UC San Diego

Diversity-generating retroelements (DGRs) are genetic modules with the ability to introduce massive sequence variation into specific target proteins. Such DGR variable proteins are able to specifically recognize and bind ligands, and their properties have several potential applications in molecular surface display technologies. The first step in such an endeavor is to show that the DGR variable proteins are compatible with molecular surface display systems. The M13 filamentous phage display system was tested for surface-display of DGR variable proteins. However, only truncated versions of the DGR variable proteins were detected in isolated phage. Further investigation strongly suggested that periplasmic proteolysis of the DGR variable proteins occurred during the phage maturation process. Next, ribosome display of DGR variable proteins was tested. Intact DGR variable proteins were detected by polyacrylamide gel electrophoresis and western blot after in vitro transcription and translation. Affinity selection of a ribosome-displayed DGR variable protein was demonstrated, which verified the presence of accompanying mRNA. This work demonstrates the incompatibility of several DGR variable proteins for M13 phage display, but also demonstrates that ribosome display is a viable alternative for surface display of DGR variable proteins.