The promise of structural biology had been to reason about biological function using the three dimensional arrangement of atoms. Over the last decades this reasoning included more and information about the dynamic movement of the structures, prominently by using molecular dynamics simulation.

Recently, cryo-EM dramatically extended the library of biomolecules structures, because it is able to capture the same biomolecule in lots of different states during their function. It is unique among structural biology techniques for its ability to produce three-dimensional structure of biomolecules in many functional sub-states instead of one structure that represents an ensemble average. The challenge remains to reconstruction how molecules move between these different states and what their relative occupations are, because these structures neither represent an average in thermodynamic equilibrium nor precise “snapshot” of an individual conformation.

Molecular dynamics simulations feature a very similar behaviour, by providing local ensemble estimates in the vicinity of a “starting structure”.

Here, I’ll show, how both techniques may be combined to move from cryo-EM as “ensemble of maps” technique to a true ensemble description that entails free energy differences between cryo-EM maps and minimum free energy paths between them.