Microsomal esterases

The intracellular localization of carboxylesterases is predominantly microsomal, the esterases being localized in the endoplasmic reticulum [73] [79] [93], They are either free in the lumen or loosely bound to the inner aspect of the membrane. The carboxylesterases in liver mitochondria are essentially identical to those of the microsomal fraction. In contrast, carboxylesterases of liver lysosomes are different, their isoelectric point being in the acidic range. Carboxylesterase activity is also found in the cytosolic fraction of liver and kidney. It has been suggested that cytosolic carboxylesterases are mere contaminants of the microsomal enzymes, but there is evidence that soluble esterases do not necessarily originate from the endoplasmic reticulum [94], In guinea pig liver, a specific cytosolic esterase has been identified that is capable of hydrolyzing acetylsalicylate and that differs from the microsomal enzyme. Also, microsomal and cytosolic enzymes have different electrophoretic properties [77]. Cytosolic and microsomal esterases in rat small intestinal mucosa are clearly different enzymes, since they hydrolyze rac-oxazepam acetate with opposite enantioselectivity [95], Consequently, studies of hydrolysis in hepatocytes reflect more closely the in vivo hepatic hydrolysis than subcellular fractions, since cytosolic and microsomal esterases can act in parallel. 

Esters, amides, hydrazides, and carbamates can all be metabolized by hydrolysis. The enzymes, which catalyze these hydrolytic reactions, carboxylesterases and amidases, are usually found in the cytosol, but microsomal esterases and amidases have been described and some are also found in the plasma. The various enzymes have different substrate specificities, but carboxylesterases have amidase activity and amidases have esterase activity. The two apparently different activities may therefore be part of the same overall activity. 

Table 9.10 Metabolism of JH analogs by microsomal esterases from CSMA houseflies...

About a decade ago, microsomal esterases were classified into four families (CES1-4) based on their sequence homology [95]. Of these, only CES2 is expressed in the... 

Hydrolysis. Hydrolysis of esters and amides is a common pathway of drug metabolism. The liver microsomes contain non-specific esterases, as do other tissues and plasma. Hydrolysis of an ester results in the formation of an alcohol and an acid hydrolysis of an amide results in the formation of an amine and an acid. The ester procaine, a local anaesthetic, is rapidly hydrolysed by plasma cholinesterases and, to a lesser extent, by hepatic microsomal esterase. An example of an amide which is hydrolysed, is the antiarrhythmic drug procainamide. Enalapril, a prodrug, is hydrolysed by esterases to the active metabolite enalapri-late, which inhibits the angiotensin-converting enzyme. 

Kor a and Ozols (1988) and Ozols (1989) established the primary structures of two microsomal esterases purified from rabbit liver and designated them as 60-kDa esterase form 1 and form 2. These two forms of CarbE have the consensus sequence of the ER retention tetra-peptide (HTEL or HIEL in the one-letter code) which recognized with the luminal side of the KDEL receptor, Robbi el al (1990) reported cDNA cloning of rat liver pi 6.1 esterase (ES-10) and pi 5.5 esterase (ES-3, egasyn). This was the... 

Fenvalerate and other a-cyano pyrethroids, however, are consistently more resistant to oxidative attack than their noncyano analogs. Liver is the predominant site of fenvalerate metabolism via hydrolysis by one or more hepatic microsomal esterases inhibition of these enzymes results in enhanced toxicity. Hydrolysis has also been demonstrated in plasma, kidney, stomach, and brain tissues. Except for brain, however, these tissues were relatively unimportant in the detoxification process. 

The microsomal fraction consists mainly of vesicles (microsomes) derived from the endoplasmic reticulum (smooth and rough). It contains cytochrome P450 and NADPH/cytochrome P450 reductase (collectively the microsomal monooxygenase system), carboxylesterases, A-esterases, epoxide hydrolases, glucuronyl transferases, and other enzymes that metabolize xenobiotics. The 105,000 g supernatant contains soluble enzymes such as glutathione-5-trans-ferases, sulfotransferases, and certain esterases. The 11,000 g supernatant contains all of the types of enzyme listed earlier. 

The metabolism of permethrin will be taken more generally as an example of the metabolism of pyrethroids (Figure 12.2). The two types of primary metabolic attack are by microsomal monooxygenases and esterases. Monooxygenase attack involves... 

Esterases form a wide family of enzymes that catalyze the hydrolysis of ester bonds. They are ubiquitously expressed in all tissues including the intestine, and are found in both microsomal and cytosolic fractions. Prueksaritonont et al. [6] have studied the metabolism of both p-nitrophenol acetate and acetylsalicylic acid by esterases from human intestinal microsomal and cytosolic systems, and the activity values were 2.76 pmol min-1 mg-1 and 0.96 nmol min-1 mg-1, respectively. Thus, the activity for the hydrolysis of p-nitrophenol acetate in human intestine approaches that in the liver. 

Drugs may also undergo hydrolysis by intestinal esterases (hydrolases), more specifically carboxylesterases (EC 3.1.1.1) in the intestinal lumen and at the brush border membrane [58, 59]. It has been shown that intestinal hydrolase activity in humans was closer to that of the rat than the dog or Caco-2 cells [60]. In these studies, six propranolol ester prodrugs and p-nitrophenylacetate were used as substrates, and the hydrolase activity found was ranked in the order human > rat Caco-2 cells > dog for intestinal microsomes. The rank order in hydrolase activity for the intestinal cytosolic fraction was rat > Caco-2 cells = human > dog. The hydrolase activity towards p-nitrophenylacetate and tenofovir disoproxil has also been reported in various intestinal segments from rats, pigs and humans. The enzyme activity in intestinal homogenates was found to be both site-specific (duodenum > jejunum > ileum > colon) and species-dependent (rat > man > Pig)-... 

It was reported that the distribution and activities of esterases that catalyze pyrethroid metabolism using several human and rat tissues, including small intestine, liver, and serum, were examined [30]. The major esterase in human intestine was hCE2. //c/n.v-Permethrin was effectively hydrolyzed by pooled human intestinal microsomes (five individuals), while deltamethrin and bioresmethrin were not. This result correlated well with the substrate specificity of recombinant hCE2. In contrast, pooled rat intestinal microsomes (five animals) hydrolyzed trans-permethrin 4.5 times slower than the human intestinal microsomes. Furthermore, pooled samples of cytosol from human or rat liver were ca. half as hydrolytically active as the corresponding microsome fraction toward pyrethroids however, the cytosolic fractions had significant amounts (ca. 40%) of the total hydrolytic activity. Moreover, a sixfold interindividual variation in hCEl protein expression in human hepatic cytosols was observed. 

Munger JS, Shi GP, Mark EA, Chin DT, Gerard C, Chapman HA (1991) A serine esterase released by human alveolar macrophages is closely related to liver microsomal carboxylesterases. J Biol Chem 266 18832-18838... 

Because the metabolism of DEHP was catalyzed by so many fractions of the trout liver homogenate, these fractions were characterized by measurement of marker enzymes to determine which organelles actually were responsible for the observed DEHP metabolism. Succinic dehydrogenase activity was used as a marker for mitochondria, whereas glucose-6-phosphatase was used as a marker for microsomes. The distribution of DEHP oxidase activity (production of polar metabolites 1 and 2 with added NADPH) and of DEHP esterase activity (production of monoester without added NADPH) were also determined. It was found (Figure 2) that the distribution of DEHP oxidase activity parallels the distribution of microsomal activity and the distribution of DEHP esterase activity parallels the distribution of microsomal activity, but is also present in the cytosol fraction. 

It was also found that paraoxon, an esterase inhibitor, substantially reduced formation of polar metabolite 1 from DEHP by trout liver microsomes with added NADPH. This suggests that polar metabolite 1 is formed via further metabolism of the monoester, the production of which was reduced by paraoxon. 

Fish liver microsomes are capable of both hydrolytic and oxidative metabolism of phthalate esters. In addition, trout liver cytosol and blood serum exhibited esterase activity against DEHP. 

Three isoenzymes of carboxylesterase were purified from rat liver micro-somes and were named RL1, RL2, and RH1. These differ from each other in their response to hormone treatment, inducibility, substrate specificity, and immunological properties [75], It was shown that RL1, RL2, and RH1 resemble hydrolases p/ 6.2/6.4, pI 6.0, and pI 5.6, respectively. Enzyme RL2 was found to be identical to egasyn, a protein with esterase activity found in the endoplasmic reticulum [76], The role of egasyn is to stabilize glucuronidase (EC 3.2.1.31) by noncovalent binding to the microsomal membrane. 

The first esterases to be characterized by sequence were identified by ES numbers [79] [80], Important efforts are now devoted to sequencing, and, as a result, correspondence to the p7 classification has appeared. Thus, microsomal carboxylesterase ES10 is probably identical to p/ 6.1, and esterases ES4 and ESI5 correspond to the p7 5.0 and p7 6.216A enzymes, respectively [74a]. 

In humans, erythrocytes contain an esterase that displays genetic polymorphism [86], This esterase has been called esterase D (ES-D), a name without connection to the above-presented A-, B-, and C-classification. Three carboxylesterases named HU1, HU2, and HU3 have been found in human liver microsomes. Other tissues where esterases have been found include brain, plasma, stomach, small intestine, and colon [79]. 

The physiological functions of carboxylesterases are still partly obscure but these enzymes are probably essential, since their genetic codes have been preserved throughout evolution [84] [96], There is some evidence that microsomal carboxylesterases play an important role in lipid metabolism in the endoplasmic reticulum. Indeed, they are able to hydrolyze acylcamitines, pal-mitoyl-CoA, and mono- and diacylglycerols [74a] [77] [97]. It has been speculated that these hydrolytic activities may facilitate the transfer of fatty acids across the endoplasmic reticulum and/or prevent the accumulation of mem-branolytic natural detergents such as carnitine esters and lysophospholipids. Plasma esterases are possibly also involved in fat absorption. In the rat, an increase in dietary fats was associated with a pronounced increase in the activity of ESI. In the mouse, the infusion of lipids into the duodenum decreased ESI levels in both lymph and serum, whereas an increase in ES2 levels was observed. In the lymph, the levels of ES2 paralleled triglyceride concentrations [92] [98],... 

Thiolester hydrolases are present in most tissues and cell compartments. High concentrations are found in liver microsomes and in brown adipose tissue mitochondria and peroxisomes. Several acyl-CoA hydrolases have shown a close relationship to the nonspecific carboxylesterases EC 3.1.1.1. Thus, palmitoyl-CoA hydrolase purified from rat liver microsomes was found to be identical to esterase pI 6.2I6A (ES4 type). An acyl-CoA hydrolase was isolated that showed high similarity to esterase pI 6.1 [74a] [129] [130]. These few examples are further illustrations of the unsatisfying situation of the traditional classification of esterases. 

Aldehyde dehydrogenase (EC 1.2.1.3) catalyzes the oxidation of aldehydes to acids (see Sect. 3.7.2). The enzyme is ubiquitously distributed, but has mainly been characterized in brain and liver, where it is found in the cytoplasm, mitochondria, and microsomes. It is not clear whether its esterase activity has a physiological role or is a surviving activity inherited from an evolutionary thiolesterase precursor. 

S. Medda, K. Takeuchi, D. Devore-Carter, O. von Diemling, E. Heymann, R. T. Swank, An Accessory Protein Identical to Mouse Egasyn Is Complexed with Rat Microsomal /1-Glucuronidase and Is Identical to Rat Esterase-3 ,. /. Biol. Chem. 1987, 262, 7248-7253. 

K. Hattori, M. Igarashi, M. Itoh, H. Tomisawa, N. Ozawa, M. Tateishi, Purification and Characterizaiton of Glucucorticoid-Inducible Steroid Esterase in Rat Hepatic Microsomes , Biochem. Pharmacol. 1992, 43, 1929-1937. 

Y. S. Huang, L. Woods, L. G. Sultatos, Solubilizaion and Purification of A-Esterase from Mouse Hepatic Microsomes , Biochem. Pharmacol. 1994, 48, 1273-1280. 

The indomethacin-hydrolyzing enzyme from pig liver microsomes was purified and partially characterized [60]. The enzyme was found to be different from known pig liver esterases, since it did not hydrolyze naphth-l-yl-acetate and (4-nitrophenyl)acetate, which are typical substrates for these car-boxy lesterases. The amino acid sequence of the enzyme showed high homology with the mouse carboxylesterase isoenzyme ES-male. Human liver car-... 

S. Hsieh, W. C. Chang, J. D. Huang, Enantioselectivity of Microsomal and Cytosolic Esterases in Rat Intestinal Mucosa , Drug Metab. Dispos. 1992, 20, 719 - 725. 

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