CFTR Protein Interactome Mapping and Off-Target Identification of Pharmacological Corrector by Mass Spectrometry

Abstract

Here we aimed to investigate the CF corrector VX-445 using affinity purification-mass spectrometry and chemical biology tools. To provide a better understanding of the underlying molecular mechanism of VX-445, protein-protein interactions were mapped. Chapter 2 defines the remodeling of the CFTR interactomics profile upon VX-445 treatment and provides a connection between corrector response and differential translational dynamics. Ribosome structural proteins were found to mediate VX-445 response in a previously minimally responsive variant of CFTR. The relationship between protein translation and corrector response is further investigated later in Chapter 5. In Chapter 3, we investigate off-targets of VX-445 to highlight the utility of understanding the full spectrum of drug binding partners. We report an uncharacterized protein, SCCPDH as an off-target of VX-445. Methods to determine drug binding site on proteins by photocrosslinking are further discussed in Chapter 5. In Chapter 4, we then apply an improved interactomics profiling method to another rare disease, deficiency of adenosine deaminase 2 (DADA2) caused by loss-of-function mutations in ADA2. We show a detailed interactome profile of several disease-causing variants and shed light on disease mechanism in the perspective of protein processing. Lastly, Chapter 5 additionally covers the determination of protein processing steps through time-resolved interactomics profiling (TRIP). The findings in this dissertation help us better understand protein misfolding diseases beyond CF, spurring deeper investigations to resolve remaining challenges for the betterment of inflicted people.