Formation of nNOS/PSD-95 PDZ dimer requires a preformed β-finger structure from the nNOS PDZ domain

Abstract

PDZ domains are modular protein units that play important roles in organizing signal transduction complexes. PDZ domains mediate interactions with both C-terminal peptide ligands and other PDZ domains. Here, we used PDZ domains from neuronal nitric oxide synthase (nNOS) and postsynaptic density protein-95 (PSD-95) to explore the mechanism for PDZ-dimer formation. The nNOS PDZ domain terminates with a ∼30 residue amino acid β-finger peptide that is shown to be required for nNOS/PSD-95 PDZ dimer formation. In addition, formation of the PDZ dimer requires this β-finger peptide to be physically anchored to the main body of the canonical nNOS PDZ domain. A buried salt bridge between the β-finger and the PDZ domain induces and stabilizes the β-hairpin structure of the nNOS PDZ domain. In apo-nNOS, the β-finger peptide is partially flexible and adopts a transient β-strand like structure that is stabilized upon PDZ dimer formation. The flexibility of the NOS PDZ β-finger is likely to play a critical role in supporting the formation of nNOS/PSD-95 complex. The experimental data also suggest that nNOS PDZ and the second PDZ domain of PSD-95 form a “head-to-tail” dimer similar to the nNOS/syntrophin complex characterized by X-ray crystallography.