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Ligand binding inhibitors of A1 adenosine receptor from Rana rugosa are phospholipase A2s
- Baek, Hwa Jin, Kwon, So Yeon, Kim, Sunkyu, Kim, Sukwon S., Oh, Utaek, Hwang, Seongbin, Chang, Hyeun Wook, Lee, Byeong Jae
- European journal of biochemistry 2000 v.267 no.5 pp. 1340-1346
- Glandirana rugosa, adenosine, capsaicin, cholinergic receptors, frogs, matrix-assisted laser desorption-ionization mass spectrometry, molecular weight, phospholipase A2, reversed-phase high performance liquid chromatography, sequence homology, snakes, venoms
- Inhibitors of the A1 adenosine receptor were isolated from the skin extract of Korean frog, Rana rugosa. The frogâskin extract was prepared by an electrical shock and fractionated with C4 followed by C18 reverseâphase HPLC. Two A1 receptor inhibitors were isolated using a filter binding assay and the molecular masses of the proteins were estimated by matrixâassisted laser desorption ionization timeâofâflight mass spectrometry to be 15â347 and 15â404âDa, respectively. The inhibitory activity was also measured against other membrane receptors, such as the A2 adenosine receptor, muscarinic acetylcholine receptor and capsaicin receptor. Ligand binding to the A2 and muscarinic receptors was also severely inhibited by these proteins. However, they did not inhibit the functional activation of the capsaicin receptor by its ligand, capsaicin, suggesting that inhibition of ligandâreceptor binding occurs specifically. Their Nâterminal sequences were determined by Edman degradation. Surprisingly, they showed sequence similarity to the secretory protein, phospholipase A2 from various organisms. The phospholipase A2 activity of both proteins was tested using Dole's assay technique. Both proteins showed phospholipase A2 activity, and therefore, they were designated as PLA2âR1 and PLA2âR2, respectively. In addition, their ligandâbinding inhibitory activity depended on their phospholipase A2 activity. This is the first finding that the frog secretes a phospholipase A2 similar to that of snake venoms, which posess inhibitory activity against the adenosine A1, adenosine A2 and muscarinic receptors.