Selection of ligands by panning of domain libraries displayed on phage lambda reveals new potential partners of synaptojanin 1

One of the goals of functional genomics is the description of reliable and complete protein interaction networks. To facilitate ligand discovery from complex protein mixtures, we have developed an improved approach that is af fected by a negligible fraction of false positives. We have combined a nove l technique based on the display of cDNA libraries on the capsid of bacteri ophage lambda and an efficient plaque assay to reveal phage displaying liga nds that are enriched after only a couple of affinity purification steps. W e show that the lambda display system has a unique ability to display, at h igh density, proteins ranging in size from a few to at least 300 amino acid residues. This characteristic permits attenuation of the size bias in the selection procedure and, at the same time, offers a sensitive plaque assay that permits us to do away with the ligand background without unduly increa sing the number of selection cycles. By using a proline-rich fragment of th e synaptojanin 1 protein as a bait, we have identified, in a brain cDNA dis play Library, seven ligands all containing either SH3 or WW domains. Four o f these correspond to proteins that have already been validated as physiolo gical partners, while the remaining three are new partners, whose physiolog ical relevance remains to be established. Two different proline-rich region s of the p21-activated protein kinase 1 (Pak1) and WAVE/SCAR2! protein retr ieve from the library different proteins containing SH3 or WW domains. (C) 2001 Academic Press.