Evidence for a peroxisomal fatty acid beta-oxidation involving D-3-hydroxyacyl-CoAs. Characterization of two forms of hydro-lyase that convert D-(-)-3-hydroxyacyl-CoA into 2-trans-enoyl-CoA.
A novel D-(-)-3-hydroxyacyl-CoA hydro-lyase, forming 2-trans-enoyl-CoA and formerly designated as epimerase (EC 5.1.2.3), was extracted from fat-degrading cotyledons of cucumber seedlings. The enzyme, called D-3-hydroxyacyl-CoA hydro-lyase or D-specific 2-trans-enoyl-CoA hydratase, is shown to be required for the degradation of unsaturated fatty acids that contain double bonds extending from even-numbered C atoms. The D-3-hydroxyacyl-CoA hydro-lyase was exclusively localized within peroxisomes. A 10,000-fold purification by chromatography on a hydrophobic matrix, a cation exchanger, on hydroxyapatite and Mono S led to two proteins of apparent homogeneity, both exhibiting Mr of 65,000. The D-3-hydroxyacyl-CoA hydro-lyases are homodimers with slightly differing isoelectric points around pH = 9.0. They catalyze the conversion of 2-trans-enoyl-CoA into D-3-hydroxyacyl-CoA. The reverse reaction was observed but no reaction with 2-cis-enoyl-CoAs or L-3-hydroxyacyl-CoAs. 2-trans-Decenoyl-CoA was converted 10-times faster than 2-trans-butenoyl-CoA. The conversion of 4-cis-decenoyl-CoA into octenoyl-CoA was demonstrated in vitro with purified proteins with an assay mixture containing acyl-CoA oxidase, multifunctional protein, thiolase and the D-3-hydroxyacyl-CoA hydro-lyase. Comparisons of enzyme activities present in the cotyledons or isolated peroxisomes clearly show that the pathway via dienoyl-CoA reductase is much less effective than the sequence involving D-3-hydroxyacyl-CoA hydro-lyase.
Engeland
K
K
Department of Biochemistry, Philipps-University, Marburg, Federal Republic of Germany.
Kindl
H
H
D015094:3-Hydroxyacyl CoA Dehydrogenases / Q000737:chemistry* / Q000378:metabolism; D000214:Acyl Coenzyme A / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004746:Enoyl-CoA Hydratase / Q000737:chemistry* / Q000378:metabolism; D005231:Fatty Acids, Unsaturated / Q000378:metabolism*; D006836:Hydro-Lyases / Q000737:chemistry* / Q000302:isolation & purification; D007526:Isoelectric Point; D007535:Isomerases / Q000737:chemistry* / Q000378:metabolism; D008830:Microbodies / Q000201:enzymology*; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000378:metabolism; D063994:Peroxisomal Bifunctional Enzyme; D010944:Plants / Q000201:enzymology; D019998:Racemases and Epimerases / Q000737:chemistry* / Q000378:metabolism; D012639:Seeds / Q000201:enzymology; D013237:Stereoisomerism
D000214:Acyl Coenzyme A; D005231:Fatty Acids, Unsaturated; D009097:Multienzyme Complexes; C034681:octanoyl-coenzyme A; C045561:decanoyl-coenzyme A; D015094:3-Hydroxyacyl CoA Dehydrogenases; C065185:D-3-hydroxyacyl CoA dehydratase; D006836:Hydro-Lyases; D004746:Enoyl-CoA Hydratase; D063994:Peroxisomal Bifunctional Enzyme; D007535:Isomerases; D019998:Racemases and Epimerases; C029601:3-hydroxybutyryl-CoA epimerase