Biosynthesis of anandamide and N-palmitoylethanolamine by sequential actions of phospholipase A2 and lysophospholipase D.
Anandamide (an endocannabinoid) and other bioactive long-chain NAEs (N-acylethanolamines) are formed by direct release from N-acyl-PE (N-acyl-phosphatidylethanolamine) by a PLD (phospholipase D). However, the possible presence of a two-step pathway from N-acyl-PE has also been suggested previously, which comprises (1) the hydrolysis of N-acyl-PE to N-acyl-lysoPE by PLA1/PLA2 enzyme(s) and (2) the release of NAEs from N-acyllysoPE by lysoPLD (lysophospholipase D) enzyme(s). In the present study we report for the first time the characterization of enzymes responsible for this pathway. The PLA1/PLA2 activity for N-palmitoyl-PE was found in various rat tissues, with the highest activity in the stomach. This stomach enzyme was identified as group IB sPLA2 (secretory PLA2), and its product was determined as N-acyl-1-acyl-lysoPE. Recombinant group IB, IIA and V of sPLA2s were also active with N-palmitoyl-PE, whereas group X sPLA2 and cytosolic PLA2a were inactive. In addition, we found wide distribution of lysoPLD activity generating N-palmitoylethanolamine from N-palmitoyl-lysoPE in rat tissues, with higher activities in the brain and testis. Based on several lines of enzymological evidence, the lysoPLD enzyme could be distinct from the known N-acyl-PE-hydrolysing PLD. sPLA2-IB dose dependently enhanced the production of N-palmitoylethanolamine from N-palmitoyl-PE in the brain homogenate showing the lysoPLD activity. N-Arachidonoyl-PE and N-arachidonoyl-lysoPE as anandamide precursors were also good substrates of sPLA2-IB and the lysoPLD respectively. These results suggest that the sequential actions of PLA2 and lysoPLD may constitute another biosynthetic pathway for NAEs, including anandamide.
Sun
Yong-Xin
YX
Department of Biochemistry, Kagawa University School of Medicine, 1750-1 Ikenobe, Miki, Kagawa 761-0793, Japan.
Tsuboi
Kazuhito
K
Okamoto
Yasuo
Y
Tonai
Takeharu
T
Murakami
Makoto
M
Kudo
Ichiro
I
Ueda
Natsuo
N
D000577:Amides; D000818:Animals; D001095:Arachidonic Acids / Q000096:biosynthesis*; D001921:Brain / Q000201:enzymology; D002460:Cell Line; D063388:Endocannabinoids; D004983:Ethanolamines / Q000378:metabolism; D054500:Group IB Phospholipases A2; D006801:Humans; D006868:Hydrolysis; D007527:Isoenzymes / Q000378:metabolism; D007668:Kidney / Q000166:cytology / Q000196:embryology; D008297:Male; D009928:Organ Specificity; D010169:Palmitic Acids / Q000378:metabolism*; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010741:Phospholipases A / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D054466:Phospholipases A1; D054467:Phospholipases A2; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D053284:Polyunsaturated Alkamides; D051381:Rats; D017208:Rats, Wistar; D013270:Stomach / Q000201:enzymology; D013379:Substrate Specificity; D000099189:Lysophospholipase D
D000577:Amides; D001095:Arachidonic Acids; D063388:Endocannabinoids; D004983:Ethanolamines; D054500:Group IB Phospholipases A2; D007527:Isoenzymes; D010169:Palmitic Acids; D010714:Phosphatidylethanolamines; D010741:Phospholipases A; D054466:Phospholipases A1; D054467:Phospholipases A2; D010727:Phosphoric Diester Hydrolases; D053284:Polyunsaturated Alkamides; D000099189:Lysophospholipase D; C022203:N-acylethanolamines; C483858:phosphatidylethanolamine; C005958:palmidrol; C516202:PLA2G1B protein, human; C516204:Pla2g1b protein, rat; C078814:anandamide