PDF(Pubmed)
Abstract：
Positive allosteric modulators (PAMs) of the cannabinoid CB1 receptor (CB1) offer potential therapeutic advantages in the treatment of neuropathic pain and addiction by avoiding the adverse effects associated with orthosteric CB1 activation. Here, molecular modeling and mutagenesis were used to identify residues central to PAM activity at CB1. Six putative allosteric binding sites were identified in silico, including novel sites previously associated with cholesterol binding, and key residues within each site were mutated to alanine. The recently determined ZCZ011 binding site was found to be essential for allosteric agonism, as GAT228, GAT229 and ZCZ011 all increased wild-type G protein dissociation in the absence of an orthosteric ligand; activity that was abolished in mutants F191A3.27 and I169A2.56. PAM activity was demonstrated for ZCZ011 in the presence of the orthosteric ligand CP55940, which was only abolished in I169A2.56. In contrast, the PAM activity of GAT229 was reduced for mutants R220A3.56, L404A8.50, F191A3.27 and I169A2.56. This indicates that allosteric modulation may represent the net effect of binding at multiple sites, and that allosteric agonism is likely to be mediated via the ZCZ011 site. This study underlines the need for detailed understanding of ligand receptor interactions in the search for pure CB1 allosteric modulators.
摘要：
大麻素CB1受体（CB1）的正变构调节剂（PAM）通过避免与正构CB1激活相关的不良反应，在治疗神经性疼痛和成瘾方面提供了潜在的治疗优势。这里,使用分子建模和诱变来鉴定对CB1处的PAM活性至关重要的残基。在计算机模拟中鉴定了六个推定的变构结合位点，包括以前与胆固醇结合相关的新位点，并且每个位点内的关键残基突变为丙氨酸。最近确定的ZCZ011结合位点被发现是必不可少的变构激动，由于GAT228，GAT229和ZCZ011在不存在正位配体的情况下都增加了野生型G蛋白的解离；突变体F191A3.27和I169A2.56中的活性被消除。在存在正构配体CP55940的情况下，ZCZ011证明了PAM活性，该活性仅在I169A2.56中被消除。相比之下,对于突变体R220A3.56、L404A8.50、F191A3.27和I169A2.56,GAT229的PAM活性降低。这表明变构调制可能代表多个位点结合的净效应，这种变构激动作用很可能是通过ZCZ011位点介导的。这项研究强调了在寻找纯CB1变构调节剂时需要详细了解配体受体相互作用。
