title: Key amino acid residues of ankyrin-sensitive phosphatidylethanolamine/phosphatidylcholine-lipid binding site of βI-spectrin.
abstract: It was shown previously that an ankyrin-sensitive, phosphatidylethanolamine/phosphatidylcholine (PE/PC) binding site maps to the N-terminal part of the ankyrin-binding domain of β-spectrin (ankBDn). Here we have identified the amino acid residues within this domain which are responsible for recognizing monolayers and bilayers composed of PE/PC mixtures. In vitro binding studies revealed that a quadruple mutant with substituted hydrophobic residues W1771, L1775, M1778 and W1779 not only failed to effectively bind PE/PC, but its residual PE/PC-binding activity was insensitive to inhibition with ankyrin. Structure prediction and analysis, supported by in vitro experiments, suggests that "opening" of the coiled-coil structure underlies the mechanism of this interaction. Experiments on red blood cells and HeLa cells supported the conclusions derived from the model and in vitro lipid-protein interaction results, and showed the potential physiological role of this binding. We postulate that direct interactions between spectrin ankBDn and PE-rich domains play an important role in stabilizing the structure of the spectrin-based membrane skeleton.
authors: Laboratory of Cytobiochemistry, Biotechnology Faculty, University of Wrocław, Wrocław, Poland.
authors: Marcin
authors: M
authors: Wolny
authors: Michał
authors: M
authors: Grzybek
authors: Ewa
authors: E
authors: Bok
authors: Anna
authors: A
authors: Chorzalska
authors: Marc
authors: M
authors: Lenoir
authors: Aleksander
authors: A
authors: Czogalla
authors: Klaudia
authors: K
authors: Adamczyk
authors: Adam
authors: A
authors: Kolondra
authors: Witold
authors: W
authors: Diakowski
authors: Michael
authors: M
authors: Overduin
authors: Aleksander F
authors: AF
authors: Sikorski
mesh_terms: D000595:Amino Acid Sequence; D017487:Ankyrins / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics / Q000502:physiology; D015153:Blotting, Western; D002942:Circular Dichroism; D031541:Fluorescence Resonance Energy Transfer; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010713:Phosphatidylcholines / Q000378:metabolism*; D010714:Phosphatidylethanolamines / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics / Q000502:physiology; D013049:Spectrin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D017487:Ankyrins; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D013049:Spectrin; C483858:phosphatidylethanolamine