title: Spontaneous, pH-dependent membrane insertion of a transbilayer alpha-helix.
abstract: A question of fundamental importance concerning the biosynthesis of integral membrane proteins is whether transmembrane secondary structure can insert spontaneously into a lipid bilayer. It has proven to be difficult to address this issue experimentally because of the poor solubility in aqueous solution of peptides and proteins containing these extremely hydrophobic sequences. We have identified a system in which the kinetics and thermodynamics of alpha-helix insertion into lipid bilayers can be studied systematically and quantitatively using simple spectroscopic assays. Specifically, we have discovered that a 36-residue polypeptide containing the sequence of the C-helix of the integral membrane protein bacteriorhodopsin exhibits significant solubility in aqueous buffers free of both detergents and denaturants. This helix contains two aspartic acid residues in the membrane-spanning region. At neutral pH, the peptide associates with lipid bilayers in a nonhelical and presumably peripheral conformation. With a pKa of 6.0, the peptide inserts into the bilayer as a transbilayer alpha-helix. The insertion reaction proceeds rapidly at room temperature and is fully reversible.
authors.affiliation: Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06511, USA. hunt@sid.bio.columbia.edu
authors.forename: J F
authors.initials: JF
authors.lastname: Hunt
authors.forename: P
authors.initials: P
authors.lastname: Rath
authors.forename: K J
authors.initials: KJ
authors.lastname: Rothschild
authors.forename: D M
authors.initials: DM
authors.lastname: Engelman
mesh_terms: D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry; D010743:Phospholipids / Q000737:chemistry; D012995:Solubility; D013050:Spectrometry, Fluorescence; D017550:Spectroscopy, Fourier Transform Infrared; D013816:Thermodynamics
chemical_list: D008051:Lipid Bilayers; D008565:Membrane Proteins; D010446:Peptide Fragments; D010743:Phospholipids