The glucagon-like peptide-1 receptor is a drug target for type 2 diabetes and obesity.  Activation of this receptor in the pancreas stimulates insulin release, leading to its glucose regulating properties. However, the receptor is widely expressed in other tissues and its roles there are less well understood. Identifying how GLP-1R signals in different cells and tissues can help understand side effects and new therapeutic benefits of GLP-1R activation beyond glucoregulatory properties. GLP-1-R is a class B GPCR that is endogenously activated by the 30 residue peptide GLP-1(7-36).  We have recently reported¹ using a combinatorial alanine scanning approach of the hydroxyl containing residues (Thr11, Ser14, Ser17, Ser18) in GLP-1(7-36) enabling us to generate new GLP-1R agonists with significant signalling bias.

Mutating any single residue to alanine removes one hydroxyl group reducing receptor affinity and cAMP ten-fold, with Ala11 or Ala14 also reducing β-arrestin-2 ten-fold while Ala17 or Ala18 also increases ERK1/2 phosphorylation five-fold. Multiple alanine mutations more profoundly bias signalling, differentially silencing or restoring one or more signaling properties. Mutating three serines silences only ERK1/2, the first example of such bias. Mutating all four residues silences β-arrestin-2, ERK1/2 and Ca²⁺, maintains ligand and receptor at the membrane, but still potently stimulates cAMP and insulin secretion in cells and mice. Utilizing recently published cryo-EM structures of GLP-R bound to agonist ligands, we show that hydrogen bonding from hydroxyl side chains cooperatively controls cell signalling. These  GLP-1R receptor agonists with signalling bias provide new tools for the study of the effects of signalling pathways in roles beyond glucose regulation.