Secondary structure, orientation, and oligomerization of phospholemman, a cardiac transmembrane protein

Human phospholemman (PLM) is a 72-residue protein, which is expressed at high density in the cardiac plasma membrane and in various other tissues. It forms ion channels selective for K+, Cl-, and taurine in lipid bilayers and colocalizes with the Na+/K+-ATPase and the Na+/Ca2+-exchanger, which may suggest a role in the regulation of cell volume. Here we present the first structural data based on synthetic peptides representing the transmembrane domain of PLM. Perfluoro-octaneoate-PAGE of reconstituted proteoliposomes containing PLM reveals a tetrameric homo-oligomerization. Infrared spectroscopy of proteoliposomes shows that the PLM peptide is completely alpha-helical, even beyond the hydrophobic core residues. Hydrogen/deuterium exchange experiments reveal that a core of 20-22 residues is not accessible to water, thus embedded in the lipid membrane. The maximum helix tilt is 17 degrees +/- 2 degrees obtained by attenuated total reflection infrared spectroscopy. Thus, our data support the idea of ion channel formation by the PLM transmembrane domain.     

The transmembrane domain of the SNARE protein VAMP2 is highly sensitive to its lipid environment

Neurotransmitter and hormone exocytosis depends on SNARE protein transmembrane domains and membrane lipids but their interplay is poorly understood. We investigated the interaction of the structure of VAMP2, a vesicular transmembrane SNARE protein, and membrane lipid composition by infrared spectroscopy using either the wild-type transmembrane domain (TMD), VAMP2_TM22, or a peptide mutated at the central residues G₁₀₀/C₁₀₃ (VAMP2_TM22VV) previously identified by us as being critical for exocytosis. Our data show that the structure of VAMP2_TM22, in terms of α-helices and β-sheets is strongly influenced by peptide/lipid ratios, by lipid species including cholesterol and by membrane surface charges. Differences observed in acyl chain alignments further underscore the role of the two central small amino acid residues G₁₀₀/C₁₀₃ within the transmembrane domain during lipid rearrangements in membrane fusion.