Prevalence of intrinsic disorder in the intracellular region of human single-pass type I proteins: The case of the Notch ligand Delta-4

Intrinsic disorder (ID) is a widespread phenomenon found especially in signaling and regulation-related eukaryotic proteins. The functional importance of flexible disordered regions often resides in their ability to allow proteins to bind different partners. The incidence and location of intrinsic disorder in 369 human single-pass transmembrane receptors with the type I topology was assessed based on both disorder predictions and amino acid physico-chemical properties. We provide evidence that ID concentrates in the receptors' cytoplasmic region. As a benchmark for this analysis, we present a structural study on the previously uncharacterized intracellular region of human Delta-4 (DLL4_1C), a single-pass transmembrane protein and a ligand of Notch receptors. DLL4_IC is required for receptor/ligand endocytosis; it undergoes regulated intramembrane proteolysis, and mediates protein-protein interactions through its C-terminal PDZ binding motif. Using a recombinant purified protein, we demonstrate using various biophysical methods that DLL4_IC is mainly disordered in solution but can form interconvertible local secondary structures in response to variations in the physico-chemical milieu. Most of these conformational changes occur in the highly conserved C-terminal segment that includes the PDZ-binding motif. On the basis of our results, we propose that global disorder, in concert with local preorganization, may play a role in Notch signaling mediated by Delta-4.