Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.3.99.3 (acyl-CoA dehydrogenase)
 1,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The role of the oxyanion hole in the reaction catalyzed by pig medium-chain acyl-CoA dehydrogenase (pMCAD) has been investigated using enzyme reconstituted with 2'-deoxy-FAD. The k(cat) (18.8 +/- 0.5 s(-1)) and K(m) (2.5 +/- 0.4 microM) values for the oxidation of n-octanoyl-CoA (C(8)-CoA) by WT pMCAD recombinantly expressed in Escherichia coli are similar to those of native pMCAD isolated from pig kidney. In agreement with previous studies [Engst et al. (1999) Biochemistry 38, 257-267], reconstitution of the WT enzyme with 2'-deoxy-FAD causes a large (400-fold) decrease in k(cat) but has little effect on K(m). To investigate the molecular basis for the alterations in activity resulting from changes in hydrogen bonding between the substrate and the enzyme's oxyanion hole, the structure of the product analogue hexadienoyl-CoA (HD-CoA) bound to the 2'-deoxy-FAD-reconstituted enzyme has been probed by Raman spectroscopy. Importantly, while WT pMCAD causes a 27 cm(-1) decrease in the vibrational frequency of the HD enone band, from 1595 to 1568 cm(-1), the enone band is only shifted 10 cm(-1) upon binding HD-CoA to 2'-deoxy-FAD pMCAD. Thus, removal of the 2'-ribityl hydroxyl group results in a substantial reduction in the ability of the enzyme to polarize the ground state of the ES complex. On the basis of an analysis of a similar system, it is estimated that ground state destabilization is reduced by up to 17 kJ mol(-1), while the activation energy for the reaction is raised 15 kJ mol(-1). In addition, removal of the 2'-ribityl hydroxyl reduces the redox potential shift that is induced by HD-CoA binding from 18 to 11 kJ mol(-1). Consequently, while ligand polarization caused by hydrogen bonding in the oxyanion hole is intimately linked to substrate turnover, additional factors must be responsible for ligand-induced changes in redox potential. Finally, while replacement of the catalytic base E376 with Gln abolishes the ability of the enzyme to catalyze substrate oxidation and to catalyze the exchange of the C(8)-CoA alpha-protons with solvent deuterium, the 2'-deoxy-FAD-reconstituted enzyme catalyzes alpha-proton exchange at a rate (k(exc)) of 0.085 s(-1), which is only 4-fold slower than k(exc) for WT pMCAD (0.35 s(-1)). Thus, either the oxyanion hole plays only a minor role in stabilizing the transition state for alpha-proton exchange, in contrast to its role in substrate oxidation, or the value of k(exc) for WT pMCAD reflects a process such as exchange of the E376 COOH proton with solvent.
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PMID:Probing hydrogen-bonding interactions in the active site of medium-chain acyl-CoA dehydrogenase using Raman spectroscopy. 1452 97

Three experiments were conducted to develop methods to control the amount of water loss and to evaluate the metabolic effects of water condition in the White Leghorn breeder eggs during incubation. One hundred twenty, 54, and 90 Julia strain White Leghorn breeder eggs were incubated at 37.8 degrees C, 60% RH in experiments 1, 2, and 3. In experiment 1, eggs were drilled with various bore diameters of 0, 0.5, 1, 2, 3, 4, and 5 mm on the blunt end of the eggshell. In experiment 2, 4 x 4 mm(2) windows were cut into the eggs or the eggs were drilled with 5 holes of bore diameter 2 mm on the blunt end of eggshell. In experiment 3, eggs were drilled with 1, 3, 5, and 7 holes of diameter 2 mm on the blunt end of eggshell. Eggs were treated on d 3 of each experiment and the amount of water loss was recorded on d 19 of incubation. Embryo growth was evaluated in experiments 2 and 3. In addition, the livers of embryos were collected in the 0-, 1-, 3-, and 5-hole treatment groups after weighing eggs to determine 3-hydroxy acyl coenzyme A dehydrogenase activity. In experiment 1, although higher water loss was observed in all windowed eggs than in control, there were no differences in amount of water loss among all bore diameters. Accordingly, that was not successful to control amount of water loss. In experiment 2, higher water loss was observed in drilled eggs at the same levels in windowed eggs as in control. Drilling holes was a more useful treatment to control amount of water loss on incubated eggs than windowing. In experiment 3, amount of water loss increased linearly with increasing number of holes on the blunt end of eggshell. Hepatic 3-hydroxy acyl coenzyme A dehydrogenase activity increased with increasing the number of drilled holes.
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PMID:Development of a method to control the water evaporation of hatching eggs during incubation. 2018 73

The multifunctional enzyme, type-1 (MFE1) is involved in several lipid metabolizing pathways. It catalyses: (a) enoyl-CoA isomerase and (b) enoyl-CoA hydratase (EC 4.2.1.17) reactions in its N-terminal crotonase part, as well as (3) a 3S-hydroxy-acyl-CoA dehydrogenase (HAD; EC 1.1.1.35) reaction in its C-terminal 3S-hydroxy-acyl-CoA dehydrogenase part. Crystallographic binding studies with rat peroxisomal MFE1, using unbranched and branched 2E-enoyl-CoA substrate molecules, show that the substrate has been hydrated by the enzyme in the crystal and that the product, 3S-hydroxy-acyl-CoA, remains bound in the crotonase active site. The fatty acid tail points into an exit tunnel shaped by loop-2. The thioester oxygen is bound in the classical oxyanion hole of the crotonase fold, stabilizing the enolate reaction intermediate. The structural data of these enzyme product complexes suggest that the catalytic base, Glu123, initiates the isomerase reaction by abstracting the C2-proton from the substrate molecule. Subsequently, in the hydratase reaction, Glu123 completes the catalytic cycle by reprotonating the C2 atom. A catalytic water, bound between the OE1-atoms of the two catalytic glutamates, Glu103 and Glu123, plays an important role in the enoyl-CoA isomerase and the enoyl-CoA hydratase reaction mechanism of MFE1. The structural variability of loop-2 between MFE1 and its monofunctional homologues correlates with differences in the respective substrate preferences and catalytic rates.
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PMID:The isomerase and hydratase reaction mechanism of the crotonase active site of the multifunctional enzyme (type-1), as deduced from structures of complexes with 3S-hydroxy-acyl-CoA. 2335 Oct 63