Provasi Davide, Bortolato Andrea, Filizola Marta
Department of Structural and Chemical Biology, Mount Sinai School of Medicine, New York, New York 10029, USA.
Biochemistry. 2009 Oct 27;48(42):10020-9. doi: 10.1021/bi901494n.
Opioid receptors are G protein-coupled receptors (GPCRs) of utmost significance in the development of potent analgesic drugs for the treatment of severe pain. An accurate evaluation at the molecular level of the ligand binding pathways into these receptors may play a key role in the design of new molecules with more desirable properties and reduced side effects. The recent characterization of high-resolution X-ray crystal structures of non-rhodopsin GPCRs for diffusible hormones and neurotransmitters presents an unprecedented opportunity to build improved homology models of opioid receptors, and to study in more detail their molecular mechanisms of ligand recognition. In this study, possible pathways for entry of the nonselective antagonist naloxone (NLX) from the water environment into the well-accepted alkaloid binding pocket of a delta opioid receptor (DOR) molecular model based on the beta2-adrenergic receptor crystal structure are explored using microsecond-scale well-tempered metadynamics simulations. Using as collective variables distances that account for the position of NLX and of the receptor extracellular loop 2 in relation to the DOR binding pocket, we were able to distinguish between the different states visited by the ligand (i.e., docked, undocked, and metastable bound intermediates) and to predict a free energy of binding close to experimental values after correcting for possible drawbacks of the sampling approach. The strategy employed herein holds promise for its application to the docking of diverse ligands to the opioid receptors as well as to other GPCRs.
阿片受体是G蛋白偶联受体（GPCRs），在开发用于治疗重度疼痛的强效镇痛药方面具有极其重要的意义。在分子水平上准确评估配体进入这些受体的结合途径，可能在设计具有更理想特性和更少副作用的新分子中发挥关键作用。最近对用于可扩散激素和神经递质的非视紫红质GPCRs的高分辨率X射线晶体结构的表征，为构建改进的阿片受体同源模型以及更详细地研究其配体识别分子机制提供了前所未有的机会。在本研究中，使用微秒级的温和元动力学模拟，探索了非选择性拮抗剂纳洛酮（NLX）从水环境进入基于β2肾上腺素能受体晶体结构的δ阿片受体（DOR）分子模型中广为人知的生物碱结合口袋的可能途径。使用作为集体变量的距离来描述NLX和受体细胞外环2相对于DOR结合口袋的位置，我们能够区分配体所经历的不同状态（即对接、未对接和亚稳结合中间体），并在校正采样方法可能存在的缺点后，预测出接近实验值的结合自由能。本文采用的策略有望应用于将各种配体对接至阿片受体以及其他GPCRs。
