Identification of a new cryptic lipid-binding domain in Ecm25. (A) Structure-based alignment of Ecm25 and five known CRAL/TRIO domain. Conserved hydrophobic sites are colored blue, polar sites are red, invariant residues are boxed, sites occupied by small residues are magenta and the ligand-binding sites of the structures are colored gray. Secondary structure elements are numbered according to the canonical C-terminal CRAL/TRIO domain structure of Sec14 (PDB:1aua), and grayed out when disrupted or absent in some of the structures. Sequences prefixed with ‘h' are from human; all others are from S. cerevisiae. (B) The truncated Ecm25-CRAL/TRIO binds to artificial membranes in flotation assays. Liposomes contain 1 mM 1-palmitoyl-2-oleoyl-snglycero-3-phosphocholine (POPC) and phosphatidylserine (PS), PtdIns or PtdIns(3,4,5)P₃, as indicated (3%, molar). Ecm25-CRAL/TRIO was analyzed by western blot.