Partial Purification and Characterization of the Porcine Brain Enzyme Hydrolyzing and Synthesizing Anandamide
Year: 1995
Type: article
Abstract: Anandamide (arachidonylethanolamide) is known as an endogenous agonist for cannabinoid receptors. An amidohydrolase, which hydrolyzed anandamide, was solubilized from the microsomal fraction of porcine brain with 1% Triton X-100. The enzyme was partially purified by Phenyl-5PW hydrophobic chromatography to a specific activity of approximately 0.37 mumol/min/mg of protein at 37 degrees C. As assayed with 14C-labeled substrates, the apparent Km value for anandamide was 60 microM, and anandamide was more active than ethanolamides of linoleic, oleic, and palmitic acids. Ceramidase and protease activities were not detected in our enzyme preparation. The purified enzyme also synthesized anandamide from free arachidonic acid in the presence of a high concentration of ethanolamine with a specific activity of about 0.16 mumol/min/mg of protein at 37 degrees C. On the basis of cochromatographies, pH dependence, heat inactivation, and effects of inhibitors such as arachidonyl trifluoromethyl ketone, p-chloromercuribenzoic acid, diisopropyl fluorophosphate, and phenylmethylsulfonyl fluoride, it was suggested that the anandamide amidohydrolase and synthase activities were attributable to a single enzyme protein.
Source: Journal of Biological Chemistry
Authors: Natsuo Ueda, Yuko Kurahashi, Shozo Yamamoto, Takashi Tokunaga
Institutions: Tokushima University, Japan Tobacco (Japan)
Language: English
Subfield: Pharmacology (medical)
Field: Medicine
Domain: Health Sciences
Open Access status: hybrid
Estimated APC paid: $3,430
OpenAlex ID: W2113920734