Accessing Transient Binding Pockets by Protein Engineering and Yeast Surface Display Screening

Abstract

The binding pocket of some therapeutic targets can acquire multiple conformations that, to some extent, depend on the protein dynamics and the interaction with other molecules. The inability to reach the binding pocket can impose a substantial or even insurmountable barrier for the de novo identification or optimization of small-molecule ligands. Herein, we describe a protocol for the engineering of a target protein and a yeast display FACS sorting strategy to identify protein variants with a stable transient binding pocket with improved binding for a cryptic site-specific ligand. This strategy may facilitate drug discovery using the resulting protein variants with accessible binding pockets for ligand screening.

Keywords: Cell cytometry; Protein engineering; Transient binding pockets; Yeast surface display.