Function of human mitochondrial 2,4-dienoyl-CoA reductase and rat monofunctional Delta3-Delta2-enoyl-CoA isomerase in beta-oxidation of unsaturated fatty acids.

Human 2,4-dienoyl-CoA reductase (2,4-reductase; DECR) and rat monofunctional ∆ 3 -∆ 2 -enoyl-CoA isomerase (rat 3,2-isomerase; ECI) are thought to be mitochondrial auxiliary enzymes involved in the β-oxidation of unsaturated fatty acids. However, their function during this process has not been demonstrated. Although they lack obvious peroxisomal targeting signals (PTSs), both proteins have been suggested previously to also occur in the mammalian peroxisomal compartment. The putative function and peroxisomal location of the two mammalian proteins can be examined in yeast, since β-oxidation of unsaturated fatty acids is a compartmentalized process in Saccharomyces cere v isiae requiring peroxisomal 2,4-dienoyl-CoA reductase (Sps19p) and peroxisomal 3,2-isomerase (Eci1p). A yeast sps19 ∆ mutant expressing human 2,4-reductase ending with the native C-terminus could not grow on petroselinic acid [ cis -C 18:1(6) ] medium but could grow when the protein was extended with a PTS tripeptide, SKL (Ser-Lys-Leu). We therefore reason that the human protein is a physiological 2,4-reductase but that it is probably not peroxisomal. Rat 3,2-isomerase expressed in a yeast eci1 ∆ strain was able to re-establish growth on oleic acid [ cis -C 18:1(9) ] medium irrespective of an SKL extension. Since we had shown that ∆ 2,4 double bonds could not be metabolized extra-peroxisomally to restore growth of the sps19 ∆ strain, we postulate that rat 3,2-isomerase acted on the ∆ 3 unsaturated metabolite of oleic acid by replacing the mutant9s missing activity from within the peroxisomes. Immunoblotting of fractionated yeast cells expressing rat 3,2-isomerase in combination with electron microscopy supported our proposal that the protein functioned in peroxisomes. The results presented here shed new light on the function and location of human mitochondrial 2,4-reductase and rat monofunctional 3,2-isomerase.