A Systematic Dissection Of Sequence Elements Determining Beta-Klotho And Fgf Interaction And Signaling

Endocrine fibroblast growth factors (FGFs) require Klotho transmembrane proteins as necessary coreceptors to activate FGF receptor (FGFR) signaling. In particular, FGF19 and FGF21 function through beta-Klotho to regulate glucose and lipid metabolism. Recent research has focused on elucidating how these two FGFs interact with beta-Klotho and FGFRs to activate downstream signaling. In this study, using hydrogen deuterium exchange coupled to mass spectrometry (HDX-MS), we identified regions on the beta-Klotho protein that likely participate in ligand interaction, and vice versa. Alanine and arginine mutagenesis were carried out to further probe the contributions of individual residues to receptor/ligand interactions. Using biochemical and cell-based signaling assays with full-length proteins, we show that both the KL1 and KL2 domains of beta-Klotho participate in ligand interaction, and these binding sites on beta-Klotho are shared by FGF19 and FGF21. In addition, we show that two highly conserved regions in the C-terminal tail of FGF19 and FGF21 are responsible for interaction with the co-receptor. Our results are consistent with recent publications on the crystal structures of the Klotho proteins and provide insight into how endocrine FGFs interact with co-receptors for signal transduction.