J Biol Chem, Vol. 274, Issue 35, 24514-24521, August 27, 1999

Alternatives to the Isomerase-dependent Pathway for the -Oxidation of Oleic Acid Are Dispensable in Saccharomyces cerevisiae
IDENTIFICATION OF YOR180c/DCI1 ENCODING PEROXISOMAL ^3,5-^2,4-DIENOYL-CoA ISOMERASE

Aner Gurvitz^§, Anu M. Mursula^§, Ahmed I. Yagi^§, Andreas Hartig, Helmut Ruis, Hanspeter Rottensteiner, and J. Kalervo Hiltunen^§

From the  Institut für Biochemie und Molekulare Zellbiologie der Universität Wien and Ludwig Boltzmann-Forschungsstelle für Biochemie, Vienna Biocenter, Dr Bohrgasse 9, A-1030 Wien, Austria and the ^§ Biocenter Oulu, Department of Biochemistry, University of Oulu, FIN-90570 Oulu, Finland

Fatty acids with double bonds at odd-numbered positions such as oleic acid can enter -oxidation via a pathway relying solely on the auxiliary enzyme ³-²-enoyl-CoA isomerase, termed the isomerase-dependent pathway. Two novel alternative pathways have recently been postulated to exist in mammals, and these additionally depend on ^3,5-^2,4-dienoyl-CoA isomerase (di-isomerase-dependent) or on ^3,5-^2,4-dienoyl-CoA isomerase and 2,4-dienoyl-CoA reductase (reductase-dependent). We report the identification of the Saccharomyces cerevisiae oleic acid-inducible DCI1 (YOR180c) gene encoding peroxisomal di-isomerase. Enzyme assays conducted on soluble extracts derived from yeast cells overproducing Dci1p using 3,5,8,11,14-eicosapentenoyl-CoA as substrate demonstrated a specific di-isomerase activity of 6 nmol × min¹ per mg of protein. Similarly enriched extracts from eci1 cells lacking peroxisomal 3,2-isomerase additionally contained an intrinsic 3,2-isomerase activity that could generate 3,5,8,11,14-eicosapentenoyl-CoA from 2,5,8,11,14-eicosapentenoyl-CoA but not metabolize trans-3-hexenoyl-CoA. Amplification of this intrinsic activity replaced Eci1p since it restored growth of the eci1 strain on petroselinic acid for which di-isomerase is not required whereas Eci1p is. Heterologous expression in yeast of rat di-isomerase resulted in a peroxisomal protein that was enzymatically active but did not re-establish growth of the eci1 mutant on oleic acid. A strain devoid of Dci1p grew on oleic acid to wild-type levels, whereas one lacking both Eci1p and Dci1p grew as poorly as the eci1 mutant. Hence, we reasoned that yeast di-isomerase does not additionally represent a physiological 3,2-isomerase and that Dci1p and the postulated alternative pathways in which it is entrained are dispensable for degrading oleic acid.