Publications

• Identification of di-(2-ethylhexyl) phthalate-induced carboxylesterase 1 in C57BL/6 mouse liver microsomes: purification, cDNA cloning, and baculovirus-mediated expression.

  Furihata T., Hosokawa M., Koyano N., Nakamura T., Satoh T., Chiba K.

  Several mouse carboxylesterase (CES) isozymes have been identified, but information about their roles in drug metabolism is limited. In this study, we purified and characterized a mouse CES1 isozyme that was induced by di-(2-ethylhexyl) phthalate. Purified mouse CES1 shared some biological charact ... >> More

  Several mouse carboxylesterase (CES) isozymes have been identified, but information about their roles in drug metabolism is limited. In this study, we purified and characterized a mouse CES1 isozyme that was induced by di-(2-ethylhexyl) phthalate. Purified mouse CES1 shared some biological characteristics with other CES isozymes, such as molecular weight of a subunit and isoelectronic point. In addition, purified mouse CES1 behaved as a trimer, a specific characteristic of CES1A subfamily isozymes. The purified enzyme possessed temocapril hydrolase activity, and it was found to contribute significantly to temocapril hydrolase activity in mouse liver microsomes. To identify the nucleotide sequences coding mouse CES1, antibody screening of a cDNA library was performed. The deduced amino acid sequence of the obtained cDNA, mCES1, exhibited striking similarity to those of CES1A isozymes. When expressed in Sf9 cells, recombinant mCES1 showed hydrolytic activity toward temocapril, as did purified mouse CES1. Based on these results, together with the findings that recombinant mouse CES1 had the same molecular weight of a subunit, the same isoelectronic point, and the same native protein mass as those of purified mouse CES1, it was concluded that mCES1 encoded mouse CES1. Furthermore, tissue expression profiles of mCES1 were found to be very similar to those of the human CES1 isozyme. This finding, together with our other results, suggests that mCES1 shares many biological properties with the human CES1 isozyme. The present study has provided useful information for study of metabolism and disposition of ester-prodrugs as well as ester-drugs. << Less

  Drug Metab. Dispos. 32:1170-1177(2004) [PubMed] [EuropePMC]

• Nonoxidative ethanol metabolism in rabbit myocardium: purification to homogeneity of fatty acyl ethyl ester synthase.

  Mogelson S., Lange L.G.

  Fatty acyl ethyl esters, previously identified in our laboratory as metabolites of ethanol in human and rabbit myocardium, arise from an esterification of free fatty acids with ethanol in the absence of ATP and coenzyme A. This study was designed to isolate and purify the enzyme(s) in rabbit myoca ... >> More

  Fatty acyl ethyl esters, previously identified in our laboratory as metabolites of ethanol in human and rabbit myocardium, arise from an esterification of free fatty acids with ethanol in the absence of ATP and coenzyme A. This study was designed to isolate and purify the enzyme(s) in rabbit myocardium that catalyze(s) this reaction. Enzyme activity in homogenates of rabbit myocardium, as assayed by the rate of synthesis of ethyl [14C]oleate from 0.4 mM [14C]oleic acid and 0.2 M ethanol, was 31 nmol/(g.h), and all of it was recovered in the 48400g supernatant. This soluble ethyl ester synthase activity bound to DEAE-cellulose at pH 8, and elution with a NaCl gradient (0-0.25 M) separated two enzyme activities accounting for 13 and 87% of recovered synthase activity. The major enzyme activity was then purified over 5000-fold to homogeneity by sequential gel permeation, hydrophobic interaction, and anti-albumin affinity chromatographies with an overall yield of 40%. Up to 45 micrograms of enzyme was present per g of myocardium. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed a single polypeptide with Mr 26 000, and gel permeation chromatography under nondenaturing conditions indicated a Mr of 50 000 for the active enzyme. Kinetic analyses using the purified enzyme indicated that greatest rates of ethyl ester synthesis were observed with unsaturated octadecanoic fatty acid substrates [Vmax = 1.9 and 1.5 nmol/(mg.s) for linoleate and oleate, respectively], with lesser rates associated with palmitate, stearate, and arachidonate substrates [0.14, 0.03, and 0.35 nmol/(mg.s), respectively].(ABSTRACT TRUNCATED AT 250 WORDS) << Less

  Biochemistry 23:4075-4081(1984) [PubMed] [EuropePMC]

• Fatty acid ethyl ester synthase in rat adipose tissue and its relationship to carboxylesterase.

  Tsujita T., Okuda H.

  Fatty acid ethyl ester (FAEE) synthase was obtained from rat adipose tissue in an electrophoretically homogeneous form. The enzyme associated with carboxylesterase activity was purified by acetone precipitation followed by successive chromatographies on DEAE-cellulose, phenyl-Sepharose, and Sephad ... >> More

  Fatty acid ethyl ester (FAEE) synthase was obtained from rat adipose tissue in an electrophoretically homogeneous form. The enzyme associated with carboxylesterase activity was purified by acetone precipitation followed by successive chromatographies on DEAE-cellulose, phenyl-Sepharose, and Sephadex G-100 gel. The two activities in rat adipose tissue were associated as judged by their co-elution profiles, co-purifications at different steps, co-precipitations by antibody raised against purified FAEE synthase, and identical profiles of inhibition by diisopropyl fluorophosphate. The enzyme catalyzed the hydrolyses of both tri- and monoacylglycerols, and the susceptibilities of substrates increase with decreasing acyl chain length of the fatty acid moiety. Ethyl oleate-hydrolyzing activity was about one-eighth of the synthesizing activity. The N-terminal amino acid sequence of the first 27 residues of the purified enzyme was identical to that of the carboxylesterase from rat liver. With a polyclonal rabbit antibody against the rat adipose tissue FAEE synthase, the enzyme was demonstrated in the liver, lung, and testis, but not in the kidney. The antibody removed the FAEE-synthesizing activities in adipose tissue (86%), liver (23%), lung (62%), and testis (82%). These results suggest that carboxylesterase contributes to the nonoxidative ethanol metabolism (FAEE synthesis) in various organs. << Less

  J. Biol. Chem. 267:23489-23494(1992) [PubMed] [EuropePMC]