Structural, in silico, and functional analysis of a Disabled-2-derived peptide for recognition of sulfatides

Disabled-2 (Dab2) is an adaptor protein that regulates numerous cellular processes. Among them, Dab2 modulates the extent of platelet aggregation by two mechanisms. In the first mechanism, Dab2 intracellularly downregulates the integrin IIb{beta}3 receptor, converting it to a low affinity state for adhesion and aggregation processes. In the second mechanism, Dab2 is released extracellularly and interacts with both the integrin IIb{beta}3 receptor and sulfatides, both of which are known to be pro-aggregatory mediators, blocking their association to fibrinogen and P-selectin, respectively. Our previous research indicated that a 35-amino acid region within Dab2, which we refer to as the sulfatide-binding peptide (SBP), contains two potential sulfatide-binding motifs represented by two consecutive polybasic regions. Using a combined methodology including molecular docking, nuclear magnetic resonance, lipid-binding assays, and surface plasmon resonance, this work identifies the critical Dab2 residues within SBP that are responsible for sulfatide binding. A hydrophilic region, primarily mediated by R42, is responsible for the interaction with the sulfatide headgroup, whereas the C-terminal polybasic region contributes to interactions with the acyl chains. Furthermore, we demonstrated that, in Dab2 SBP, R42 significantly contributes to the inhibition of platelet P-selectin surface expression. The interacting Dab2 SBP residues with sulfatide resemble those described for sphingolipid-binding in other proteins, suggesting that sulfatide-binding proteins share common binding mechanisms.