Protein folding remains a formidable challenge despite significant advances, particularly in sequence-to-structure prediction. Accurately capturing thermodynamics and intermediates via simulations dem...

The beta-1 adrenergic receptor (ADRB1) is a critical target for cardiovascular drugs, yet our understanding of how it is activated remains incomplete. Capturing the concerted interplay of agonists, solvent, ions, and protein microswitches is a significant challenge for conventional simulation methods and is essential for unraveling this process. Here, we address this challenge by implementing a powerful enhanced sampling framework that integrates the OneOPES enhanced sampling algorithm with a set of biologically motivated collective variables (CVs). These CVs are designed to track several key features of the activation process simultaneously, including rearrangement of conserved microswitches, the state of the sodium ion binding pocket, and dynamics of critical water molecules. Using this framework, we mapped the multidimensional free energy landscapes of the ADRB1 receptor in both its apo- and adrenaline-bound holo states. Our analysis reveals a detailed, stepwise activation pathway that quantifies the known modulatory roles of sodium ions and protonation states and identifies essential water-mediated networks that stabilize the active conformation. This work provides a detailed overview of ADRB1 activation and establishes the robustness of our OneOPES approach for investigating complex activation mechanisms with the potential for application to other Class A GPCRs.