REQUIMTE, Departamento de Química, Faculdade de Farmácia, Universidade do Porto, Rua Aníbal Cunha, Porto, Portugal.
Chem Phys Lipids. 2011 May;164(4):292-9. doi: 10.1016/j.chemphyslip.2011.03.003. Epub 2011 Mar 22.
Phospholipase A(2) (PLA(2)) lipolytic activity can be regarded as a limiting factor for the development of inflammatory processes by restricting the production of pro-inflammatory mediators, hence representing a valuable therapeutic target for drugs that are able to modulate the activity of this enzyme. In the current work, the hydrolysis of phospholipids by PLA(2) was monitored with acrylodan-labelled intestinal fatty acid binding protein (ADIFAB) and this fluorescence based technique was also used to access the enzymatic inhibitory effect of non-steroidal anti-inflammatory drugs (NSAIDs). The intrinsic fluorescence of PLA(2) tryptophan residues was further used to gain complementary information regarding the accessibility of these residues on the PLA(2) structure upon interaction with the NSAIDs tested; and to calculate the NSAIDs-PLA(2) binding constants. Finally, circular dichroism (CD) measurements were performed to evaluate changes in PLA(2) conformation resultant from the inhibitory effect of the drugs tested. Overall, results gathered in this study point to the conclusion that the studied NSAIDs inhibit PLA(2) activity due to a disturbance of the enzyme binding efficiency to membrane interface possibly by a shielding effect of the Trp residues required for the membrane interfacial binding step that precedes lipolysis process.
磷脂酶 A2（PLA2）的脂解活性可以通过限制促炎介质的产生，被视为炎症过程发展的限制因素，因此成为能够调节这种酶活性的药物的有价值的治疗靶点。在目前的工作中，通过丙烯酰基标记的肠脂肪酸结合蛋白（ADIFAB）监测 PLA2 对磷脂的水解，并且还使用这种基于荧光的技术来评估非甾体抗炎药物（NSAIDs）的酶抑制作用。进一步使用 PLA2 色氨酸残基的固有荧光来获得关于这些残基在与所测试的 NSAIDs 相互作用时在 PLA2 结构上的可及性的补充信息；并计算 NSAIDs-PLA2 结合常数。最后，进行圆二色性（CD）测量以评估药物测试的抑制作用导致的 PLA2 构象变化。总的来说，本研究的结果表明，所研究的 NSAIDs 抑制 PLA2 活性是由于酶与膜界面的结合效率受到干扰，可能是由于脂解过程之前的膜界面结合步骤所需的色氨酸残基的屏蔽效应。
