Asymmetric dipping of bacteriophage M13 coat protein with increasing lipid bilayer thickness

D. Stopar, R.B.M. Koehorst, R.B. Spruijt, M.A. Hemminga

Research output: Contribution to journalArticleAcademicpeer-review

1 Citation (Scopus)

Abstract

Knowledge about the vertical movement of a protein with respect to the lipid bilayer plane is important to understand protein functionality in the biological membrane. In this work, the vertical displacement of bacteriophage M13 major coat protein in a lipid bilayer is used as a model system to study the molecular details of its anchoring mechanism in a homologue series of lipids with the same polar head group but different hydrophobic chain length. The major coat proteins were reconstituted into 14:1PC, 16:1PC, 18:1PC, 20:1PC, and 22:1PC bilayers, and the fluorescence spectra were measured of the intrinsic tryptophan at position 26 and BADAN attached to an introduced cysteine at position 46, located at the opposite ends of the transmembrane helix. The fluorescence maximum of tryptophan shifted for 700 cm-1 on going from 14:1PC to 22:1PC, the corresponding shift of the fluorescence maximum of BADAN at position 46 was approximately 10 times less ( 70 cm-1). Quenching of fluorescence with the spin label CAT 1 indicates that the tryptophan is becoming progressively inaccessible for the quencher with increasing bilayer thickness, whereas quenching of BADAN attached to the T46C mutant remained approximately unchanged. This supports the idea that the BADAN probe at position 46 remains at the same depth in the bilayer irrespective of its thickness and clearly indicates an asymmetrical nature of the protein dipping in the lipid bilayer. The anchoring strength at the C-terminal domain of the protein (provided by two phenylalanine residues together with four lysine residues) was estimated to be roughly 5 times larger than the anchoring strength of the N-terminal domain
Original languageEnglish
Pages (from-to)2217-2221
JournalBiochimica et Biophysica Acta. Biomembranes
Volume1788
Issue number10
DOIs
Publication statusPublished - 2009

Keywords

  • membrane-proteins
  • tryptophan residues
  • amino-acids
  • peptides
  • topology
  • helix
  • spectroscopy
  • orientation

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