| Most eukaryotic proteins that receive and process signals are constructed from a combination of interaction and catalytic domains. Among the many interaction domains identified in the past decade, PDZ domains are one of the most frequently encountered. They are often found in combination with other interaction modules and play a role in directing the specificity of receptor tyrosine kinase-mediated signaling, in establishing cell polarity, in directing protein trafficking, and in coordinating synaptic signaling. The enormous diversity of PDZ domain function is manifest in their abundance; there are over 240 such domains encoded in the mouse genome. To understand their individual roles, it is necessary first to define their recognition properties in a comprehensive fashion.;First, we describe an effort to provide a genome-wide perspective on how PDZ domains recognize C-terminal peptides. Previous efforts to define PDZ domain binding selectivity have been based on experimental data that are specific to the domains being studied and so are not applicable to other members of the domain family. Here, we report the construction of a three-dimensional extension of a position-specific scoring matrix that predicts to which peptides a PDZ domain will bind, given only the primary sequences of the PDZ domain and the peptides. The model, which was trained using interaction data from 82 PDZ domains and 93 peptides encoded in the mouse genome, successfully predicts interactions involving other mouse PDZ domains, as well as PDZ domains from Drosophila melanogaster and Caenorhabditis elegans. The model is also able to predict the effects of mutations in peptide ligands on their PDZ domain-binding affinities. Overall, we show that our approach is a viable way to capture, in a single model, the binding selectivity of the PDZ domain family.;We then report a family-wide investigation of a less well-characterized mechanism of binding: PDZ domain dimerization. To date, fewer than fifteen PDZ domain dimers have been identified and the frequency of this binding mode remains unclear. With the goal of determining whether PDZ domain dimerization is a commonly used mode of PDZ domain-mediated protein-protein interaction, we devised a strategy to examine all possible PDZ-PDZ interactions among a proteome-sized set of PDZ domains. Our approach combines the throughput of protein microarrays and the fidelity of fluorescence polarization with biological validation as a method of discovering PDZ-PDZ interactions. In our unbiased study, we identified 33 novel PDZ-PDZ interactions revealing that PDZ-PDZ dimerization is not a frequently encountered mode of PDZ domain-mediated interaction. Nevertheless, this study uncovered numerous previously unrecognized protein-protein interactions and serves as a launching point for future, more in-depth investigations of their biological relevance. |
