AlphaScreen ® is a technology particularly suitable for bi-molecular inhibitor screening assays, e.g. using protein–protein interactions with purifi ed recombinant proteins. Each binding partner of the bi-molecular interaction is coupled either to donor or to acceptor beads. The technology is based on the quantifi able transfer of oxygen singlets from donor to acceptor microbeads brought together by a specifi c interaction between the partners. We identifi ed the conserved interaction between WW domains of cellular ubiquitin ligases of the Nedd4 family and a short peptide motif (PPxY) present in several structural and nonstructural viral proteins as a potential drug target. Using an AlphaScreen assay recapitulating the interaction between Nedd4.2 and the PPxY motif of the adenoviral capsid protein VI, we screened a library of small molecules and identifi ed specifi c inhibitors of this interaction. © Springer Science+Business Media New York 2016.