Conformational changes on the third PDZ domain of PSD95 upon phosphorylation of Tyr397

PSD95, a member of the membrane-associated guanylate kinase family, plays a key role in synaptic transmission. In this multidomain protein, the third PDZ domain has a complex regulatory mechanism that modulates its binding of carboxyl-terminal sequences. Phosphorylation of Tyr397, located in the additional α3 helix of this PDZ domain, has been shown to affect the domain's binding affinity. To explore the molecular basis of these changes in affinity, the crystal structure of the mutant Tyr397Glu, a point mutation intended to mimic phosphorylated tyrosine, has been determined. The crystal structure of this mutant reveals conformational changes induced by the introduction of a negative charge into the extra-domain α3 helix, suggesting communication between distant secondary-structure elements that may affect the binding affinity of this domain. Additionally, DSC folding studies show a noticeable decrease in the mutant's stability, indicating significant conformational changes. Altogether, the experimental results included in this work demonstrate that α3 is part of an electrostatic network that regulates stability and conformational changes at distant sites, including the β-hairpin at the binding site.