Abstract
2,4-Dienoyl-CoA reductases, enzymes of the beta-oxidation of unsaturated fatty acids which were purified from bovine liver and oleate-induced cells of Escherichia coli, revealed very similar substrate specificities but distinctly different molecular properties. The subunit molecular weights, estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were 32,000 and 73,000 for the mammalian and the bacterial enzyme, respectively. The native molecular weights, calculated from sedimentation coefficients and Stokes radii yielded 124,000 for the bovine liver and 70,000 for the bacterial enzyme. Thus, bovine liver 2,4-dienoyl-CoA reductase is a tetramer consisting of four identical subunits. The E. coli 2,4-dienoyl-CoA reductase, however, possesses a monomeric structure. The latter enzyme contains 1 mol of FAD/mol of enzyme, whereas the former reductase is not a flavoprotein. The bovine liver reductase reduced 2-trans, 4-cis- and 2-trans,4-trans-decadienoyl-CoA to 3-trans-decenoyl-CoA. The E. coli reductase catalyzed the reduction of the same two substrates but in contrast yielded 2-trans-decenoyl-CoA as reaction product. Certain other properties of the two 2,4-dienoyl-CoA reductases are also presented. The localization of the reductase step within the degradation pathway of 4-cis-decenoyl-CoA, a metabolite of linoleic acid, is discussed.
Highlights
From the Institutf u r Physiologische Chenie, Abteilung fur NaturwissenschftlicheMedizirz, Ruhr-Uniuersitiit Bochum, Postfach 102 148, 0-4630 Bochum 1, Federal Republic of Germany
Bovine liver 2,4-dienoyl-CoA reductase is a tetramer consisting of four identical subunits
Since thevalues obtained by the two methods did not agree closely, the molecularweights were calculated by the method of Siegel and Monty[28] from small Sephadex G-25 column, and storeadt -18 “C. This purification sedimentation coefficients ( s ~,o~f)the two 2,4-dienoyl-CoA
Summary
From the Institutf u r Physiologische Chenie, Abteilung fur NaturwissenschftlicheMedizirz, Ruhr-Uniuersitiit Bochum, Postfach 102 148, 0-4630 Bochum 1, Federal Republic of Germany. From bovine liver and oleate-induced cells of Esche- trans,4-cis-dienoyl-CoA esters according tothe epimerase richia coli, revealed very simsiulabrstrate specificities but distinctlydifferent molecular properties. Fatty Acids and Acyl-CoA Esters tivity has been determined in liver [1,4,5,6,7] and heart [2,7,8,9] 2-trans-Decen-4-ynoic acid was synthesized according to Crombie of different mammalian species as well as ineucaryotic Stokes radii of the of 2,4-dienoyl-CoA reductase activity)were prepared in mM potassium phosphate buffer, pH 7.4, containing 1mM EDTA and 0.5 mM dithioerythritol (buffer A), and theDEAE-cellulose column (2.2 X 12 cm, Whatman DE52) was performed as describedpreviously [10]. The DE52 fraction was applied to a 2’,5’-ADP-Sepharose 4B column containing 1 mM EDTA and 0.5 mM dithioerythritol (buffer B), and standard proteins were taken from the literature [28].
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