Alternatives to the Isomerase-dependent Pathway for the β-Oxidation of Oleic Acid Are Dispensable inSaccharomyces cerevisiae

doi:10.1074/jbc.274.35.24514

Alternatives to the Isomerase-dependent Pathway for the β-Oxidation of Oleic Acid Are Dispensable inSaccharomyces cerevisiae

IDENTIFICATION OF YOR180c/DCI1 ENCODING PEROXISOMAL Δ^3,5-Δ^2,4-DIENOYL-CoA ISOMERASE*

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

Abstract

Fatty acids with double bonds at odd-numbered positions such as oleic acid can enter β-oxidation via a pathway relying solely on the auxiliary enzyme Δ³-Δ²-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 Δ^3,5-Δ^2,4-dienoyl-CoA isomerase (di-isomerase-dependent) or on Δ^3,5-Δ^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 × min⁻¹ per mg of protein. Similarly enriched extracts from eci1Δ 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 theeci1Δ 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 theeci1Δ 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 eci1Δ 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.

Footnotes

↵* This work was supported in part by Fonds zur Förderung der Wissenschaftlichen Forschung, Vienna, Austria, Grants P12061 (to B. Hamilton) and P12118 (to A. H.), by Jubiläumsfonds derÖsterreichischen Nationalbank, Austria, Grant 6517 (to H. R.), and by the Sigrid Juselius Foundation, Finland, and the Academy of Finland (to J. K. H.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¶ To whom correspondence should be addressed. Present address: Inst. für Biochemie, Limonenstrasse 7, 12203 Berlin, Germany. Tel.: 49-30-838-2937; Fax: 49-30-838-2936; E-mail: hpr@zedat.fu-berlin.de
↵2 Reserved in the Saccharomyces Genome Data Base as an enoyl-CoA hydratase homologue; I. V. Karpichev, J. Lopez, and G. M. Small, manuscript in preparation.
Abbreviations:

MFE

multifunctional enzyme

2,4-reductase

2,4-dienoyl-CoA reductase

3,2-isomerase

Δ³-Δ²-enoyl-CoA isomerase

di-isomerase

Δ^3,5-Δ^2,4-dienoyl-CoA isomerase

PCR

polymerase chain reaction

kb

kilobase

GFP

green fluorescent protein

DAPI

4′,6-diamidino-2-phenylindole

ORE

oleate response element

PTS

peroxisomal targeting signal
- Received January 11, 1999.
- Revision received May 12, 1999.
The American Society for Biochemistry and Molecular Biology, Inc.