Screening libraries to identify proteins with desired binding activities using a split-GFP reassembly assay

Designer protein modules, which bind specifically to a desired target, have numerous potential applications. One approach to creating such proteins is to construct and screen libraries. Here we present a detailed description of using a split-GFP reassembly assay to screen libraries and identify proteins with novel binding properties. Attractive features of the split-GFP based screen are the absence of false positives and the simplicity, robustness, and ease of automation of the screen. Here, we describe both the construction of a näive protein library, and screening of the library using the split-GFP assay to identify proteins that bind specifically to chosen peptide sequences.