proteomics approach was utilized.Results2-D gel electrophoresis of mitochondria from working rat hearts subjected to 25 minutes of global no flow ischemia followed by 40 minutes of aerobic reperfusion identified 32 changes in protein abundance compared to aerobic controls. Of the five proteins with the greatest change in abundance, two were increased (long chain acyl-coenzyme A dehydrogenase (48 +/- 1 versus 39 +/- 3 arbitrary units, n = 3, P < 0.05) and alpha subunit of ATP synthase (189 +/- 15 versus 113 +/- 23 arbitrary units, n = 3, P < 0.05)), while two were decreased (24 kDa subunit of NADH-ubiquinone oxidoreductase (94 +/- 7 versus 127 +/- 9 arbitrary units, n = 3, P < 0.05) and D subunit of ATP synthase (230 +/- 11 versus 368 +/- 47 arbitrary units, n = 3, P < 05)). Two forms of pyruvate dehydrogenase betaE1 subunit, the rate-limiting enzyme for glucose oxidation, were also identified. The protein level of the more acidic form of pyruvate dehydrogenase was reduced during reperfusion (37 +/- 4 versus 56 +/- 7 arbitrary units, n = 3, P < 05), while the more basic form remained unchanged. The more acidic isoform was found to be O-palmitoylated, while both isoforms exhibited ischemia/reperfusion-induced phosphorylation. In silico analysis identified the putative kinases as the insulin receptor kinase for the more basic form and protein kinase Czeta or protein kinase A for the more acidic form. These modifications of pyruvate dehydrogenase are associated with a 35% decrease in glucose oxidation during reperfusion.ConclusionsCardiac ischemia/reperfusion induces significant changes to a number of metabolic proteins of the mitochondrial proteome. In particular, ischemia/reperfusion induced the post-translational modification of pyruvate dehydrogenase, the rate-limiting step of glucose oxidation, which is associated with a 35% decrease in glucose oxidation during reperfusion. Therefore these post-translational modifications may have important implications in the regulation of myocardial energy metabolism.

Dataset Information

Assessing the Thiamine Diphosphate Dependent Pyruvate Dehydrogenase E1 Subunit for Carboligation Reactions with Aliphatic Ketoacids.

Publications

Assessing the Thiamine Diphosphate Dependent Pyruvate Dehydrogenase E1 Subunit for Carboligation Reactions with Aliphatic Ketoacids.

Similar Datasets

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