Delta3,5-delta2,4-dienoyl-CoA isomerase from rat liver. Molecular characterization.
rECH1, a recently identified rat cDNA (FitzPatrick, D. R., Germain-Lee, E., and Valle, D. (1995) Genomics 27, 457-466) encodes a polypeptide belonging to the hydratase/isomerase superfamily. We modeled the structure of rECH1 based on rat mitochondrial 2-enoyl-CoA hydratase 1. The model predicts that rECH1p has the hydratase fold in the core domain and two domains for interaction with other subunits. When we incubated 3,5,8,11, 14-eicosapentaenoyl-CoA with purified rECH1p, the spectral data suggested a switching of the double bonds from the Delta3-Delta5 to the Delta2-Delta4 positions. This was confirmed by demonstrating that the product was a valid substrate for 2,4-dienoyl-CoA reductase. These results indicate that rECH1p is Delta3,5-Delta2,4-dienoyl-CoA isomerase. Subcellular fractionation and immunoelectron microscopy using antibodies to a synthetic polypeptide derived from the C terminus of rECH1p showed that rECH1p is located in the matrix of both mitochondria and peroxisomes in rat liver. Consistent with these observations, the 36,000-Da rECH1p has a potential N-terminal mitochondrial targeting signal as well as a C-terminal peroxisomal targeting signal type 1. Transport of the protein into the mitochondria with cleavage of the targeting signal results in a mature mitochondrial form with a molecular mass of 32,000 Da; transport to peroxisomes yields a protein of 36,000 Da.
Filppula
S A
SA
Biocenter Oulu and Department of Biochemistry, University of Oulu, Linnanmaa, FIN-90570 Oulu, Finland.
Yagi
A I
AI
Kilpeläinen
S H
SH
Novikov
D
D
FitzPatrick
D R
DR
Vihinen
M
M
Valle
D
D
Hiltunen
J K
JK
D000595:Amino Acid Sequence; D000818:Animals; D019748:Carbon-Carbon Double Bond Isomerases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D016253:Microscopy, Immunoelectron; D008930:Mitochondria, Liver / Q000201:enzymology* / Q000648:ultrastructure; D008958:Models, Molecular; D008969:Molecular Sequence Data; D051381:Rats; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions / Q000201:enzymology
D019748:Carbon-Carbon Double Bond Isomerases; C085575:delta(3,5),delta(2,4)-dienoyl-CoA isomerase