Structure/Function of the CB2 Receptor Binding and G-protein Recognition Pockets

CB2 受体结合和 G 蛋白识别口袋的结构/功能

• 批准号: 8851758
• 负责人: Xiang-Qun Xie
• 金额: $ 9.6万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8851758
• 关键词: Adenylate Cyclase; Adverse effects; Affinity; Agonist; Algorithms; Anti-Obesity Agents; Appetite Depressants; Autoimmune Diseases; Autoimmune Process; Binding; Binding Sites; Biochemical; Biological Assay; C-terminal; CNR2 gene; Cannabinoids; Cell Signaling Process; Chemicals; Chronic; Complication; Computer Simulation; Coupled; Coupling; Data; Depression and Suicide; Disease; Docking; Drug Design; Drug Targeting; Enzymes; European; Event; Fluorescence; Future; G Protein-Coupled Receptor Genes; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; GTP-Binding Proteins; Glioma; Goals; Guanosine Triphosphate Phosphohydrolases; Health; Human; Immune; Immune system; Isotope Labeling; Length; Ligand Binding; Ligands; Light; Marketing; Mediating; Metabolism; Methods; Molecular; Mutagenesis; Mutation; Neurons; Outcomes Research; Pain; Patients; Pharmaceutical Preparations; Pilot Projects; Proteins; Publishing; Recombinants; Reporting; Research; Research Design; Resolution; Roentgen Rays; Signal Transduction; Site; Specificity; Structure; Structure-Activity Relationship; System; Techniques; Therapeutic; Therapeutic Studies; Transmembrane Domain; Validation; Work; base; biophysical techniques; cannabinoid receptor; design; drug discovery; endogenous cannabinoid system; experience; extracellular; interest; novel; pharmacophore; protein expression; receptor; receptor binding; receptor structure function; research study; rimonabant; three dimensional structure; tumor

DESCRIPTION (provided by applicant): The discovery of the endogenous cannabinoid (CB) system, i.e. the CB receptors (the subtypes CB1 and CB2), endogenous ligands and enzymes for CB ligand metabolism, has triggered intensive pharmacological research into the CB receptors and the therapeutic potential of cannabinergic ligands. The CB2 receptor is known to be involved in the signal transduction cascades in the immune system, and has a great therapeutic potential for developing CB2 drugs without CB1-related psychotropic side effects for treatment of chronic neuro-pain, neuronal disorders, autoimmune diseases, and gliomas and other tumors of immune origin, thereby benefiting the millions of patients who suffer from the autoimmune/immune-related diseases for which we have no cure. Over the years, however, structural and functional studies of CB receptors have focused on predicting structures by computer modeling, identification of the specific sites for ligand binding and G-protein coupling whereas studies on experiment 3D CB structures are limited, in particular for the CB2 receptor. To understand the molecular mechanism behind these pharmacological and biochemical events, it is important to characterize the contacting points and binding domains and then elucidate the specific CB2/ligand recognition and CB2/G-protein coupling mechanisms at molecular structural level. In our published and pilot studies, we have successfully investigated the structural and conformational features of several CB2 protein functional domains; CB ligand structures and active pharmacophoric features; and agonist/antagonist recognition sites in the CB2 receptor. However, many questions still remain about CB2 receptor structure-function relationship as well as CB2 ligands and G-protein recognition mechanisms. The objective of this proposal is to identify/characterize the key residues/functional domains and elucidate their 3D structures of recognition pockets important to agonist and antagonist binding as well as G-protein coupling recognitions in the CB2 receptor by the combined biophysical and biochemical approaches. Our long term goal is to understand, in structural and functional terms, the molecular mechanisms of human CB2 activation and G-protein cell signaling process in order to facilitate the structure-based design for novel CB2 ligands. Having completed the "proof-of-concept" research work, we propose the specific goals and in-depth research to three aims. Aim 1: Characterize the functional domains and key residues important to the CB2 ligand recognition and derive the structural determinants of the agonist/antagonist binding domains in the transmembrane and extra-cellular segments. Aim 2: Investigate and define the structural and functional features of the important CB2 intracellular segments and key residues involving G-protein coupling and intracellular cell signaling by the biophysical approaches developed and validated in Aim 1. Aim 3: Explore full-length CB2 receptor and confirm the key residues determined in Aims 1 and 2 and verify their importance to CB2 ligand recognition and G-protein coupling by functional binding assays and site-directed mutations of the native CB2 receptor. The elucidated CB2 agonist/antagonist recognition pockets and G-protein coupling domains will be further examined by our established computer modeling and receptor docking algorithms. Our proposed research and the outcomes will shed light onto a better understanding of CB2 structure/function and its mechanism of actions, and provide the structural bases for CB2-specific drug design in future. The techniques and methods developed from the proposed CB2 receptor research will also have a significant impact to other GPCRs.

描述（由申请人提供）：内源性大麻素（CB）系统，即CB受体（CB1和CB2亚型）、内源性配体和CB配体代谢酶的发现，引发了对CB受体和CB配体代谢的深入药理学研究。大麻能配体的治疗潜力。已知 CB2 受体参与免疫系统中的信号转导级联，并且具有开发 CB2 药物的巨大治疗潜力，该药物没有 CB1 相关的精神副作用，用于治疗慢性神经疼痛、神经元疾病、自身免疫性疾病和神经胶质瘤和其他免疫来源的肿瘤，从而使数百万患有我们无法治愈的自身免疫/免疫相关疾病的患者受益。然而，多年来，CB 受体的结构和功能研究主要集中在通过计算机建模预测结构、识别配体结合和 G 蛋白偶联的特定位点，而对实验 3D CB 结构的研究却很有限，特别是对于 CB2受体。为了了解这些药理学和生化事件背后的分子机制，重要的是表征接触点和结合域，然后在分子结构水平上阐明特定的 CB2/配体识别和 CB2/G 蛋白偶联机制。在我们已发表的试点研究中，我们成功研究了多个 CB2 蛋白功能域的结构和构象特征； CB配体结构和活性药效特征； CB2 受体中的激动剂/拮抗剂识别位点。然而，关于CB2受体结构-功能关系以及CB2配体和G蛋白识别机制仍然存在许多问题。该提案的目的是通过生物物理和生化相结合的方法来识别/表征关键残基/功能域，并阐明其识别口袋的 3D 结构，这些识别口袋对激动剂和拮抗剂结合以及 CB2 受体中的 G 蛋白偶联识别很重要。我们的长期目标是从结构和功能角度了解人类 CB2 激活和 G 蛋白细胞信号传导过程的分子机制，以促进新型 CB2 配体的基于结构的设计。在完成“概念验证”研究工作后，我们提出了具体目标并针对三个目标进行深入研究。目标 1：表征对 CB2 配体识别重要的功能域和关键残基，并推导跨膜和细胞外片段中激动剂/拮抗剂结合域的结构决定因素。目标 2：通过目标 1 中开发和验证的生物物理方法，研究和定义涉及 G 蛋白偶联和细胞内细胞信号传导的重要 CB2 胞内片段和关键残基的结构和功能特征。 目标 3：探索全长 CB2 受体和确认目标 1 和 2 中确定的关键残基，并通过功能结合测定和天然 CB2 的定点突变验证它们对 CB2 配体识别和 G 蛋白偶联的重要性受体。阐明的 CB2 激动剂/拮抗剂识别口袋和 G 蛋白耦合结构域将通过我们建立的计算机建模和受体对接算法进一步检查。我们提出的研究和结果将有助于更好地理解 CB2 结构/功能及其作用机制，并为未来 CB2 特异性药物设计提供结构基础。拟议的 CB2 受体研究开发的技术和方法也将对其他 GPCR 产生重大影响。

项目成果

Structural insight into tetrameric hTRPV1 from homology modeling, molecular docking, molecular dynamics simulation, virtual screening, and bioassay validations.

• DOI: 10.1021/ci5007189
• 发表时间: 2015-03-23
• 期刊: Journal of chemical information and modeling
• 影响因子: 5.6
• 作者: Feng Z;Pearce LV;Xu X;Yang X;Yang P;Blumberg PM;Xie XQ
• 通讯作者: Xie XQ

Metal binding mediated conformational change of XPA protein:a potential cytotoxic mechanism of nickel in the nucleotide excision repair.

• DOI: 10.1007/s00894-016-3017-x
• 发表时间: 2016-07
• 期刊: Journal of molecular modeling
• 影响因子: 2.2
• 作者: Hu J;Hu Z;Zhang Y;Gou X;Mu Y;Wang L;Xie XQ
• 通讯作者: Xie XQ

Computational target fishing: what should chemogenomics researchers expect for the future of in silico drug design and discovery?

• DOI: 10.4155/fmc.14.5
• 发表时间: 2014-03
• 期刊: Future medicinal chemistry
• 影响因子: 4.2
• 作者: Wang L;Xie XQ
• 通讯作者: Xie XQ

Comparative normal/failing rat myocardium cell membrane chromatographic analysis system for screening specific components that counteract doxorubicin-induced heart failure from Acontium carmichaeli.

• DOI: 10.1021/ac500287e
• 发表时间: 2014-05-20
• 期刊: ANALYTICAL CHEMISTRY
• 影响因子: 7.4
• 作者: Chen, Xiaofei;Cao, Yan;Zhang, Hai;Zhu, Zhenyu;Liu, Min;Liu, Haibin;Ding, Xuan;Hong, Zhanying;Li, Wuhong;Lv, Diya;Wang, Lirong;Zhuo, Xianyi;Zhang, Junping;Xie, Xiang-Qun;Chai, Yifeng
• 通讯作者: Chai, Yifeng

Cardiovascular Disease Chemogenomics Knowledgebase-guided Target Identification and Drug Synergy Mechanism Study of an Herbal Formula.

心血管疾病化学基因组学知识库引导的中药方靶点识别及药物协同机制研究

• DOI: 10.1038/srep33963
• 发表时间: 2016-09-28
• 期刊: Scientific reports
• 影响因子: 4.6
• 作者: Zhang H;Ma S;Feng Z;Wang D;Li C;Cao Y;Chen X;Liu A;Zhu Z;Zhang J;Zhang G;Chai Y;Wang L;Xie XQ
• 通讯作者: Xie XQ