MULTIENZYME COMPLEXES OF FATTY ACID OXIDATION FROM WILD TYPE ESCHERICHIA COLI AND FROM FATTY ACID OXIDATION MUTANTS.
Item
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Title
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MULTIENZYME COMPLEXES OF FATTY ACID OXIDATION FROM WILD TYPE ESCHERICHIA COLI AND FROM FATTY ACID OXIDATION MUTANTS.
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Identifier
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AAI8401951
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identifier
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8401951
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Creator
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PRAMANIK, AJAY.
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Contributor
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Horst Schulz
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Date
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1983
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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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Biology, Microbiology
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Abstract
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An E. coli mutant (fadB64) with a defective L-3-hydroxyacyl-CoA dehydrogenase (EC 1.1.1.35) which is unable to grow on long chain fatty acids as the sole carbon source, was shown to possess a fatty acid oxidation complex that contains five (beta)-oxidation enzymes including L-3-hydroxyacyl-CoA dehydrogenase. The fatty acid oxidation complex from this mutant was purified to near homogeneity and the complex from its parental strain was highly purified to near homogeneity. A comparative study of these two complexes and that from E. coli B demonstrated the immunological and gross structural identity of all three fatty acid oxidation complexes despite significant differences between their specific activities. A kinetic evaluation of the complexes led to the conclusion that the mutation has affected the active site of L-3-hydroxyacyl-CoA dehyrogenase so that it is inactive with acetoacetyl-CoA as a substrate, but exhibits an increasing percentage of the parental dehydrogenase activity with increasing chain length of the substrate. However, even with 3-ketodecanoyl-CoA as a substrate the mutant complex is 20-times less active than the wild-type complex. This behavior is apparently due to a strong dependence of the maximal velocity on the chain length of the substrate whereas the K(,m) value for the substrate seems to be little or not at all affected by the mutation.;An E. coli strain (RS 3084) with a mutation in the fadABC region was found to contain the parental level of long-chain enoyl-CoA hydratase while being devoid of any of the enzyme activities associated with the fatty acid oxidation complex. However, other fadABC mutants including mutant fad5 did neither contain long-chain enoyl-CoA hydratase nor the fatty acid oxidation complex. These observations together suggest that long-chain enoyl-CoA is a gene product of the fadABC operon but is not associated with the fatty acid oxidation complex.;Evidence is also presented for the absence of cis-(DELTA)('3)-trans-(DELTA)('2)-enoyl-CoA isomerase and 3-hydroxyacyl-CoA epimerase activities in the peroxisomal bifunctional enzyme which exhibits enoyl-CoA hydratase and L-3-hydroxyacyl-CoA dehydrogenase activities.
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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.
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Program
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Biochemistry
