Defining the conformational landscape of glycogen synthase kinase 3 beta (GSK3β)

Glycogen synthase kinase 3 beta (GSK3β) is a serine/threonine kinase implicated in Alzheimer's disease, type 2 diabetes, oncology and bipolar disorder, and represents an extensively pursued therapeutic target. Despite the substantial accumulation of GSK3β crystal structures in the Protein Data Bank (PDB), no systematic consolidation and conformational analysis of this dataset have been performed. Here, we manually classified all deposited GSK3β crystal structures, comprising 111 unique PDB entries and 88 distinct co-crystallized ligands (data extracted March 11th, 2026). Using the pre-existing KinCoRe dataset and standardized nomenclature, we assigned conformational states across the complete GSK3β structural ensemble and evaluated their relationships with bound ligands. All structures adopt the DFG-in kinase conformation but populate eight distinct conformational substates defined by combinations of activation-loop geometry, αC-helix orientation, regulatory salt-bridge status, αC-β5-loop and phosphorylation state. Our structural analysis reveals that chemically distinct Type I inhibitors stabilize distinct conformational substates, including variation in activation-loop arrangement and activity-like states. These relationships have not previously been characterized across the complete GSK3β crystal structure ensemble. This work provides the first comprehensive conformational landscape of resolved GSK3β crystal structures, a curated structural reference to guide the selection of potentially biologically relevant conformations for structure-based drug design and a systematic framework for analogous kinase-wide conformational analyses.