The study of phospholipase A2 and inhibitors interaction

• Main Authors: Yi-Hsun Tsao, 曹藝薰
• Other Authors: Wei-Ning Huang
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/35r64s

碩士 === 元培科技大學 === 生物技術研究所 === 99 === Phospholipase A2 (PLA2, E.C. 3.1.1.4) converts phospholipids into free fatty acids and lysophospholipids by hydrolyzing the ester bond of phospholipids at sn-2 position. Physiologically, PLA2 activation in immune cells will pass the message to initiate the inflammation mechanism. In this study, we used NBD fluorescent agent and liposome coated phenol red as indicators to compare inhibitory mechanims of various inhibitors to different PLA2s. The two indicators showed that the inhibitor MJ33 strongly suppressed the activites of bee venom phospholipase A2 (bvPLA2) while the inhibitor PK03 strongly suppressed the activites of and human nonpancreatic phospholipase A2 (hnpPLA2); however, the two methods showed contrary results when MJ33 and PK03 were applied to Naja atra phospholipase A2 (aPLA2). As this was the first time to utilize liposome coated phenol red as the indicator for studying PLA2 activity assay, more studies would be needed to characterize properties of this method. We also used X-ray diffractions and the co-crystals of MJ33 and aPLA2 for the three-dimensional structural determination. We found that calcium ion was far away from MJ33 and the ability of inhibition of MJ33 might not be enhanced by calcium ion. Based on computational results on docking, the fatty-acid-like tail of PK03 would enhance hydrophobic interactions with PLA2. However, the profile of lowest binding energy of MJ33 and aPLA2 showed the carbon chain of MJ33 dock into the active site of aPLA2 and it was not matching the result of X-ray crystallgraphy. It may provide more possibility of interaction on PLA2/inhibitor.