Hydrolysis of 2-arachidonoylglycerol in Tetrahymena thermophila. Identification and partial characterization of a Monoacylglycerol Lipase-like enzyme.
Tetrahymena thermophila is a model organism for molecular and cellular biology. Previous studies from our group showed that Tetrahymena contains major components of the endocannabinoid system, such as various endocannabinoids and FAAH. In mammalian cells the endocannabinoid 2-arachidonoylglycerol is inactivated mainly by MAGL. In this study we showed that 2-arachidonoylglycerol and 2-oleoylglycerol are hydrolyzed by the combined actions of MAGL and FAAH. MAGL-like activity was examined in the presence of FAAH specific inhibitors, URB597 or AM374 and showed optimum pH of 8-9, apparent K(M) of 14.1μM and V(max) of 5.8nmol/min×mg. The enzyme was present in membrane bound and cytosolic isoforms; molecular mass was determined at ∼45 and ∼40kDa. MAGL and FAAH could also inactivate endogenous signaling lipids, which might play an important role in Tetrahymena as suggested in mammals. Tetrahymena could be used as a model system for testing drugs targeting enzymes of the endocannabinoid system.
Evagorou
Andri
A
National and Kapodistrian University of Athens, Department of Chemistry (Biochemistry), University of Athens, Panepistimioupolis, 15771 Athens, Greece.
Anagnostopoulos
Dimitrios
D
Farmaki
Elena
E
Siafaka-Kapadai
Athanasia
A
D000581:Amidohydrolases / Q000037:antagonists & inhibitors; D001095:Arachidonic Acids / Q000378:metabolism*; D001549:Benzamides / Q000494:pharmacology; D002219:Carbamates / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D063388:Endocannabinoids; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005989:Glycerides / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008970:Molecular Weight; D008994:Monoacylglycerol Lipases / Q000737:chemistry / Q000378:metabolism*; D010168:Palmitates / Q000494:pharmacology; D015800:Protozoan Proteins / Q000737:chemistry / Q000378:metabolism*; D016808:Tetrahymena thermophila / Q000378:metabolism*; D000099252:Fatty Acid Amide Hydrolases
D000581:Amidohydrolases; D001095:Arachidonic Acids; D001549:Benzamides; D002219:Carbamates; D063388:Endocannabinoids; D004791:Enzyme Inhibitors; D005989:Glycerides; D008994:Monoacylglycerol Lipases; D010168:Palmitates; D015800:Protozoan Proteins; D000099252:Fatty Acid Amide Hydrolases; C104963:AM 374; C500528:cyclohexyl carbamic acid 3'-carbamoylbiphenyl-3-yl ester; C094503:glyceryl 2-arachidonate; C505247:2-oleoylglycerol