A conformational free energy landscape of the biological macromolecule adenylyl-3’,5’-adenylyl-3’,5’-adenosine is presented. The aim was to explore the stacking-unstacking process of the trimer. The free energy landscape was created from two-dimensional potential of mean force calculations based on 324 molecular dynamics simulations in aqueous solution. A large number of conformational states and substrates with various transition barriers were observed. In the free energy landscape we can follow the changes of the stacking to unstacking process for the trimer. The lowest minimum in the free energy landscape was observed for the trimer having all three bases stacked. Local minima were found for the trimer having two bases stacked and one unstacked.