P-Hydroxyacyl CoA-dehydrogenase

A second oxidation reaction is catalyzed by P-hydroxyacyl-CoA dehydrogenase, an NAD -dependent enzyme. The product is a p-ketoacyl-GoA. 

Vallejo-Cordoba, B., Mazorra-Manzano, M. A., and Gonzalez-Cordova, A. R, Determination of P-hydroxyacyl CoA-dehydrogenase activity in meat by electrophoretically mediated microanalysis, J. Cap. Elec. Micro. Tech., 8, 81, 2003. 

The enzyme P-hydroxyacyl-CoA-dehydrogenase (HADH, EC 1.1.1.35) is also suitable for the detection of frozen meat or fish. In the oxidation of fatty acids, HADH catalyzes the reaction shown in Formula 12.29. This enzyme is bound to the iimer membrane of mitochondria and is liberated in the freeze/thaw process. Its activity can then be measured in the issuing sap with acetoacetyl CoA or with the artificial substrate N-acetylacetoacetylcysteamine. 

Feeney, R.E., Yin Yeh Antifreeze proteins from fish bloods. Adv. Protein Chem. 32, 191 (1978) Fernandez, M., Mano, S., Garcia de Fernando, G.D., Ordonez, J.A., Hoz, L. Use of P-hydroxyacyl-CoA-dehydrogenase (HADH) activity to differentiate frozen from unfrozen fish and shellfish. Eur. Food Res. Technol. 209, 205 (1999)... 

In the third step, 1, -/3-hydroxyacyl-CoA is dehydrogenated to form /3-ketoacyl-CoA, by the action of /3-hydroxyacyl-CoA dehydrogenase NAD+ is the electron acceptor. This enzyme is absolutely specific for the l stereoisomer of hydroxyacyl-CoA The NADH formed in the reaction donates its electrons to NADH dehydrogenase, an electron carrier of the respiratory chain, and ATP is formed from ADP as the electrons pass to 02. The reaction catalyzed by /3-hydroxyacyl-CoA dehydrogenase is closely analogous to the malate dehydrogenase reaction of the citric acid cycle (p. XXX). 

Enzyme L-3-hydroxyacyI CoA dehydrogenase (which is specific for the L-isomer of the P-hydroxyacyl CoA)... 

Strauss AW, Beimett MJ, Rinaldo P, Sims HF, O Brien LK, Zhao Y, et al. Inherited long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency and a fetal-maternal interaction cause maternal fiver disease and other pregnancy complications. Sem Perinatol 1999 23 100-12. 

Yeasts are able to degrade long-chain alkanes. The initial hydroxylation is carried out in micrososmes by cytochrome P-450, while degradation of the alkanoate is carried out in peroxisomes that contain the P-oxidation enzymes alkanoate oxidase, enoyl-CoA hydratase, and 3-hydroxyacyl-CoA dehydrogenase. Further details are given in Chapter 4, Sections 4.4.1.2 and 4.4.4. 

Reaction 4. In this oxidation reaction the hydroxyl group of the p-carbon is now dehydrogenated. NAD+ is reduced to form NADH that is subsequently used to produce three ATP molecules by oxidative phosphorylation. L- -Hydroxyacyl-CoA dehydrogenase catalyzes this reaction. 

Hydroxyacyl-CoA dehydrogenase EC 1.1.1.35/36 Reversible NAD(P) -dependent oxidation of alcohol to ketone... 

Fig. 8. P-Oxidation of fatty acids in E. coli. Long-chain fatty acids are transported into the cell by FadL and converted to their CoA thioesters by FadD (not shown). The acyl-CoAs are substrates for the (1) acyl-CoA dehydrogenase (YafH) to form a trans-2-enoyl-CoA. The double bond is reduced by (2) rrans-2-enoyl-hydratase (crotonase) activity of FadB. The P-hydroxyacyl-CoA is then a substrate for the NADP -dependent dehydrogenase activity of FadB (3). A thiolase, FadA (4), releases acetyl-CoA from the P-ketoacyl-CoA to form an acyl-CoA for subsequent cycles. (5) Polyunsaturated fatty acyl-CoAs are reduced by the 2,4-dienoyl-CoA reductase (FadH). (6) FadB also catalyzes the isomerization of c/s-unsaturated fatty acids to trans. (7) The epimerase activity of FadB converts O-P-hydroxy thioesters to their L-enantiomers via the /rans-2-enoyl-CoA.

Fig. 2. Model of the functional and physical organization of P-oxidation enzymes in mitochondria. (A) P-Oxidation system active with long-chain (LC) acyl-CoAs (B) P-oxidation system active with medium-chain (MC) and short-chain (SC) acyl-CoAs. Abbreviations T, camitineiacylcamitine translocase CPT 11, carnitine palmitoyltransferase 11 AD, acyl-CoA dehydrogenase EH, enoyl-CoA hydratase HD, t-3-hydroxyacyl-CoA dehydrogenase KT, 3-ketoacyl-CoA thiolase VLC, very-long-chain.

The third reaction in the P-oxidation cycle is the reversible dehydrogenation of L-3-hydroxyacyl-CoA to 3-ketoacyl-CoA catalyzed by L-3-hydroxyacyl-CoA dehydrogenase as shown in the following equation. 

CoA oxidase, 3-hydroxyacyl-CoA dehydrogenase, enoyl-CoA hydratase, and 3-ketoacyl-CoA thiolase. Dietary sesamin also increased the activity of 2,4-dienoyl-CoA reductase and A, A -enoyl-CoA isomerase, enzymes involved in the auxiliary pathway for p-oxidation of unsaturated fatty acids [246], On the other hand, the results obtained by Fukuda et al. [247] suggest that increased fatty acid oxidation by dietary sesamin leads to decreased esterification of fatty acids and reduces the synthesis and secretion of triacylglycerol. 

Figure 4. Alignment of the amino acid sequence between His and Glu of the coli multifunctional protein (MP) with those of homologous regions of pig mitochondrial long-chain-specific bifunctional enzyme (LT), plant glyoxysomal tetrafunctional protein (PT), rat peroxisomal trifunctional enzyme (TE), and pig liver L-3-hydroxyacyl-CoA dehydrogenase (LD). The conserved histidine and glutamate residues are indicated by asterisks. The large subunit of human mitochondrial trifunctional P-oxidation complex has the same amino acid sequence as that of LT in this region. Qlu of MP corresponds to Glu of the large subunits of these mammalian p-oxidation complexes and to Glu of their precursors.

The majority of the CoA-moiety of L-3-hydroxyacyl-CoA is in contact with the bulk medium while the fatty acid tail is inserted into the cleft and buried by the enzyme. This orientation of the substrate is consistent with the fact that the length of the acyl chain has a significant effect on the turnover number of pig heart L-3-hydroxyacyl-CoA dehydrogenase. Since detailed structural information about the liganded active site is not available, it is not known why the dehydrogenase displays the top catalytic efficiency with substrates of medium acyl chain length. A more intensive study of the substrate chain length specificity, a characteristic which contributes enormously to the complexity of fatty acid P-oxidation systems, shall be made in the future. 

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