Conformational dynamics has an established role in enzyme catalysis, but its contribution to ligand binding and specificity is largely unexplored. Here we used the Tiam1 PDZ domain and an engineered variant (QM PDZ) with broadened specificity to investigate the role of structure and conformational dynamics in molecular recognition. Crystal structures of the QM PDZ domain both free and bound to ...

Many G protein-coupled receptors possess carboxyl-terminal motifs ideal for interaction with PDZ scaffold proteins, which can control receptor trafficking and signaling in a cell-specific manner. To gain a panoramic view of beta1-adrenergic receptor (beta AR) interactions with PDZ scaffolds, the beta1AR carboxyl terminus was screened against a newly developed proteomic array of PDZ domains. The...

PDZ domain-containing proteins play an important role in the targeting and localization of synaptic membrane proteins. Here, we report an interaction between the PDZ domain-containing protein PICK1 and monoamine neurotransmitter transporters in vitro and in vivo. In dopaminergic neurons, PICK1 colocalizes with the dopamine transporter (DAT) and forms a stable protein complex. Coexpression of PI...

Postsynaptic density 95/discs large/zonus occludens-1 (PDZ) domain-interacting motifs, in addition to their well-established roles in protein scaffolding at the cell surface, are proposed to act as cis-acting determinants directing the molecular sorting of transmembrane cargo from endosomes to the plasma membrane. This hypothesis requires the existence of a specific trans-acting PDZ protein tha...

We report a general method for light-assisted control of interactions of PDZ domain binding motifs with their cognate domains by the incorporation of a photolabile caging group onto the essential C-terminal carboxylate binding determinant of the motif. The strategy was implemented and validated for both simple monovalent and biomimetic divalent ligands, which have recently been established as p...

PDZ domains are ubiquitous protein interaction modules that play a key role in cellular signaling. Their binding specificity involves recognition of the carboxyl-terminus of various proteins, often belonging to receptor and ion channel families. PDZ domains also mediate more complicated molecular networks through PDZ-PDZ interactions, recognition of internal protein sequences or phosphatidylino...

PDZ proteins usually contain multiple protein–protein interaction domains and act as molecular scaffolds that are important for the generation and maintenance of cell polarity and cell signaling. Here, we identify and characterize TIP-1 as an atypical PDZ protein that is composed almost entirely of a single PDZ domain and functions as a negative regulator of PDZ-based scaffolding. We found that...

One of the principal challenges in systems biology is to uncover the networks of protein-protein interactions that underlie most biological processes. To date, experimental efforts directed at this problem have largely produced only qualitative networks that are replete with false positives and false negatives. Here, we describe a domain-centered approach--compatible with genome-wide investigat...

Some membrane proteins must be clustered at target sites to efficiently perform their functions. PDZ domain-containing scaffold proteins bind to the tails of target membrane proteins and promote their localization and clustering on the cell surface. This chapter describes the experimental details of the clustering assay, using the interaction between potassium channels and PSD-95, an abundant P...

The Na1yH1 exchanger regulatory factor (NHERF) binds to the tail of the b2-adrenergic receptor and plays a role in adrenergic regulation of Na1yH1 exchange. NHERF contains two PDZ domains, the first of which is required for its interaction with the b2 receptor. Mutagenesis studies of the b2 receptor tail revealed that the optimal C-terminal motif for binding to the first PDZ domain of NHERF is ...