In vivo conformational space and defects of misfolded CFTR variants by covalent protein painting.

In vivo characterization of protein structures and structural changes after perturbation is still a major challenge and has impacted our understanding of the molecular events involved in protein misfolding diseases. To identify the true conformational space occupied by proteins in their native state in vivo, we recently developed a structural proteomics method named Covalent Protein Painting (CPP). Here, we show how CPP can be used to identify and quantify the conformational defects of proteins in the misfolding disease Cystic Fibrosis. We first report the discovery of a previously unreported opening mechanism for the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) as well as its conformational changes during biogenesis. Then we further reveal how misfolding of different CFTR variants in Cystic Fibrosis disturbs these conformational changes even upon treatment with current approved drugs and suggest possibilities to stabilize misfolded CFTR variants not or less responsive to these drugs such as N1303K CFTR.