Low-affinity analytical chromatography for measuring inhaled anesthetic binding to isolated proteins.
Low-affinity analytical chromatography for measuring inhaled anesthetic binding to isolated proteins.
复制标题
用于测量吸入麻醉剂与分离蛋白质的结合的低亲和力分析色谱。
DOI：
10.1006/abio.2001.5506
复制
发表时间：
2002
影响因子：
2.9
通讯作者：
Eckenhoff,RodericG
中科院分区：
文献类型：
--
作者：
Chan,Kin;Meng,QingC;Johansson,JonasS;Eckenhoff,RodericG
挥发性麻醉剂与蛋白质结合的直接测量由于弱亲和力和因此的快速动力学而变得复杂。因此，这些临床重要药物的几个假定靶点只有功能数据来支持直接作用模式。虽然有几种方法可以测量结合的某些方面，但都有很大的局限性。我们介绍了使用分析色谱法的目的，直接测量挥发性麻醉剂结合蛋白质，并表明，它可以提供估计的亲和力和化学计量的蛋白质，可以获得相当高的纯度和质量。使用这种方法，我们的特点是氟烷结合血清白蛋白的低亲和力和多位点，和肌红蛋白或细胞色素C的严格非特异性。这种方法将是有用的，在直接表征平衡，溶液结合分离的蛋白质在准备更耗时的方法与结构分辨率。
The direct measure of volatile anesthetic binding to protein is complicated by weak affinity and therefore rapid kinetics. Consequently, several puted targets for these clinically important drugs have only functional data to support a direct mode of action. While several methods for measuring some aspects of binding are available, all have significant limitations. We introduce the use of analytical chromatography for the purpose of directly measuring volatile anesthetic binding to protein, and show that it can provide estimates of both affinity and stoichiometry for proteins that can be obtained in fairly high purity and mass. Using this approach we characterize halothane binding to serum albumin as low affinity and multisite, and to myoglobin or cytochrome C as strictly nonspecific. This approach will be useful in directly characterizing equilibrium, solution binding to isolated proteins in preparation for more time-consuming methods with structural resolution.