Computer Modeling of G Protein-Coupled Receptors

G 蛋白偶联受体的计算机建模

• 批准号: 8939540
• 负责人: Kenneth Alan Jacobson
• 金额: $ 32.49万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8939540
• 关键词: ADORA3 gene; Adenosine; Adenosine A1 Receptor; Adenosine A2A Receptor; Adenosine A2B Receptor; Affinity; Agonist; Binding; Binding Proteins; Binding Sites; Biochemical; Biological Assay; CXCR4 gene; Characteristics; Charge; Clinical; Complex; Computational Technique; Computer Simulation; Data; Databases; Development; Docking; Drug Design; Elements; Family; Fibrinolytic Agents; G-Protein-Coupled Receptors; Homologous Gene; Homology Modeling; Human; Inflammation; Lead; Ligand Binding; Ligands; Light; Maps; Measures; Mediating; Membrane; Modeling; Molecular; Molecular Conformation; Mutagenesis; Nucleosides; Nucleotides; Performance; Pharmaceutical Preparations; Pharmacology; Platelet aggregation; Property; Protein Databases; Purinergic P1 Receptors; Purines; Pyrimidines; Receptor Activation; Reporting; Research; Resolution; Retrieval; Roentgen Rays; Sequence Alignment; Series; Shapes; Signal Transduction; Structure; Structure-Activity Relationship; Techniques; Testing; Triazoles; X-Ray Crystallography; analog; base; chemokine receptor; design; extracellular; farnesyl pyrophosphate; improved; inorganic phosphate; insight; molecular dynamics; molecular recognition; novel; nucleotide receptor; purine; purinoceptor P2Y1; radioligand; receptor; receptor binding; scaffold; screening; small molecule; small molecule libraries; three dimensional structure; three-dimensional modeling; tool; usability; virtual

Structure-based drug design strategies have been used to elucidate specific ligand recognition determinants and ultimately lead to the design of new small molecules for a specific target. The structure of the target receptor protein is the first requirement in structure-based drug design approaches. In the absence of a high resolution structure of a given G protein-coupled receptor (GPCR), computational techniques like homology modeling can be used to build a 3D model. Briefly, the best structural template is chosen from the Protein Database (PDB) mainly considering sequence identity and similarity with the target receptor and the quality of the crystallographic structure (e.g. resolution). The sequence of the target receptor is aligned to the template structure using highly conserved residues and the known shared structural features to guide the automated or semi-automated alignment. The sequence alignment and the template structures are the input for the homology modeling. Energy minimization or molecular dynamics can be used to further refine and optimize the resulting 3D models. The ligand-receptor interactions can be identified by means of structure-based approaches, e.g. molecular docking. The information contained in the ligand-receptor complexes from the docking can clarify structural elements for molecular recognition and lead to a further optimization of the compounds and the design of new derivatives. The binding site of a given GPCR can be mapped for each class of small molecule ligands. Also, virtual searching of chemically diverse databases for novel chemotypes to bind to a given GPCR structure has been productive using both structure-based and ligand-based strategies. We have applied mutagenesis and homology modeling to the study of GPCR families for extracellular purines and pyrimidines and used the structural insights gained to assist in the design of novel ligands. These families consist of the adenosine receptors (ARs) and the P2Y (nucleotide) receptors. The structures of the human A2AAR were recently reported in the antagonist-bound state and in the agonist-bound state. These structures can reliably serve as modeling templates, with some adjustment, for other ARs due to the relatively high sequence identity between ARs (average 47% between human subtypes). We achieved the structure-function analysis of P2YRs by indirect means, using mutagenesis and homology modeling based on a template of the high-resolution structure of similar GPCRs, such as the CXCR4 chemokine receptor. We have collaborated with one of the premier centers for X-ray crystallography of membrane-bound proteins, i.e. the lab of Prof. Ray Stevens (Scripps Research Inst.), to report the first X-ray structure of an agonist-bound A2AAR. Automatic docking of known potent nucleosides to the agonist-bound A2AAR crystallographic structure, and to homology models of other subtypes, was performed, resulting in new predictions of stabilizing interactions and a structural basis for previous empirical structure activity relationships. We predicted binding of novel C2 terminal and 5' derivatives of adenosine and used the models to interpret effects on measured binding affinity and efficacy of newly-synthesized agonists. Structures of G protein-coupled receptors (GPCRs) have a proven utility in the discovery of new antagonists and inverse agonists, which may modulate signaling of this important family of clinical targets. However, applicability of active-state GPCR structures to virtual screening and rational optimization of agonists, remains to be assessed. We have studied adenosine 5′ derivatives and evaluated the performance of an agonist-bound A2A adenosine receptor (AR) structure in retrieval of known agonists, and then employed the structure to screenfor new fragments optimally fitting the corresponding subpocket. Biochemical and functional assays demonstrate high affinity of new derivatives that include polar heterocycles. The binding models also explain a modest selectivity gain for some substituents toward the closely related A1AR subtype and the modified agonist efficacy of some of these ligands. The study suggested further applicability of in silico fragment screening to rational lead optimization for GPCRs in general. In order to investigate the usability of homology models and the inherent selectivity of a particular model in relation to close homologs, we constructed multiple homology models for the A1 adenosine receptor (A1AR) and docked, 2.2 M lead-like compounds. High-ranking molecules were tested on the A1AR as well as the close homologs A2AAR and A3AR. While the screen yielded numerous potent and novel ligands (hit rate 21% and highest affinity of 400 nM), it delivered few selective compounds. Moreover, most compounds appeared in the top ranks of only one model. The structure− activity relationship (SAR) for a novel class of 1,2,4- triazole antagonists of the human A2A adenosine receptor (hA2A AR) was explored. Thirty-three analogs of a ligand that was discovered in a structure-based virtual screen against the hA2A AR were tested in AR radioligand binding assays and in functional assays for the A2B AR subtype. As a series of closely related analogs of the initial lead did not display improved binding affinity or selectivity, molecular docking was used to guide the selection of more distantly related molecules. This resulted in the discovery of novel AR antagonists (Ki ≥200 nM) with high ligand efficiency. In light of the SAR for the 1,2,4-triazole scaffold, we also investigated the binding mode of these compounds based on docking to several A2AAR crystal structures The P2Y1 receptor (P2Y1R) is a G protein-coupled receptor naturally activated by extracellular ADP. Its stimulation is an essential requirement of ADP-induced platelet aggregation, thus making antagonists highly sought compounds for the development of antithrombotic agents. Here, through a virtual screening campaign based on a pharmacophoric representation of the common characteristics of known P2Y1R ligands and the putative shape and size of the receptor binding pocket, we have identified novel antagonist hits of microM affinity derived from a N,N-bis-arylurea chemotype. Unlike the vast majority of known P2Y1R antagonists, these drug-like compounds do not have a nucleotidic scaffold or highly negatively charged phosphate groups. Hence, our compounds may provide a direction for the development of receptor probes with altered physicochemical properties. The P2Y12R is an ADP-activated GPCR that is well validated clinically as an important target for antithrombotic drugs. Our new P2Y12R structure is the most predictive of the P2Y14R, a target for treating inflammation. Ligand docking to the P2Y12R-based model of the P2Y14R predicted poses of both reversibly-binding small molecules, consistent with observed pharmacology. We are using this approach to discover novel ligands of this receptor. Farnesyl pyrophosphate was recently identified as an insurmountable antagonist of ADP-induced platelet aggregation mediated by the P2Y12R. Docking of farnesyl pyrophosphate in a P2Y12R model revealed molecular similarities with ADP and a good fit into the binding pocket for ADP.

基于结构的药物设计策略已用于阐明特定的配体识别决定因素，并最终导致针对特定靶标的新小分子设计。目标受体蛋白的结构是基于结构的药物设计方法中的第一个要求。在没有给定G蛋白偶联受体（GPCR）的高分辨率结构的情况下，可以使用同源模型之类的计算技术来构建3D模型。简而言之，最佳结构模板是从蛋白质数据库（PDB）中选择的，主要考虑序列身份和与靶受体的相似性以及晶体学结构的质量（例如分辨率）。使用高度保守的残基和已知的共享结构特征将目标受体的序列与模板结构对齐，以指导自动化或半自动对齐。序列比对和模板结构是同源性建模的输入。能量最小化或分子动力学可用于进一步完善和优化所得的3D模型。可以通过基于结构的方法，例如分子对接。来自对接的配体受体复合物中包含的信息可以阐明分子识别的结构元素，并导致化合物的进一步优化和新衍生物的设计。可以为每个类别的小分子配体映射给定GPCR的结合位点。同样，使用基于结构的基于结构和基于配体的策略，对新型化学型的化学数据库进行虚拟搜索以与给定的GPCR结构结合的新型化学型与给定的GPCR结构结合。 我们已将诱变和同源性建模应用于GPCR家族的细胞外嘌呤和嘧啶家族的研究，并使用了获得的结构见解来协助设计新型配体的设计。这些家族由腺苷受体（ARS）和P2Y（核苷酸）受体组成。最近在拮抗剂结合的状态和激动剂结合的状态下报道了人类A2AAR的结构。由于ARS之间的序列身份相对较高（人类亚型之间的平均47％），这些结构可以可靠地充当其他AR的建模模板，并进行一些调整。我们使用基于诱变和同源性建模基于基于类似GPCR的高分辨率结构（例如CXCR4趋化因子受体的高分辨率结构）的诱变和同源性建模来实现P2YRS的结构 - 功能分析。 我们已经与膜结合蛋白的X射线晶体学的主要中心之一，即Ray Stevens教授（Scripps Research Inst。），以报告激动剂结合的A2AAR的第一个X射线结构。进行了已知有效的核苷的自动对接，以与激动剂结合的A2AAR晶体结构以及其他亚型的同源模型进行自动对接，从而产生了稳定相互作用的新预测和以前的经验结构活动关系的结构基础。我们预测了新型C2末端和腺苷的5'衍生物的结合，并使用模型来解释对新近合成激动剂的结合亲密关系和功效的影响。 G蛋白偶联受体（GPCR）的结构在发现新的拮抗剂和反激动剂的发现方面具有证实的效用，这可能会调节这一重要的临床靶标家族的信号传导。但是，活跃状态GPCR结构的适用性在虚拟筛查和对激动剂的合理优化中的适用性仍有待评估。我们已经研究了腺苷5'衍生物，并评估了已知的激动剂的腺苷结合A2a腺苷受体（AR）结构的性能，然后采用该结构来筛选新的片段，以最佳地拟合相应的子货。生化和功能分析表明，包括极性杂环在内的新衍生物的高亲和力。结合模型还解释了某些取代基对紧密相关的A1AR亚型的适度选择性增益以及其中某些配体的修饰激动剂功效。该研究表明，在硅片段筛选中进一步适用于GPCR的合理铅优化。 为了研究同源模型的可用性以及与关闭同源物相关的特定模型的固有选择性，我们为A1腺苷受体（A1AR）（A1AR）构建了多个同源性模型，并停靠了2.2 M铅样化合物。在A1AR以及近距离同源物A2AAR和A3AR上测试了高级分子。尽管屏幕产生了许多有效和新颖的配体（命中率为21％，最高亲和力为400 nm），但几乎没有选择性化合物。此外，大多数化合物仅出现在一个模型的最高排名中。 新型1,2,4-的结构 - 活性关系（SAR） 探索了人A2a腺苷受体（HA2A AR）的三唑拮抗剂。在基于结构的虚拟屏幕针对HA2A AR中发现的配体的三十三个类似物在AR放射性结合测定中测试了A2B AR亚型的功能分析。由于初始铅的一系列紧密相关的类似物没有显示出改善的结合亲和力或选择性，因此使用分子对接来指导选择更遥远相关的分子。这导致了具有高配体效率的新型AR拮抗剂（Ki≥200nm）的发现。鉴于1,2,4-三唑支架的SAR，我们还根据对接与几个A2AAR晶体结构的结合模式研究了这些化合物的结合模式 P2Y1受体（P2Y1R）是G蛋白偶联受体自然通过细胞外ADP激活的受体。它的刺激是ADP诱导的血小板聚集的必不可少的要求，因此使拮抗剂高度寻求抗血栓形成剂的化合物。在这里，通过基于已知P2Y1R配体的共同特征的药物表达的虚拟筛选运动以及受体结合口袋的假定形状和大小，我们已经确定了源自N，N，N-BIS-芳族芳烃化学型的新型Microm亲和力。与绝大多数已知的P2Y1R拮抗剂不同，这些类似药物的化合物没有核苷酸支架或高度负电荷的磷酸基团。因此，我们的化合物可能为开发具有改变理化特性的受体探针的发展提供了方向。 P2Y12R是ADP激活的GPCR，在临床上被很好地证明是抗血栓药物的重要靶标。我们的新P2Y12R结构是P2Y14R的最预测性，这是治疗炎症的靶标。 P2Y14R基于P2Y12R的模型的配体对接两种可逆的小分子的姿势，与观察到的药理学一致。我们正在使用这种方法来发现该受体的新型配体。 Farnesyl焦磷酸最近被确定为由P2Y12R介导的ADP诱导的血小板聚集的替代拮抗剂。 P2Y12R模型中Farnesyl焦磷酸的对接显示出与ADP的分子相似性，并且适合ADP的结合口袋。