Conformational thermostabilization of the β1-adrenergic receptor in a detergent-resistant form
Conformational thermostabilization of the β1-adrenergic receptor in a detergent-resistant form
复制标题
DOI：
10.1073/pnas.0711253105
复制
发表时间：
2008-01-22
影响因子：
11.1
通讯作者：
Tate, Christopher G.
中科院分区：
文献类型：
--
作者：
Serrano-Vega, Maria J.;Magnani, Francesca;Tate, Christopher G.
人类基因组编码的无气味G蛋白偶联受体(GPCRs)大约有350个，其中许多被预测为潜在的治疗靶点，但只有两个结构可代表整个家族。我们假设，提高这些受体的洗涤剂稳定性，并同时将它们锁定在一种首选构象中，将大大提高结晶的机会。我们开发了一种通用策略，用于分离β1-肾上腺素能受体GPCR的洗涤剂增溶耐热突变体。最稳定的突变体受体βAR-M23包含6个点突变，导致明显的T-m比天然蛋白高21摄氏度，在结合拮抗剂存在的情况下，βAR-M23与牛视紫质一样稳定。此外，βAR-M23在结晶理想的多种洗涤剂中明显更稳定，并且在没有配体的情况下优先以拮抗剂构象存在。
There are approximate to 350 non-odorant G protein-coupled receptors (GPCRs) encoded by the human genome, many of which are predicted to be potential therapeutic targets, but there are only two structures available to represent the whole of the family. We hypothesized that improving the detergent stability of these receptors and simultaneously locking them into one preferred conformation will greatly improve the chances of crystallization. We developed a generic strategy for the isolation of detergent-solubilized thermostable mutants of a GPCR, the beta 1-adrenergic receptor. The most stable mutant receptor, beta AR-m23, contained six point mutations that led to an apparent T-m 21 degrees C higher than the native protein, and, in the presence of bound antagonist, beta AR-m23 was as stable as bovine rhodopsin. In addition, beta AR-m23 was significantly more stable in a wide range of detergents ideal for crystallization and was preferentially in an antagonist conformation in the absence of ligand.