title: Membrane-anchoring interactions of M13 major coat protein.
abstract: The response to hydrophobic mismatch of membrane-bound M13 major coat protein is measured using site-directed fluorescence and ESR spectroscopy. For this purpose, we investigate the membrane-anchoring interactions of M13 coat protein in model systems consisting of phosphatidylcholine bilayers that vary in hydrophobic thickness. Mutant coat proteins are prepared with an AEDANS-labeled single cysteine residue in the hinge region of the protein or at the C-terminal side of the transmembrane helix. In addition, the fluorescence of the tryptophan residue is studied as a monitor for the N-terminal side of the transmembrane helix. The fluorescence results show that the hinge region and C-terminal side of the transmembrane helix hardly respond to hydrophobic mismatch. In contrast, the N-terminal side of the helical transmembrane domain shifts to a more apolar environment, when the hydrophobic thickness is increased. The apparent strong membrane-anchoring interactions of the C-terminus are confirmed using a mutant that contains a longer transmembrane domain. As a result of this mutation, the tryptophan residue at the N-terminal side of the helical domain clearly shifts to a more polar environment, whereas the labeled position 46 at the C-terminal side is not affected. The phenylalanines in the C-terminal part of the protein play an important role in these apparent strong anchoring interactions. This is demonstrated with a mutant in which both phenylalanines are replaced by alanine residues. The phenylalanine residues in the C-terminus affect the location in the membrane of the entire transmembrane domain of the protein.
authors.affiliation: Laboratory of Biophysics, Wageningen University, Dreijenlaan 3, 6703 HA Wageningen, The Netherlands.
authors.forename: A B
authors.initials: AB
authors.lastname: Meijer
authors.forename: R B
authors.initials: RB
authors.lastname: Spruijt
authors.forename: C J
authors.initials: CJ
authors.lastname: Wolfs
authors.forename: M A
authors.initials: MA
authors.lastname: Hemminga
mesh_terms: D000595:Amino Acid Sequence; D017104:Bacteriophage M13 / Q000235:genetics / Q000378:metabolism*; D002213:Capsid / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D036022:Capsid Proteins; D003545:Cysteine / Q000235:genetics; D004134:Dimyristoylphosphatidylcholine / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D005456:Fluorescent Dyes / Q000378:metabolism; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009282:Naphthalenesulfonates / Q000378:metabolism; D010713:Phosphatidylcholines / Q000378:metabolism; D013113:Spin Labels; D014364:Tryptophan / Q000235:genetics
chemical_list: C089667:1-O-eicosanoyl-2-O-dodecanoylphosphatidylcholine; D036022:Capsid Proteins; D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D008565:Membrane Proteins; D009282:Naphthalenesulfonates; D010713:Phosphatidylcholines; D013113:Spin Labels; C025668:coat protein, Bacteriophage M13; D014364:Tryptophan; C017251:1,2-oleoylphosphatidylcholine; D003545:Cysteine; D004134:Dimyristoylphosphatidylcholine