Neural precursor cell expressed, developmentally down regulated protein 4-2 (Nedd4-2/NEDD4L) is a HECT type E3 ubiquitin ligase that controls ubiquitination and endocytosis of many membrane proteins, including epithelial sodium channels and other ion transporters, thereby influencing blood pressure, neuronal excitability and kidney function. As a multidomain protein composed of a Ca²⁺ binding C2 domain, four WW domains and a catalytic HECT domain, Nedd4-2 is tightly regulated by its own intramolecular architecture as well as by calcium, phosphorylation and binding of adaptor proteins such as 14-3-3.

In our work, we use integrative structural and biochemical approaches to dissect this regulation. We show that full length Nedd4-2 adopts a compact, autoinhibited conformation in which the C2 and WW domains restrict access to key surfaces of the HECT domain, and that Ca²⁺ dependent membrane binding can unlock this state and enhance catalytic activity. We further demonstrate that phosphorylation dependent binding of 14-3-3 to motifs flanking WW2–WW3 remodels the domain arrangement, alters the mobility and solvent exposure of selected WW domains and the catalytic site, and thereby modulates Nedd4-2 ubiquitination of physiological substrates.

Contact at IPHYS: Veronika Obšilová, Laboratory of Structural Biology of Signaling Proteins, veronika.obsilova@fgu.cas.cz