Epoxide cytosolic, microsomal

Reductions Epoxide hydroplase Azo and nitro reduction Carbonyl reductase Disulfide reduction Sulfoxide reduction Quinone reduction Reductive dehalogenation Microsomes, cytosol Gut microflora Cytosol Cytosol Cytosol Cytosol, microsomes Microsomes... 

Glutathione conjugation Glutathione (GSH) GSH-S-transferase (cytosol, microsomes) Epoxides, arene oxides, nitro groups, hydroxylamines Acetaminophen, ethacrynic acid, bromobenzene... 

According to biochemical separation, location, and substrate specificity, epoxide hydrolases (EH) have been divided into a number of groups. In mammals, the insoluble microsomal epoxide hydrolases and the soluble cytosolic epoxide hydrolases are enzymes of broad and complementary substrate specificity. 

A critical input in unraveling the catalytic mechanism of epoxide hydrolases has come from the identification of essential residues by a variety of techniques such as analysis of amino acid sequence relationships with other hydrolases, functional studies of site-directed mutated enzymes, and X-ray protein crystallography (e.g., [48][53][68 - 74]). As schematized in Fig. 10.6, the reaction mechanism of microsomal EH and cytosolic EH involves a catalytic triad consisting of a nucleophile, a general base, and a charge relay acid, in close analogy to many other hydrolases (see Chapt. 3). 

Fig. 10.6. Simplified representation of the postulated catalytic cycle of microsomal and cytosolic epoxide hydrolases, showing the roles played by the catalytic triad (i.e., nucleophile, general base, and charge relay acid) and some other residues, a) Nucleophilic attack of the substrate to form a /3-hydroxyalkyl ester intermediate, b) Nucleophilic attack of the /Thydroxyal-kyl ester by an activated H20 molecule, c) Tetrahedral transition state in the hydrolysis of the /f-hydroxyalkyl ester, d) Product liberation, with the enzyme poised for a further catalytic... 

An unusual case of intramolecular competition (chemoselectivity, see Chapt. 1 in [la]) between ester and oxirane occurs in the detoxification of (oxiran-2-yl)methyl 2-ethyl-2,5-dimethylhexanoate (10.49), one of the most abundant isomers of an epoxy resin. The compound is chemically very stable, i.e., resistant to aqueous hydrolysis, but is rapidly hydrolyzed in cytosolic and microsomal preparations by epoxide hydrolase and carboxylesterase, which attack the epoxide and ester groups, respectively [129], The rate of overall enzymatic hydrolysis was species dependent, decreasing in the order mouse > rat > human, but was relatively fast in all tissues examined (lung and skin as portals of entry, and liver as a further barrier). In mouse and rat lung microsomes, ester hydrolysis was 3-4 times faster than epoxide hydration, whereas the opposite was true in human lung microsomes. 

Polyunsaturated fatty acids in general and arachidonic acid in particular are especially of interest. With its four C=C bonds, arachidonic acid (10.51) can be oxidized by cytochrome P450 to four regioisomeric monoepoxides, namely the epoxyeicosatrienoic acids (EETs). The four epoxides, although chemically stable, were shown to be hydrolyzed to the corresponding vicinal diols by mouse liver cytosolic EH but not by microsomal EH... 

Interestingly, there is a marked species difference in the in vitro hydrolysis of carbamazepine 10,11-epoxide, such that the reaction was observed only in liver microsomes from humans but not in liver microsomal or cytosolic preparations from dogs, rabbits, hamsters, rats, or mice [181][196], Thus, carbamazepine appears to be a very poor substrate for EH, in analogy with the simpler analogues 10.129 (X = RN, RCH, or RCH=C). The human enzyme is exceptional in this respect, but not, however, in the steric course of the reaction. The diol formed (10.131, X = H2NCON) is mostly the trans-(10.S, 11. S )-enaniiomer [196], In other words, the product enantioselectivity of the hydration of carbamazepine epoxide catalyzed by human EH is the same as that of di benzol a,oxide catalyzed by rabbit microsomal EH, discussed above. 

F. Waechter, P. Bentley, F. Bieri, S. Muakkassah-Kelly, W. Staubli, M. Villermain, Organ Distribution of Epoxide Hydrolases in Cytosolic and Microsomal Fractions of Normal and Nafenopin-Treated Male DBA/2 Mice , Biochem. Pharmacol. 1988, 37, 3897 -3903. 

G. Bellucci, C. Chiappe, F. Marioni, M. Benetti, Regio- and Enantioselectivity of the Cytosolic Epoxide Hydrolase-Catalysed Hydrolysis of Racemic Monosubstituted Alkyloxiranes ,./. Chem. Soc., Perkin Trans. 1 1991, 361 - 363 G. Bellucci, C. Chiappe, L. Conti, F. Marioni, G. Pierini, Substrate Enantioselection in the Microsomal Epoxide Hydrolase Catalyzed Hydrolysis of Monosubstituted Oxiranes. Effects of Branching of Alkyl Chains ,./. Org. Chem. 1989, 54, 5978 - 5983. 

J. Magdalou, B. D. Hammock, 1,2-Epoxycycloalkanes Substrates and Inhibiors of Microsomal and Cytosolic Epoxide Hydrolases in Mouse Liver , Biochem. Pharmacol. 1988, 37, 2717 - 2722. 

G. Bellucci, C. Chiappe, F. Marioni, Enantioselectivity of the Enzymatic Hydrolysis of Cyclohexene Oxide and ( )-l-Methylcyclohexene Oxide A Comparison between Microsomal and Cytosolic Epoxide Hydrolases , J. Chem. Soc., Perkin Trans. 1 1989, 2369 -2373. 

Enzyme catalysed hydrolysis of racemic epoxides is interesting from a practical point of view. This reaction is catalysed by epoxide hydrolases (EHs, EC 3.3.2.3) (Archelas and Furstoss, 1998). Mammalian EHs are the most widely studied and they are divided into five groups among which the soluble (cytosolic) epoxide hydrolases (sEH) and microsomal epoxide hydrolases (mEH) are best charactelised. The mechanism of sEH from rat starts with a nucleophilic attack by Asp333 on a carbon of the epoxide (usually the least hindered one) to form a glycol monoester intermediate which is stabilised by an oxyanion hole. A water molecule activated by His523 releases the 1,2-diol product. An... 

Other types of reduction catalyzed by non-microsomal enzymes have also been described for xenobiotics. Thus, reduction of aldehydes and ketones may be carried out either by alcohol dehydrogenase or NADPH-dependent cytosolic reductases present in the liver. Sulfoxides and sulfides may be reduced by cytosolic enzymes, in the latter case involving glutathione and glutathione reductase. Double bonds in unsaturated compounds and epoxides may also be reduced. Metals, such as pentavalent arsenic, can also be reduced. 

Investigations in vitro have demonstrated that epoxybutene is eliminated by microsomal epoxide hydrolase and by cytosolic glutathione V-transferase (GST). Epoxide hydro-... 

Diepoxybutane, like epoxybutene, is eliminated by microsomal epoxide hydrolase in liver and lung of mouse, rat and man (Boogard Bond, 1996) and by cytosolic GST in liver and lung of mouse and rat and in liver of man (Boogard et al., 1996). 

Bentley, P., Bieri, F., Kuslcr. H., Muakkassah-Kelly, S., Sagelsdorff. P, Staubli, W. Waechter, F. (1989) Hydrolysis of bisphenol A diglycidylether by epoxide hydrolases in cytosolic and microsomal fractions of mouse liver and skin inhibition by bis epoxycyclopentylether and the effects upon the covalent binding to mouse skin DNA. Carcinogenesis, 10, 321-327... 

Conversion of arene oxides to dihydrodiols by mouse liver cytosol epoxide hydrolase and microsomal epoxide hydrolase has been compared, and it is found that the former is less active than the latter.200... 

Phase I reactions include microsomal monooxygenations, cytosolic and mitochondrial oxidations, co-oxidations in the prostaglandin synthetase reaction, reductions, hydrolyses, and epoxide hydration. All of these reactions, with the exception of reductions, introduce polar groups to the molecule that, in most cases, can be conjugated during phase II metabolism. The major phase I reactions are summarized in Table 7.1. 

Hydration of epoxides catalyzed by epoxide hydrolase is involved in both detoxication and intoxication reactions. With high concentrations of styrene oxide as a substrate, the relative activity of hepatic microsomal epoxide hydrolase in several animal species is rhesus monkey > human = guinea pig > rabbit > rat > mouse. With some substrates, such as epoxidized lipids, the cytosolic hydrolase may be much more important than the microsomal enzyme. 

Mukhtar, H., R.M. Philpot, and J.R. Bend. The postnatal development of microsomal epoxide hydrase, cytosolic glutathione S-transferase, and mitochondrial and microsomal cytochrome P-450 in adrenals and ovaries of female rats. Drug Metab. Dispos. 6 577-583, 1978. 

Microsomal epoxide hydrolase is widely distributed, having been described from plants, invertabrates, and vertebrates. In vertebrates it has wide organ distribution for example, in the rat, the most studied species, the enzyme has been found in essentially every organ and tissue. Although predominantly located in the endoplasmic reticulum (microsomes), epoxide hydrolase is also found in the plasma and nuclear membranes and, to some extent, in the cytosolic fraction. 

The existence of a cytosolic epoxide hydrolase was first indicated by its ability to hydrolyze analogs of insect juvenile hormone not readily hydrolyzed by microsomal epoxide hydrolase. Subsequent studies demonstrated a unique cytosolic enzyme catalytically and structurally distinct from the microsomal enzyme. It appears probable that the cytosolic enzyme is peroxisomal in origin. Both enzymes are broadly nonspecific and have many substrates in common. It is clear, however, that many substrates hydrolyzed well by cytosolic epoxide hydrolase are hydrolyzed poorly by microsomal epoxide hydrolase and vice versa. For example, l-(4 -ethylphenoxy)-3,7-dimethy I -6,7-epoxy-//7//i,v-2-octene, a substituted geranyl epoxide insect juvenile hormone mimic, is hydrolyzed 10 times more rapidly by the cytosolic enzyme than by the microsomal one. In any series, such as the substituted styrene oxides, the trans configuration is hydrolyzed more rapidly by the cytosolic epoxide hydrolase than is the cis isomer. At the same time, it should remembered that in this and other series,... 

The assay method described by Eaton and Stapleton (1989), measures the activities of both cytosolic glutathione 5-transferase and microsomal epoxide hydrolase toward benzo[a]pyrene-4,5-oxide as a substrate. These enzymes are important in the biotransformation of many epoxide xenobiotics, including potentially carcinogenic arene oxides. 

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