Stainbrook Sarah C, Yu Jessica S, Reddick Michael P, Bagheri Neda, Tyo Keith E J
Interdisciplinary Biological Sciences Program, Northwestern University, 2145 Sheridan Road, Evanston, IL 60208, USA.
Chemical and Biological Engineering, Northwestern University, 2145 Sheridan Road, Evanston, IL 60208, USA.
Protein Eng Des Sel. 2017 Jun 1;30(6):455-465. doi: 10.1093/protein/gzx018.
The promiscuity of G-protein-coupled receptors (GPCRs) has broad implications in disease, pharmacology and biosensing. Promiscuity is a particularly crucial consideration for protein engineering, where the ability to modulate and model promiscuity is essential for developing desirable proteins. Here, we present methodologies for (i) modifying GPCR promiscuity using directed evolution and (ii) predicting receptor response and identifying important peptide features using quantitative structure-activity relationship models and grouping-exhaustive feature selection. We apply these methodologies to the yeast pheromone receptor Ste2 and its native ligand α-factor. Using directed evolution, we created Ste2 mutants with altered specificity toward a library of α-factor variants. We then used the Vectors of Hydrophobic, Steric, and Electronic properties and partial least squares regression to characterize receptor-ligand interactions, identify important ligand positions and properties, and predict receptor response to novel ligands. Together, directed evolution and computational analysis enable the control and evaluation of GPCR promiscuity. These approaches should be broadly useful for the study and engineering of GPCRs and other protein-small molecule interactions.
G蛋白偶联受体（GPCRs）的多配体 promiscuity 在疾病、药理学和生物传感领域具有广泛影响。对于蛋白质工程而言，多配体 promiscuity 是一个特别关键的考量因素，在该领域中，调节和模拟多配体 promiscuity 的能力对于开发理想的蛋白质至关重要。在此，我们展示了以下方法：（i）利用定向进化来修饰GPCR的多配体 promiscuity，以及（ii）使用定量构效关系模型和分组穷举特征选择来预测受体反应并识别重要的肽特征。我们将这些方法应用于酵母信息素受体Ste2及其天然配体α因子。通过定向进化，我们创建了对α因子变体文库具有改变特异性的Ste2突变体。然后，我们使用疏水、空间和电子性质向量以及偏最小二乘回归来表征受体 - 配体相互作用，识别重要的配体位置和性质，并预测受体对新型配体的反应。总之，定向进化和计算分析能够实现对GPCR多配体 promiscuity 的控制和评估。这些方法对于GPCR以及其他蛋白质 - 小分子相互作用的研究和工程应用应该具有广泛的用途。
