Moser J, Hye A, Lovell W W, Earl L K, Castell J V, Miranda M A
SEAC Toxicology Unit, Unilever Research, Colworth House, Sharnbrook, Bedford MK44 1LQ, UK.
Toxicol In Vitro. 2001 Aug-Oct;15(4-5):333-7. doi: 10.1016/s0887-2333(01)00033-9.
Photobinding of drugs to biomolecules constitutes an important early event in the onset of photoallergy. In the present work, UV irradiation of human serum albumin in the presence of either suprofen (SUP) or its major photoproduct, decarboxylated suprofen (DSUP), has been studied as a model system for drug-photosensitised protein binding. Both dark binding and binding in the presence of light were investigated since this will affect the mode, site and mechanism of drug interaction with the protein. In order to determine the binding features of SUP to albumin, competitive binding experiments were carried out using fluorescent probes specific for site I and II. Suprofen was found to selectively dark bind to site II on HSA. Photobinding of DSUP to HSA was more efficient than SUP. Parallel to this, the intrinsic tryptophan fluorescence of HSA decreased when the protein was previously irradiated in the presence of the photoactive compounds, again being DSUP more efficient compared with SUP. As fluorescence quenching involves electron transfer from the excited Trp to the ground state DSUP, it follows that the photoactive compound binding to HSA must be on (or in close proximity to) site I Trp(214) residue. It appears that photobinding of SUP is largely preceded by its photodecomposition to DSUP which, in turn, associates and photobinds to HSA.
药物与生物分子的光结合是光过敏反应发生过程中的一个重要早期事件。在本研究中，以在舒洛芬（SUP）或其主要光产物脱羧舒洛芬（DSUP）存在下对人血清白蛋白进行紫外线照射作为药物光敏化蛋白质结合的模型系统进行了研究。对黑暗条件下的结合以及光照条件下的结合均进行了研究，因为这会影响药物与蛋白质相互作用的方式、位点和机制。为了确定SUP与白蛋白的结合特性，使用针对位点I和位点II的荧光探针进行了竞争性结合实验。发现舒洛芬在黑暗中选择性地与HSA上的位点II结合。DSUP与HSA的光结合比SUP更有效。与此平行的是，当蛋白质在光活性化合物存在下预先照射时，HSA的固有色氨酸荧光降低，同样，与SUP相比，DSUP的效率更高。由于荧光猝灭涉及从激发态色氨酸到基态DSUP的电子转移，因此可以得出结论，与HSA结合的光活性化合物必定位于位点I色氨酸（214）残基上（或其附近）。似乎SUP的光结合很大程度上是在其光分解为DSUP之后发生的，而DSUP继而与HSA缔合并发生光结合。
