Elucidating the detailed mechanism of activation of membrane protein receptors and their ligand binding is essential for structure-based drug design. Membrane protein crystal structure analysis successfully aids in understanding these fundamental molecular interactions. However, protein crystal structure analysis of the G-protein-coupled receptor (GPCR) remains challenging, even for the class of GPCRs which have been included in the majority of structure analysis reports among membrane proteins, due to the substantial instability of these receptors when extracted from lipid bilayer membranes. It is known that increased thermostability tends to decrease conformational flexibility, which contributes to the generation of diffraction quality crystals. However, this is still not straightforward, and significant effort is required to identify thermostabilized mutants that are optimal for crystallography. To address this issue, a versatile screening platform based on a label-free ligand binding assay combined with transient overexpression in virus-like particles was developed. This platform was used to generate thermostabilized GPR40 [also known as free fatty acid receptor 1 (FFAR1)] for fasiglifam (TAK-875). This demonstrated that the thermostabilized mutant GPR40 (L42A/F88A/G103A/Y202F) was successfully used for crystal structure analysis.

阐明膜蛋白受体及其配体结合的激活的详细机制对于基于结构的药物设计至关重要。膜蛋白晶体结构分析成功地帮助理解了这些基本的分子相互作用。然而，即使对于膜蛋白中大多数结构分析报告中已包括的一类GPCR，G-蛋白偶联受体（GPCR）的蛋白晶体结构分析仍具有挑战性，因为这些受体在使用时非常不稳定。从脂质双层膜中提取。已知增加的热稳定性趋于降低构象柔韧性，这有助于产生衍射品质的晶体。但是，这仍然不是一件容易的事，并且需要大量的努力来鉴定对于晶体学最合适的热稳定突变体。为了解决这个问题，开发了一种基于无标记配体结合测定并结合病毒样颗粒中瞬时过表达的多功能筛选平台。该平台用于为法西格列姆（TAK-875）生成热稳定的GPR40 [也称为游离脂肪酸受体1（FFAR1）]。这表明热稳定的突变体GPR40（L42A / F88A / G103A / Y202F）已成功用于晶体结构分析。该平台用于为法西格列姆（TAK-875）生成热稳定的GPR40 [也称为游离脂肪酸受体1（FFAR1）]。这表明热稳定的突变体GPR40（L42A / F88A / G103A / Y202F）已成功用于晶体结构分析。该平台用于为法西格列姆（TAK-875）生成热稳定的GPR40 [也称为游离脂肪酸受体1（FFAR1）]。这表明热稳定的突变体GPR40（L42A / F88A / G103A / Y202F）已成功用于晶体结构分析。