The glucagon-like peptide 1 receptor (GLP-1R) is a class B G-protein coupled receptor (GPCR) and diabetes drug target expressed mainly in pancreatic β-cells that, when activated by its agonist glucagon-like peptide 1 (GLP-1) after a meal, stimulates insulin secretion and β-cell survival and proliferation. The N-terminal region of GLP-1 interacts with membrane-proximal residues of GLP-1R, stabilizing its active conformation to trigger intracellular signaling. The best-studied agonist peptides, GLP-1 and exendin-4, share sequence homology at their N-terminal region; however, modifications that can be tolerated here are not fully understood. In this work, a functional screen of GLP-1 variants with randomized N-terminal domains reveals new GLP-1R agonists and uncovers a pattern whereby a negative charge is preferred at the third position in various sequence contexts. We further tested this sequence–structure–activity principle by synthesizing peptide analogues where this position was mutated to both canonical and noncanonical amino acids. We discovered a highly active GLP-1 analogue in which the native glutamate residue three positions from the N-terminus was replaced with the sulfo-containing amino acid cysteic acid (GLP-1-CYA). The receptor binding and downstream signaling properties elicited by GLP-1-CYA were similar to the wild type GLP-1 peptide. Computational modeling identified a likely mode of interaction of the negatively charged side chain in GLP-1-CYA with an arginine on GLP-1R. This work highlights a strategy of combinatorial peptide screening coupled with chemical exploration that could be used to generate novel agonists for other receptors with peptide ligands.

中文翻译：

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使用饱和诱变和化学设计鉴定 N 端多样化的 GLP-1R 激动剂

胰高血糖素样肽 1 受体 (GLP-1R) 是 B 类 G 蛋白偶联受体 (GPCR) 和糖尿病药物靶标，主要在胰腺 β 细胞中表达，当被其激动剂胰高血糖素样肽 1 (GLP- 1）饭后，刺激胰岛素分泌和β细胞的存活和增殖。GLP-1 的 N 末端区域与 GLP-1R 的膜近端残基相互作用，稳定其活性构象以触发细胞内信号传导。研究得最好的激动剂肽 GLP-1 和 exendin-4 在它们的 N 端区域具有序列同源性；但是，这里可以容忍的修改尚未完全理解。在这项工作中，具有随机 N 末端结构域的 GLP-1 变体的功能筛选揭示了新的 GLP-1R 激动剂，并揭示了一种模式，即在各种序列环境中，负电荷优先位于第三位。我们通过合成肽类似物进一步测试了这种序列-结构-活性原理，其中该位置突变为规范和非规范氨基酸。我们发现了一种高活性 GLP-1 类似物，其中 N 末端三个位置的天然谷氨酸残基被含磺基的氨基酸半胱氨酸 (GLP-1-CYA) 取代。GLP-1-CYA 引发的受体结合和下游信号传导特性与野生型 GLP-1 肽相似。计算模型确定了 GLP-1-CYA 中带负电荷的侧链与 GLP-1R 上的精氨酸相互作用的可能模式。