The tachykinin 2 receptor (NK2R) plays critical roles in gastrointestinal, respiratory and mental disorders and is a well-recognized target for therapeutic intervention. To date, therapeutics targeting NK2R have failed to meet regulatory agency approval due in large part to the limited characterization of the receptor-ligand interaction and downstream signaling. Herein, we report a protein engineering strategy to improve ligand-binding- and signaling-competent human NK2R that enables a yeast-based NK2R signaling platform by creating chimeras utilizing sequences from rat NK2R. We demonstrate that NK2R chimeras incorporating the rat NK2R C-terminus exhibited improved ligand-binding yields and downstream signaling in engineered yeast strains and mammalian cells, where observed yields were better than 4-fold over wild type. This work builds on our previous studies that suggest exchanging the C-termini of related and well-expressed family members may be a general protein engineering strategy to overcome limitations to ligand-binding and signaling-competent G protein-coupled receptor yields in yeast. We expect these efforts to result in NK2R drug candidates with better characterized signaling properties.

中文翻译：

---

使用具有大鼠NK2R C末端的嵌合体，可提高酵母中配体结合和信号传导能力的人类NK2R产量，从而使NK2R-G蛋白信号传导平台发挥作用。

速激肽2受体（NK2R）在胃肠道，呼吸道和精神疾病中起关键作用，并且是公认的治疗干预目标。迄今为止，靶向NK2R的治疗药物未能获得监管机构的批准，这在很大程度上是由于受体-配体相互作用和下游信号传导的有限表征。在本文中，我们报告了一种蛋白质工程策略，旨在通过利用来自大鼠NK2R的序列创建嵌合体来改善配体结合和信号传递能力的人类NK2R，从而使基于酵母的NK2R信号传递平台成为可能。我们证明，掺入大鼠NK2R C端的NK2R嵌合体在工程酵母菌株和哺乳动物细胞中表现出提高的配体结合产量和下游信号传导，其中观察到的产量比野生型高4倍。这项工作建立在我们以前的研究的基础上，该研究表明，交换相关的和表达良好的家族成员的C末端可能是克服酵母中配体结合和信号传导G蛋白偶联受体产量限制的通用蛋白质工程策略。我们期望这些努力将导致具有更好表征的信号特性的NK2R候选药物。