Refinement of a Recent CFTR Homology Model Guided by Pore Formation

Cystic Fibrosis, which affects nearly 1 in 2,500 births in the Caucasian population, is caused by various mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) protein. CFTR is a member of the ATP-binding-cassette (ABC) protein superfamily. While most ABC proteins are importers or exporters, CFTR is the only one that has evolved to function as an ion channel. Currently no crystal structure of full length CFTR exists, but due to its biological relevance, many CFTR homology models have been proposed. Now, using a recently published and experimentally supported model of CFTR, we have carried out all-atom Molecular Dynamics (MD) in an attempt to further characterize the dynamics of this recalcitrant protein. First, we refined the model utilizing MD flexible fitting (MDFF) and structural data from a 9-A cryo-EM map of CFTR. Next, roughly 1 microsecond of equilibrium MD simulations of the model within a membrane was used to probe three physically relevant states: the apo state, the proposed semi-apo state (1 ATP bound), and the bound state (2 ATP bound). Results show both experimentally demonstrated protein interactions as well as new, untested ones. For instance, an ATP-dependent interaction between the X-loop and Q-loop motifs was observed. A mechanistic picture of how ATP binding leads to channel activity will be reported. Specifically, structural metrics of protein motion are defined and used to explain the formation of a pore within CFTR in simulations when ATP is bound.