Interaction oh inhalational anesthetics with macromolecules

描述（改编自申请人申请）：吸入麻醉剂是目前医生使用的毒性最大、了解最少的药物。只有通过详细了解它们的结合相互作用、位点和目标，才能实现改进。在这个子项目中，我们将继续使用等温滴定量热法和酰胺氢交换法，将候选蛋白靶点的结构特征与吸入麻醉剂结合的化学计量学和能量学联系起来（目的1）。每个靶都有结构上的定义，因此，诸如腔数、形状、体积和极性等特征可以与结合能相关联。这些特征将被参数化成一种预测的麻醉剂结合算法，使用现有的高分辨率载铁蛋白和血清白蛋白复合物进行训练，从蛋白质数据库和蛋白质结构计划中挖掘进一步的潜在靶点。在目标2中，目标1中显示特异性结合的蛋白质将与各种吸入麻醉剂共结晶，以获得进一步的结构细节，以纳入结合算法，用于亲和力的相关分析，并探索结合的结构后果。除了载铁蛋白和血清白蛋白外，在最后一个周期中还鉴定了4种蛋白，将在最初进行检查。最后，在目标3中，麻醉药光标记的强大方法将在两个方面进一步完善。首先，结合光标记与晶体学将允许验证揭示的位置，并估计选择性和结合机制。重氮和重氮挥发性化合物将继续发展，重点将放在内合检测方法上。其他4个项目都将存在重要的互动；这些目标将有助于通过揭示亲和力和化学计量学的基础-改变蛋白质构象的能量驱动，并通过揭示新的和意想不到的吸入麻醉蛋白靶标-无论是从现有数据库的挖掘，还是通过光标记鉴定。

DESCRIPTION (adapted from applicant's application): Inhaled anesthetics are the most toxic and least understood drugs that physicians currently use. Improvements can only occur via a detailed understanding of their binding interactions, sites and targets. In this subproject of the program, we will continue to relate structural features of candidate protein targets with the stoichiometry and energetics of inhaled anesthetic binding using both isothermal titration calorimetry and amide hydrogen exchange (aim 1). Each target is structurally defined, so that features, such as cavity number, shape, volume, and polarity can be correlated with binding energetics. These features will be parameterized into a predictive, anesthetic binding algorithm, using the existing high-resolution apoferritin and serum albumin complexes for training, to mine further potential targets from the Protein Data Bank and the Protein Structure Initiative. In aim 2, proteins from aim 1 that show specific binding will be co-crystallized with various inhaled anesthetics to gain further structural detail to be incorporated into the binding algorithm, to use in correlative analyses of affinity, and to probe for the structural consequences of binding. In addition to apoferritin and serum albumin, 4 proteins were identified in the last cycle that will be examined initially. Finally, in aim 3, the powerful method of anesthetic photolabeling will be further refined in two ways. First, combining photolabeling with crystallography will allow validation of revealed location, and an estimate of selectivity and mechanism of incorporation. Continued development of diazirine and diazo volatile compounds will occur, and emphasis will be placed on adduction detection methods. Important interactions will each of the other 4 projects exist; these aims will contribute to the overall program goals by revealing the basis for affinity and stoichiometry- the energetic driver for altering protein conformation, and by revealing new and unanticipated inhaled anesthetic protein targets - either from the mining of existing databases, or through identification via photolabeling.