Abstract
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.
Footnotes
-
This work was supported in part by a grant from the Ministry of Education, Sciences, Sports and Culture of Japan (14572090).
-
doi:10.1124/dmd.104.000620.
-
ABBREVIATIONS: CES, carboxylesterase; CES D1, dog carboxylesterase D1; CES ML1, mouse carboxylesterase ML1; CES ML2, mouse carboxylesterase ML2; CES RL1, rat carboxylesterase RL1; CES P1, porcine carboxylesterase P1; DEHP, di-(2-ethylhexyl) phthalate; Ms, microsomes; PAGE, polyacrylamide gel electrophoresis; PNPA, p-nitrophenylacetate; PNPester, p-nitrophenylester; PNPB, p-nitrophenylbutylate; PNPP, p-nitrophenylpropionate; RT-PCR, reverse transcription-polymerase chain reaction; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; mTGH, mouse triacylglycerol hydrolase; PP, peroxisome proliferator; PPARα, peroxisome proliferator-activated receptor.
- Received May 14, 2004.
- Accepted July 16, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
DMD articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|