Secondary Structure, Orientation, Oligomerization, and Lipid Interactions of the Transmembrane Domain of Influenza Hemagglutinin

• Published in: Biochemistry (Easton) 2000-01, Vol.39 (3), p.496-507
• Main Authors: Tatulian, Suren A, Tamm, Lukas K
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Circular Dichroism; Conserved Sequence; Dimyristoylphosphatidylcholine; Disulfides; Glycosylphosphatidylinositols; Hemagglutinin Glycoproteins, Influenza Virus - chemistry; Hemagglutinin Glycoproteins, Influenza Virus - metabolism; Humans; Influenza A virus; Influenza virus; Lipid Bilayers; Macromolecular Substances; Molecular Sequence Data; Peptide Fragments - chemical synthesis; Peptide Fragments - chemistry; Phosphatidylglycerols; Protein Structure, Quaternary; Protein Structure, Secondary; Sequence Alignment; Sequence Homology, Amino Acid; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; transmembrane domain
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi991594p

Influenza virus hemagglutinin (HA), the viral envelope glycoprotein that mediates fusion between the viral and cellular membranes, is a homotrimer of three subunits, each containing two disulfide-linked polypeptide chains, HA1 and HA2. Each HA2 chain spans the viral membrane with a single putative transmembrane α-helix near its C-terminus. Fusion experiments with recombinant HAs suggest that this sequence is required for a late step of membrane fusion, as a glycosylphosphatidylinositol-anchored analogue of HA only mediates “hemifusion” of membranes, i.e., the merging of the proximal, but not distal, leaflets of the two juxtaposed lipid bilayers [Kemble et al. (1994) Cell 76, 383−391]. To find a structural explanation for the function of the transmembrane domain of HA2 in membrane fusion, we have studied the secondary structure, orientation, oligomerization, and lipid interactions of a synthetic peptide representing the transmembrane segment of X:31 HA (TMX31) by circular dichroism and attenuated total reflection Fourier transform infrared spectroscopy and by gel electrophoresis. The peptide was predominantly α-helical in detergent micelles and in phospholipid bilayers. The helicity was increased in lipid bilayers composed of acidic lipids compared to pure phosphatidylcholine bilayers. In planar lipid bilayers, the helices were oriented close to the membrane normal. TMX31 aggregated into small heat-resistant oligomers composed of two to five subunits in SDS micelles. Amide hydrogen exchange experiments indicated that a large fraction of the helical residues were accessible to water, suggesting the possibility that TMX31 forms pores in lipid bilayers. Finally, the peptide increased the acyl chain order in lipid bilayers, which may be related to the preferential association of HA with lipid “rafts” in the cell surface and which may be an important prerequisite for complete membrane fusion.