Effect of Mutagenesis on the Stereochemistry of Enoyl-CoA Hydratase

Feng, Yixiang; Hofstein, Hilary A.; Zwahlen, Jacque; Tonge, Peter J.

doi:10.1021/bi020382g

Cited by 16 publications

(22 citation statements)

References 30 publications

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“…Two active site glutamic acids have been proposed to act in catalysis (62,63), and an oxyanion hole has been proposed to polarize the thioester carbonyl group. The N-terminal Glu activates a water molecule for nucleophilic attack at the C3 position (hereafter called Glu N (Glu 119 in AtMFP2)).…”

Section: Discussionmentioning

confidence: 99%

The Multifunctional Protein in Peroxisomal β-Oxidation

Arent¹,

Christensen²,

Pye³

et al. 2010

Journal of Biological Chemistry

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Plant fatty acids can be completely degraded within the peroxisomes. Fatty acid degradation plays a role in several plant processes including plant hormone synthesis and seed germination. Two multifunctional peroxisomal isozymes, MFP2 and AIM1, both with 2-trans-enoyl-CoA hydratase and L-3-hydroxyacyl-CoA dehydrogenase activities, function in mouse ear cress (Arabidopsis thaliana) peroxisomal ␤-oxidation, where fatty acids are degraded by the sequential removal of two carbon units. A deficiency in either of the two isozymes gives rise to a different phenotype; the biochemical and molecular background for these differences is not known. Structure determination of Arabidopsis MFP2 revealed that plant peroxisomal MFPs can be grouped into two families, as defined by a specific pattern of amino acid residues in the flexible loop of the acylbinding pocket of the 2-trans-enoyl-CoA hydratase domain. This could explain the differences in substrate preferences and specific biological functions of the two isozymes. The in vitro substrate preference profiles illustrate that the Arabidopsis AIM1 hydratase has a preference for short chain acyl-CoAs compared with the Arabidopsis MFP2 hydratase. Remarkably, neither of the two was able to catabolize enoyl-CoA substrates longer than 14 carbon atoms efficiently, suggesting the existence of an uncharacterized long chain enoyl-CoA hydratase in Arabidopsis peroxisomes.Fatty acids are degraded by the sequential removal of two carbon units in a process known as ␤-oxidation (Fig. 1). This process is ubiquitous and takes its name from the oxidation taking place at the carbon atom ␤ to the carboxyl group. The discovery of a peroxisomal ␤-oxidation system was made in plants (1), and whereas peroxisomal ␤-oxidation in animals appears to be merely a fatty acid chain-shortening machine feeding mitochondrial ␤-oxidation, plant and fungal ␤-oxidation takes place almost entirely in the peroxisomes. The products of the reaction are H 2 O 2 , acetyl-CoA, reducing equivalents, and a variety of chain length-shortened acyl-containing molecules like the plant hormones jasmonic acid (2) and indole-3-acetic acid (3, 4).The conversion of storage triacylglycerols by ␤-oxidation provides metabolic energy and carbon skeletons for germination and early post-germinative seedling growth in oil seed plants (5) and metabolic energy and carbon for the production of hydrolytic enzymes in cereals (6). It is a salvage pathway for fatty acids during foliar senescence (7), supplies respiratory substrates to carbohydrate-deprived tissue (8), and at a lower level, is a constitutive property of all plant tissues most likely involved with membrane lipid turnover (7).Three proteins (each present as several isozymes) that host a total of four enzyme activities constitute the core of peroxisomal ␤-oxidation. Acyl-CoA oxidase (ACX) 6 oxidizes acyl-CoA to 2-trans-enoyl-CoA using FAD as co-enzyme. A multifunctional protein (MFP) adds water over the 2-trans-enoyl-CoA double bond and oxidizes the resultant L-3-hydroxyacylCoA usin...

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Section: Discussionmentioning

confidence: 99%

The Multifunctional Protein in Peroxisomal β-Oxidation

Arent¹,

Christensen²,

Pye³

et al. 2010

Journal of Biological Chemistry

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“…However, DmdD is not nearly as efficient as an ECH at catalyzing the hydration of crotonyl-CoA, with a catalytic efficiency of 2100 mM -1 s -1 compared with the typical values of 45000–119000 mM -1 s -1 of other crotonases. DmdD instead displays a K m of 0.008 mM for MTA-CoA and a high catalytic efficiency, 5400 mM -1 s -1 (Kiema et al, 1999; Feng et al, 2002; Tan et al, 2013). This greater catalytic efficiency only applies to reactions with MTA-CoA and appears to be due, at least in part, to the structure of MTA-CoA.…”

Section: Dmsp Specific Enoyl-coa Hydratases: Dmdd and Acuhmentioning

confidence: 99%

Evolution of Dimethylsulfoniopropionate Metabolism in Marine Phytoplankton and Bacteria

2017

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The elucidation of the pathways for dimethylsulfoniopropionate (DMSP) synthesis and metabolism and the ecological impact of DMSP have been studied for nearly 70 years. Much of this interest stems from the fact that DMSP metabolism produces the climatically active gas dimethyl sulfide (DMS), the primary natural source of sulfur to the atmosphere. DMSP plays many important roles for marine life, including use as an osmolyte, antioxidant, predator deterrent, and cryoprotectant for phytoplankton and as a reduced carbon and sulfur source for marine bacteria. DMSP is hypothesized to have become abundant in oceans approximately 250 million years ago with the diversification of the strong DMSP producers, the dinoflagellates. This event coincides with the first genome expansion of the Roseobacter clade, known DMSP degraders. Structural and mechanistic studies of the enzymes of the bacterial DMSP demethylation and cleavage pathways suggest that exposure to DMSP led to the recruitment of enzymes from preexisting metabolic pathways. In some cases, such as DmdA, DmdD, and DddP, these enzymes appear to have evolved to become more specific for DMSP metabolism. By contrast, many of the other enzymes, DmdB, DmdC, and the acrylate utilization hydratase AcuH, have maintained broad functionality and substrate specificities, allowing them to carry out a range of reactions within the cell. This review will cover the experimental evidence supporting the hypothesis that, as DMSP became more readily available in the marine environment, marine bacteria adapted enzymes already encoded in their genomes to utilize this new compound.

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“…ECH (XI) was the first CS member for which a crystal structure was solved by Engel and co-workers in 1996 [32]; the structure rationalized previous kinetic and mutagenesis studies. Two glutamyl residues (Glu144 and Glu164) are essential for ECH (XI) catalysis [33,34]. One proposed mechanism for ECH (XI) involves conjugate addition of water with general acid/base catalysis from Glu144 and Glu164 with polarization of the thioester carbonyl group by the OAH (Fig.…”

Section: Active Sitesmentioning

confidence: 99%

“…Glu196 is also hydrogen bonded to Asp176, and this interaction may help Glu196 to achieve the appropriate pKa for catalysis. Glu196 of DCI (XII) is similarly positioned to the catalytically important Glu164 of ECH (XI) [33,34].…”

Section: Active Sitesmentioning

confidence: 99%

Mechanisms and structures of crotonase superfamily enzymes – How nature controls enolate and oxyanion reactivity

Hamed

Batchelar

Clifton

et al. 2008

Cell. Mol. Life Sci.

112

145

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Structural and mechanistic studies on the crotonase superfamily (CS) are reviewed with the aim of illustrating how a conserved structural platform can enable catalysis of a very wide range of reactions. Many CS reactions have precedent in the 'carbonyl' chemistry of organic synthesis; they include alkene hydration/isomerization, aryl-halide dehalogenation, (de)carboxylation, CoA ester and peptide hydrolysis, fragmentation of beta-diketones and C-C bond formation, cleavage and oxidation. CS enzymes possess a canonical fold formed from repeated betabetaalpha units that assemble into two approximately perpendicular beta-sheets surrounded by alpha-helices. CS enzymes often, although not exclusively, oligomerize as trimers or dimers of trimers. Two conserved backbone NH groups in CS active sites form an oxyanion 'hole' that can stabilize enolate/oxyanion intermediates. The range and efficiency of known CS-catalyzed reactions coupled to their common structural platforms suggest that CS variants may have widespread utility in biocatalysis.

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Effect of Mutagenesis on the Stereochemistry of Enoyl-CoA Hydratase

Cited by 16 publications

References 30 publications

The Multifunctional Protein in Peroxisomal β-Oxidation

The Multifunctional Protein in Peroxisomal β-Oxidation

Evolution of Dimethylsulfoniopropionate Metabolism in Marine Phytoplankton and Bacteria

Mechanisms and structures of crotonase superfamily enzymes – How nature controls enolate and oxyanion reactivity

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