Analysis of the beta-oxidation of trans-unsaturated fatty acid in recombinant Saccharomyces cerevisiae expressing a peroxisomal PHA synthase reveals the involvement of a reductase-dependent pathway.
The degradation of fatty acids having cis- or trans-unsaturated bond at an even carbon was analyzed in Saccharomyces cerevisiae by monitoring polyhydroxyalkanoate production in the peroxisome. Polyhydroxyalkanaote is synthesized by the polymerization of the beta-oxidation intermediates 3-hydroxy-acyl-CoAs via a bacterial polyhydroxyalkanoate synthase targeted to the peroxisome. The synthesis of polyhydroxyalkanoate in cells grown in media containing 10-cis-heptadecenoic acid was dependent on the presence of 2,4-dienoyl-CoA reductase activity as well as on Delta3,Delta2-enoyl-CoA isomerase activity. The synthesis of polyhydroxyalkanoate from 10-trans-heptadecenoic acid in mutants devoid of 2,4-dienoyl-CoA reductase revealed degradation of the trans fatty acid directly via the enoyl-CoA hydratase II activity of the multifunctional enzyme (MFE), although the level of polyhydroxyalkanoate was 10-25% to that of wild type cells. Polyhydroxyalkanoate produced from 10-trans-heptadecenoic acid in wild type cells showed substantial carbon flux through both a reductase-dependent and a direct MFE-dependent pathway. Flux through beta-oxidation was more severely reduced in mutants devoid of Delta3,Delta2-enoyl-CoA isomerase compared to mutants devoid of 2,4-dienoyl-CoA reductase. It is concluded that the intermediate 2-trans,4-trans-dienoyl-CoA is metabolized in vivo in yeast by both the enoyl-CoA hydratase II activity of the multifunctional protein and the 2,4-dienoyl-CoA reductase, and that the synthesis of the intermediate 3-trans-enoyl-CoA in the absence of the Delta3,Delta2-enoyl-CoA isomerase leads to the blockage of the direct MFE-dependent pathway in vivo.
Robert
Julien
J
Département de Biologie Moléculaire Végétale, Bâtiment de Biologie, Université de Lausanne, CH-1015 Lausanne, Switzerland.
Marchesini
Silvia
S
Delessert
Syndie
S
Poirier
Yves
Y
D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry; D005231:Fatty Acids, Unsaturated / Q000737:chemistry*; D008968:Molecular Conformation; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism*; D020675:Peroxisomes / Q000737:chemistry / Q000201:enzymology; D011550:Pseudomonas aeruginosa / Q000201:enzymology; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics
D003470:Culture Media; D005231:Fatty Acids, Unsaturated; D010088:Oxidoreductases; D000217:Acyltransferases; C077225:poly(3-hydroxyalkanoic acid) synthase