Evaluation of indole-based probes for high-throughput screening of drug binding to human serum albumin: Analysis by high-performance affinity chromatography
Evaluation of indole-based probes for high-throughput screening of drug binding to human serum albumin: Analysis by high-performance affinity chromatography
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DOI：
10.1002/jssc.200800567
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发表时间：
2009-04-01
影响因子：
3.1
通讯作者：
Hage, David S.
中科院分区：
文献类型：
--
作者：
Conrad, Mandi L.;Moser, Annette C.;Hage, David S.
近年来，人们越来越关注使用快速和选择性分离方法，如高效亲和色谱（HPAC）或亲和毛细管电泳（ACE）来表征药物-蛋白质相互作用。L-色氨酸在这些方法和其他方法中通常用作位点选择性探针，用于检查小溶质和药物在蛋白质HSA上的Sudlow位点II的结合。然而，L-色氨酸溶液可能不稳定，通常每天新鲜制备用于这些研究。在这份报告中，HPAC被用来检查其他吲哚化合物作为可能的替代L-色氨酸作为一个位点选择性探针，用于高通量筛选药物结合到HSA的影响，这些结果在使用这些化合物在ACE中也被考虑。测试的探针候选物包括吲哚-3-乙酸、吲哚-3-羧酸、吲哚-3-丁酸、吲哚-3-丙酸、吲哚-3-甲醇、3-乙酰基吲哚和3-甲基吲哚。所有这些化合物被发现通过H-1 NMR和UV-Vis光谱是稳定的提示3周在室温下，当保持在pH 7.4，0.067 M磷酸盐缓冲液。通过使用含有固定化HSA的柱来检查这些化合物的结合。3-发现乙酰吲哚是该组中用作Sudlow位点II的L-色氨酸的替代探针的最佳候选物。该探针与L-色氨酸具有相同的结合位点和相似的亲和力，但在水溶液中更稳定，使其适合于在HPAC和ACE中进行药物-HSA结合的高通量筛选。
There has been growing interest in the use of rapid and selective separation methods such as high-performance affinity chromatography (HPAC) or affinity capillary electrophoresis (ACE) for the characterization of drug-protein interactions. L-Tryptophan is commonly used in these and other methods as a site-selective probe for examining the binding of small solutes and drugs at Sudlow site II on the protein HSA. However, solutions of L-tryptophan can be unstable and are generally prepared fresh daily for these studies. In this report, HPAC was used to examine other indole compounds as possible replacements for L-tryptophan as a site-selective probe for use in the high-throughput screening of drug binding to HSA; the implications OF these results in the Use Of such compounds in ACE were also considered. The probe candidates that were tested included indole-3-acetic acid, indole-3-carboxylic acid, indole-3-butyric acid, indole-3-propionic acid, indole-3-methanol, 3-acetylindole, and 3-methylindole. All of these compounds were found by H-1 NMR and UV-Vis spectroscopy to be stable for tip to 3 wk at room temperature when kept in a pH 7.4, 0.067 M phosphate buffer. The binding of these compounds was examined by using columns that contained immobilized HSA. 3-Acetylindole was found to be the best candidate in this group for use as in alternative probe to L-tryptophan for Sudlow site II. This probe had the same binding site and a similar affinity to L-tryptophan but Was More stable in aqueous solution, making it suitable for high-throughput screening of drug-HSA binding in both HPAC and ACE.