DELTA(3,5)-DELTA(2,4)-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 stru cture 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 d ata suggested a switching of the double bonds from the Delta(3)-Delta( 5) to the Delta(2)-Delta(4) positions. This was confirmed by demonstra ting that the product was a valid substrate for 2,4-dienoyl-CoA reduct ase, These results indicate that rECH1p is Delta(3,5)-Delta(2,4)-dieno yl-CoA isomerase, Subcellular fractionation and immunoelectron microsc opy using antibodies to a synthetic polypeptide derived from the C ter minus of rECH1p showed that rECH1p is located in the matrix of both mi tochondria and peroxisomes in rat liver, Consistent with these observa tions, the 36,000-Da rECH1p has a potential N-terminal mitochondrial t argeting signal as well as a C-terminal peroxisomal targeting signal t ype 1, Transport of the protein into the mitochondria with cleavage of the targeting signal results in a mature mitochondrial form with a mo lecular mass of 32,000 Da; transport to peroxisomes yields a protein o f 36,000 Da.