| Molecular interactions drive all processes in life. They determine the molecular crosstalk and build the basic language of biological processes. By developing a combined approach of atomic force microscopy and single-molecule force spectroscopy (SMFS) we image native membrane proteins and locate their molecular interactions at submolecular resolution. The approach observes how molecular interactions guide a membrane protein along unfolding pathways, how they fold a polypeptide into the functional protein, and how they stabilize structural intermediates of the protein. We show examples of how mutations, temperature, electrolyte and pH change the interactions of and between secondary structure elements and guide the membrane protein along different unfolding and folding pathways. However SMFS can also be applied to measure protein-protein interactions, interactions switching on and off ion channels, ligand- or inhibitor-binding, the functional states of receptors, and the supramolecular assembly of molecular machines as functional units. Dynamic SMFS (DFS) obtains insights into the mechanical rigidity, transition state, lifetime, and free energy stabilizing the structural regions of a membrane protein. Using DFS we reveal mechanistic insights how molecular interactions modulates these energetic parameters to precisely tune the folding and function of a membrane protein. |
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