Enzymology and mechanisms of double bond isomerizations during the beta -oxidation of unsaturated fatty acids.
Item
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Title
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Enzymology and mechanisms of double bond isomerizations during the beta -oxidation of unsaturated fatty acids.
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Identifier
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AAI3037166
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identifier
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3037166
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Creator
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Zhang, Dongyan.
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Contributor
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Adviser: Horst Schulz
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Date
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2002
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Language
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English
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Publisher
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City University of New York.
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Subject
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Chemistry, Biochemistry
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Abstract
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Delta3,5, Delta2,4-Dienoyl-CoA isomerase (dienoyl-CoA isomerase) is required in the alternative pathway for the degradation of unsaturated fatty acids with odd-numbered double bonds. Mitochondrial and peroxisomal dienoyl-CoA isomerases are products of the same gene with slightly different N-terminal sequences. Mutations of any of the three acidic amino acid residues located at the active site (Modis et al. (1998) Structure 6, 957--970) caused activity losses. In contrast to only a 10-fold decrease in activity upon replacement of Asp 176 by Ala, substitutions of Asp204 by Asn and of Glu196 by Gln resulted in 105-fold lower activities. Such activity losses are consistent with the direct involvement of these latter two residues in the proposed proton transfers at carbons 2 and 6 or 8 of the substrates. Probing of the wild-type and mutant forms of the enzyme with 2,5octadienoyl-CoA as substrate revealed low Delta2,Delta 3-enoyl-CoA isomerase and Delta5,Delta4 -enoyl-CoA isomerase activities catalyzed by Glu196 and Asp204, respectively. Altogether, this data reveals that positional isomerizations of the diene and triene are facilitated by simultaneous proton transfers involving Glu196 and Asp204, whereas each residue alone can catalyze, albeit less efficiently, a monoene isomerization.;Enoyl-CoA isomerase is required for the degradation of unsaturated fatty acids with double bonds at both odd-numbered and even-numbered positions. A functional analysis of rat liver Delta3,Delta 2-enoyl-CoA isomerases (enoyl-CoA isomerases) revealed the presence of mitochondrial enoyl-CoA isomerase (MECI) and enoyl-CoA isomerase (ECI) in mitochondria while peroxisomes contain ECI and multifunctional enzyme 1 (MFE1). ECI, which previously had been described as peroxisomal enoyl-CoA isomerase (PECI), was shown to be present in both mitochondria and peroxisomes. This enzyme seems to be identical with mitochondrial long-chain enoyl-CoA isomerase. All three known hepatic enoyl-CoA isomerases, MECI, ECI and MFE1, have broad chain length specificities but are distinguishable by their catalytic efficiencies that are optimal for 3-cis → 2-trans, 3-trans → 2-trans, and 2,5 → 3,5 isomerizations, respectively. An analysis based on their total tissue activities and substrate specificities prompts the conclusion that in mitochondria, MECI is responsible for the 3-cis → 2-trans and 2,5 → 3,5 isomerizations, while ECI is responsible for the 3-trans → 2-trans isomerization. In the peroxisomes, ECI may be the dominant enzyme for catalyzing 3-trans → 2-trans and 3-cis → 2-trans isomerizations of long-chain substrates, while MFE1 may play a key role in the 2,5 → 3,5 isomerization.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
