title: Binding kinetics of sulfatide with influenza A virus hemagglutinin.
abstract: Association of a sulfated galactosyl ceramide, sulfatide, with the viral envelope glycoprotein hemagglutinin (HA) delivered to the cell surface is required for influenza A virus (IAV) replication through efficient translocation of the newly synthesized viral nucleoprotein from the nucleus to the cytoplasm. To determine whether the ectodomain of HA can bind to sulfatide, a secreted-type HA (sHA), in which the transmembrane region and cytoplasmic tail were deleted, was generated by using a baculovirus expression system. The receptor binding ability and antigenic structure of sHA were evaluated by a hemagglutination assay, solid-phase binding assay and hemagglutination inhibition assay. sHA showed subtype-specific antigenicity and binding ability to both sulfatide and gangliosides. Kinetics of sHA binding to sulfatide and GD1a was demonstrated by quartz crystal microbalance (QCM) analysis. QCM analysis showed that the sHA bound with the association rate constant (k on) of 1.41 × 10(4) M(-1) sec(-1), dissociation rate constant (k off) of 2.03 × 10(-4) sec(-1) and K d of 1.44 × 10(-8) M to sulfatide immobilized on a sensor chip. The k off values of sHA were similar for sulfatide and GD1a, whereas the k on value of sHA binding to sulfatide was 2.56-times lower than that of sHA binding to GD1a. The results indicate that sulfatide directly binds to the ectodomain of HA with high affinity.
authors.affiliation: Department of Biochemistry, School of Pharmaceutical Sciences and Global COE Program for Innovation in Human Health Sciences, University of Shizuoka, Shizuoka, 422-8526, Japan.
authors.forename: Tadanobu
authors.initials: T
authors.lastname: Takahashi
authors.forename: Sawako
authors.initials: S
authors.lastname: Kawagishi
authors.forename: Midori
authors.initials: M
authors.lastname: Masuda
authors.forename: Takashi
authors.initials: T
authors.lastname: Suzuki
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D005732:Gangliosides / Q000378:metabolism; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013433:Sulfoglycosphingolipids / Q000378:metabolism*
chemical_list: D005732:Gangliosides; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D013433:Sulfoglycosphingolipids