Streiff John H, Allen Thomas W, Atanasova Elena, Juranic Nenad, Macura Slobodan, Penheiter Alan R, Jones Keith A
Departments of Anesthesiology and Molecular Pharmacology and Experimental Therapeutics and Biochemistry and Molecular Biology, Mayo College of Medicine, Rochester, Minnesota, USA.
Biophys J. 2006 Nov 1;91(9):3405-14. doi: 10.1529/biophysj.106.082586. Epub 2006 Jul 28.
Computational methods designed to predict and visualize ligand protein binding interactions were used to characterize volatile anesthetic (VA) binding sites and unoccupied pockets within the known structures of VAs bound to serum albumin, luciferase, and apoferritin. We found that both the number of protein atoms and methyl hydrogen, which are within approximately 8 A of a potential ligand binding site, are significantly greater in protein pockets where VAs bind. This computational approach was applied to structures of calmodulin (CaM), which have not been determined in complex with a VA. It predicted that VAs bind to Ca(2+)-CaM, but not to apo-CaM, which we confirmed with isothermal titration calorimetry. The VA binding sites predicted for the structures of Ca(2+)-CaM are located in hydrophobic pockets that form when the Ca(2+) binding sites in CaM are saturated. The binding of VAs to these hydrophobic pockets is supported by evidence that halothane predominantly makes contact with aliphatic resonances in Ca(2+)-CaM (nuclear Overhauser effect) and increases the Ca(2+) affinity of CaM (fluorescence spectroscopy). Our computational analysis and experiments indicate that binding of VA to proteins is consistent with the hydrophobic effect and the Meyer-Overton rule.
旨在预测和可视化配体与蛋白质结合相互作用的计算方法，被用于表征与血清白蛋白、荧光素酶和脱铁铁蛋白结合的挥发性麻醉剂（VA）已知结构内的结合位点和未占据口袋。我们发现，在VA结合的蛋白质口袋中，位于潜在配体结合位点约8埃范围内的蛋白质原子和甲基氢的数量显著更多。这种计算方法被应用于尚未确定与VA复合物结构的钙调蛋白（CaM）。它预测VA与Ca(2+)-CaM结合，但不与脱辅基CaM结合，我们通过等温滴定量热法证实了这一点。预测的Ca(2+)-CaM结构的VA结合位点位于CaM中Ca(2+)结合位点饱和时形成的疏水口袋中。VA与这些疏水口袋的结合得到了以下证据的支持：氟烷主要与Ca(2+)-CaM中的脂肪族共振发生接触（核Overhauser效应），并增加了CaM的Ca(2+)亲和力（荧光光谱法）。我们的计算分析和实验表明，VA与蛋白质的结合符合疏水效应和迈耶-欧弗顿规则。
