Functional analysis of human microsomal epoxide hydrolase genetic variants

Abstract

Human microsomal epoxide hydrolase (EPHX1) is active in the metabolism of many potentially carcinogenic or otherwise genotoxic epoxides, such as those derived from the oxidation of polyaromatic hydrocarbons. EPHX1 is polymorphic and encodes allelic variation at least two amino acid positions, Y113H and H139R. In a number of recent molecular epidemiological investigations, EPHX1 polymorphism has been suggested as a susceptibility factor for several human diseases. To better evaluate the functional contribution of EPHX1 genetic polymorphism, we characterized the enzymatic properties associated with each of the respective variant proteins. Enzymatic profiles were evaluated with cis-stilbene oxide (cSO) and benzo[a]pyrene-4,5-epoxide (BaPO), two prototypical substrates for the hydrolase. In one series of experiments, activities of recombinant EPHX1 proteins were analyzed subsequent to their expression using the pFastbac ® baculovirus vector in Spodoptera frugiperda-9 (Sf9) insect cells, and purification by column chromatography. In parallel studies, EPHX1 activities were evaluated with human liver microsomes derived from individuals of known EPHX1 genotype. Using the purified protein preparations, rates of cSO and BaPO hydrolysis for the reference protein, Y113/H139, were approximately 2-fold greater than those measured with the other EPHX1 allelic variants. However, when activities were analyzed using human liver microsomal fractions, no major differences were evident in the reaction rates generated among preparations representing the different EPHX1 alleles. Collectively, these results suggest that the structural differences encoded by the Y113H and H139R variant alleles exert only modest impact on EPHX1-specific enzymatic activities in vivo.

System review and metaanalysis of the relationships between five metabolic gene polymorphisms and colorectal adenoma risk

“…Previous in vitro study found that EPHX1 Tyr113His was associated with 40% of decreased enzyme activity, while His139Arg was associated with 25% of increased enzyme activity [40]. Recent experimental studies have demonstrated that EPHX1 genetic polymorphisms had limited effect on the activity of the enzyme and that interindividual differences of EPHX1 gene expression greatly varied, ranging from 2 to 10 folds [41,42]. A recent study by Yang et al [43] found that the upstream E1-b promoter of EPHX1 was the major regulator of EPHX1 expression in human tissues, and that polymorphism in this region might contribute to an interindividual risk determinant to xenobiotic-induced toxicities.…”

Associations between Smoking, Polymorphisms in Polycyclic Aromatic Hydrocarbon (PAH) Metabolism and Conjugation Genes and PAH-DNA Adducts in Prostate Tumors Differ by Race

“…The CYP1A1 Ile 462 Val and CYP3A4 A(À392)G polymorphisms have variant alleles with higher enzymatic activity compared with their respective wild-type alleles (23,24), and the activity of the CYP1B1 Ala 119 Ser and Leu 432 Val variants is substrate dependent with 432 Val/ 119 Ala, having slightly higher activity in metabolizing benzo(a)pyrene-7,8-dihydrodiols but slightly lower activity in metabolizing parent benzo(a)pyrene than 432 Leu/ 119 Ala (15). Effects of the mEH Try 113 His and mEH His 139 Arg polymorphisms remain unclear; an earlier study reported that 113 Tyr/ 139 Arg had the most activity in hydrolyzing benzo(a)pyrene-epoxides to benzo(a)-pyrene-dihydrodiols (25), but recent work shows the 113 Tyr/ 139 His combination may be the most active (26). The CYP1A1 Ile 462 Val, CYP1B1 Leu 432 Val, CYP3A4 A(À392)G polymorphisms have been equivocally associated with prostate cancer (27)(28)(29)(30)(31), which may be attributed, in part, to heterogeneity in PAH exposure.…”

Integrating Field Analyses with Laboratory Exposures to Assess Ecosystems Health

“…Microsomal epoxide hydrolase catalyses the hydrolysis of various epoxides and reactive epoxide intermediates of numerous endobiotic and xenobiotic chemicals into less reactive and more polar dihydrodiols Hosagrahara et al, 2004). Although it is mainly known to be a protective enzyme by decomposing numerous epoxide intermediates, it also mediates activation of PAHs to highly reactive diol epoxides, in conjunction with CYPs Gonzalez et al, 1991;Hecht, 2002a).…”

“…(vi) Genetic variability of epoxide hydrolase Two genetic polymorphisms of the mEH gene that encodes microsomal epoxide hydrolase have been reported to affect its catalytic activities Kiyohara et al, 2002a;Gsur et al, 2003;Hosagrahara et al, 2004). The Tyr113His variant (EH3) was found in exon 3 and caused a decrease in catalytic activities.…”

“…However one study showed that the specific activity of microsomal epoxide hydrolase was similar for each variant . The structural differences encoded by the Tyr113His and His139Arg variants have been reported to exert only a modest impact on the activities of microsomal epoxide hydrolase, using cis-stilbene oxide and benzo [a]pyrene-4,5-oxide as substrates Omiecinski et al, 2000;Hosagrahara et al, 2004).…”