Post-Translational Protein Switches: Novel and Selective Control in Human Cells

Abstract

Controlling enzymes in a monospecific fashion is significant in the study and treatment of metabolic diseases and is a fundamental scientific investigation. There exists a gap in our ability to isolate the function of any one protein within complex signaling networks. We lack adequate tools to specifically probe a single protein functioning in a dynamic network without off-target interactions or crosstalk with other cellular proteins.Here, we present two turn-on switches to interrogate protein signaling pathways. The first switch is a domain insertion turn-on switch that we engineer toward selective interactions. In the process, we highlight discrepancies between observed selectivity in vitro and a lack of correlation in the specificity within cellular environments. The second switch is a first-in-class chemically-induced dimerization complex that we use to reassemble split enzymes. Both switches are built using the protein Bcl-xL. Our last pursuit is to generate a Bcl-xL phage-displayed library that can be used to select for potentially-novel interactions with BH3 peptide binding partners. We hypothesize that there is potential to engineer selective protein switches within human cells based on these technologies.