The HA protein of influenza virus binds virus particles to susceptible cells and is the major antigen against which neutralizing (protective) antibodies are directed. Variability in HA is primarily responsible for the continual evolution of new strains and subsequent influenza epidemics. HA derives its name from its ability to agglutinate erythrocytes under certain conditions.
The primary sequence of HA contains 566 amino acids (Figure 1A). A short signal sequence at the amino terminal inserts the polypeptide into the endoplasmic reticulum; the signal is then removed. The HA protein is cleaved into two subunits, HA1 and HA2, that remain tightly associated by a disulfide bridge. A hydrophobic stretch near the carboxyl terminal of HA2 anchors the HA molecule in the membrane, with a short hydrophilic tail extending into the cytoplasm. Oligosaccharide residues are added at several sites.
Fig1. Influenza virus hemagglutinin (HA) and neuraminidase (NA) surface glycoproteins. A: Primary structures of HA and NA polypeptides. The cleavage of HA into HA1 and HA2 is necessary for virus to be infectious. HA1 and HA2 remain linked by a disulfide bond (S–S). No posttranslational cleavage occurs with NA. Carbohydrate attachment sites • are shown. The hydrophobic amino acids that anchor the proteins in the viral membrane are located near the carboxyl terminal of HA and the amino terminal of NA. B: Folding of the HA1 and HA2 polypeptides in an HA monomer. Five major antigenic sites (sites A–E) that undergo change are shown as shaded areas. The amino terminal of HA2 provides fusion activity (fusion peptide). The fusion particle is buried in the molecule until it is exposed by a conformational change induced by a low pH. C: Structure of the HA trimer as it occurs on a virus particle or the surface of infected cells. Some of the sites involved in antigenic variation are shown (A). Carboxyl terminal residues (C) protrude through the membrane. D: Structure of the NA tetramer. Each NA molecule has an active site on its upper surface. The amino terminal region (N) of the polypeptides anchors the complex in the membrane. (Redrawn with permission from [A, B] Murphy BR, Webster RG: Influenza viruses, pp. 1185 and 1186, and [C, D] Kingsbury DW: Orthomyxo-and paramyxoviruses and their replication, pp. 1163 and 1172. In Fields BN [editor-in-chief] Virology. Raven Press, 1985.)
The three-dimensional structure of the HA protein has been revealed by x-ray crystallography. The HA molecule is folded into a complex structure (Figure 1B). Each linked HA1 and HA2 dimer forms an elongated stalk capped by a large globule. The base of the stalk anchors it in the membrane. Five antigenic sites on the HA molecule exhibit extensive mutations. These sites occur at regions exposed on the surface of the structure, are apparently not essential to the molecule’s stability, and are involved in viral neutralization. Other regions of the HA molecule are conserved in all iso lates, presumably because they are necessary for the molecule to retain its structure and function.
The HA spike on the virus particle is a trimer composed of three intertwined HA1 and HA2 dimers (Figure 1C). The trimerization imparts greater stability to the spike than could be achieved by a monomer. The cellular receptor binding site (viral attachment site) is a pocket located at the top of each large globule. The pocket is inaccessible to antibody.
The cleavage that separates HA1 and HA2 is necessary for the virus particle to be infectious and is mediated by cellular proteases. Influenza viruses normally remain confined to the respiratory tract because the protease enzymes that cleave HA are expressed only at those sites. Examples have been noted of more virulent viruses that have adapted to use a more ubiquitous enzyme, such as plasmin, to cleave HA and promote widespread infection of cells. The amino terminal of HA2, generated by the cleavage event, is necessary for the viral envelope to fuse with the cell membrane, an essential step in the process of viral infection. Low pH triggers a con formational change that activates the fusion activity.
