Purification of human liver cytosolic epoxide hydrolase and comparison to the microsomal enzyme.
Epoxide hydrolase (EC 3.3.2.3) was purified to electrophoretic homogeneity from human liver cytosol by using hydrolytic activity toward trans-8-ethylstyrene 7,8-oxide (TESO) as an assay. The overall purification was 400-fold. The purified enzyme has an apparent monomeric molecular weight of 58 000, significantly greater than the 50 000 found for human (or rat) liver microsomal epoxide hydrolase or for another TESO-hydrolyzing enzyme also isolated from human liver cytosol. Purified cytosolic TESO hydrolase catalyzes the hydrolysis of cis-8-ethylstyrene 7,8-oxide 10 times more rapidly than does the microsomal enzyme, catalyzes the hydrolysis of TESO and trans-stilbene oxide as rapidly as the microsomal enzyme, but catalyzes the hydrolysis of styrene 7,8-oxide, p-nitrostyrene 7,8-oxide, and naphthalene 1,2-oxide much less effectively than does the microsomal enzyme. Purified cytosolic TESO hydrolase does not hydrolyze benzo[a]pyrene 4,5-oxide, a substrate for the microsomal enzyme. The activities of the purified enzymes can explain the specific activities observed with subcellular fractions. Anti-human liver microsomal epoxide hydrolase did not recognize cytosolic TESO hydrolase in purified form or in cytosol, as judged by double-diffusion immunoprecipitin analysis, precipitation of enzymatic activity, and immunoelectrophoretic techniques. Cytosolic TESO hydrolase and microsomal epoxide hydrolase were also distinguished by peptide mapping. The results provide evidence that physically different forms of epoxide hydrolase exist in different subcellular fractions and can have markedly different substrate specificities.
D000293:Adolescent; D000328:Adult; D000818:Animals; D003600:Cytosol / Q000201:enzymology*; D000939:Epitopes / Q000276:immunology; D004851:Epoxide Hydrolases / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D004852:Epoxy Compounds / Q000378:metabolism; D005260:Female; D006801:Humans; D008099:Liver / Q000201:enzymology*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D008970:Molecular Weight; D010446:Peptide Fragments; D051381:Rats; D013343:Styrenes; D013379:Substrate Specificity
D000939:Epitopes; D004852:Epoxy Compounds; D010446:Peptide Fragments; D013343:Styrenes; C033332:4-nitrostyrene oxide; C027035:beta-ethylstyrene oxide; C013690:styrene oxide; D004851:Epoxide Hydrolases