Post-Translational Modifications Modulate Ligand Recognition by the Third PDZ Domain of the MAGUK Protein PSD-95 Murciano Calles, Javier Corbi Verge, Carlos Candel Ramón, Adela María Luque Fernández, Irene Martínez Herrerías, José C. Biochemical simulations Mutation Nuclear magnetic resonance Peptides Phosphates Phosphorylation Post-translational modification Salt bridges The relative promiscuity of hub proteins such as postsynaptic density protein-95 (PSD-95) can be achieved by alternative splicing, allosteric regulation, and post-translational modifications, the latter of which is the most efficient method of accelerating cellular responses to environmental changes in vivo. Here, a mutational approach was used to determine the impact of phosphorylation and succinimidation post-translational modifications on the binding affinity of the postsynaptic density protein-95/discs large/zonula occludens-1 (PDZ3) domain of PSD-95. Molecular dynamics simulations revealed that the binding affinity of this domain is influenced by an interplay between salt-bridges linking the α3 helix, the β2–β3 loop and the positively charged Lys residues in its high-affinity hexapeptide ligand KKETAV. The α3 helix is an extra structural element that is not present in other PDZ domains, which links PDZ3 with the following SH3 domain in the PSD-95 protein. This regulatory mechanism was confirmed experimentally via thermodynamic and NMR chemical shift perturbation analyses, discarding intra-domain long-range effects. Taken together, the results presented here reveal the molecular basis of the regulatory role of the α3 extra-element and the effects of post-translational modifications of PDZ3 on its binding affinity, both energetically and dynamically. 2014-03-12T08:47:40Z 2014-03-12T08:47:40Z 2014 journal article Murciano-Calles, J.; et al. Post-Translational Modifications Modulate Ligand Recognition by the Third PDZ Domain of the MAGUK Protein PSD-95. Plos One, 9(2): e90030 (2014). [http://hdl.handle.net/10481/30789] 1932-6203 doi: 10.1371/journal.pone.0090030 http://hdl.handle.net/10481/30789 eng http://creativecommons.org/licenses/by-nc-nd/3.0/ open access Creative Commons Attribution-NonCommercial-NoDerivs 3.0 License Public Library of Science (PLOS)