Hemagglutinin binding

It has been reported that the antigenic shifts are manifested in hemagglutinin and neuraminidase, two glycoproteins found on the surface of the influenza virion. It is suggested that the hemagglutinin binds the virus to t he target cell and when the hemagglutinin function is inhibited... 

The influenza virus is one of the most extensively studied vimses at present. The influenza virus recognizes sialic acid on the host cell surface, followed by infection and transmission [109]. Two spike-formed proteins (hemagglutinin and neuraminidase) are found on the surface of the influenza virus [110,111]. Hemagglutinin binds to the host cells through recognizing the ligand containing sialic acid on the surface of infected cells, while neuraminidase eliminates sialic acid from the cell surface to avoid inhibition of the transmission of newly formed virus particles. 

Sharma, S.K. and Singh, B.R. 1998. Hemagglutinin binding mediated protection of botuhnum neurotoxin from proteolysis. J. Nat. Toxins. 7 239-253. 

Neuraminidase inhibitors are analogs of sialic acid (7). They work by binding to the active, catalytic site of neuraminidase that protrudes from the surface of influenza viruses. Viral hemagglutinin binds to the intact sialic acid residues, which results in viral aggregation at the host cell surface and a reduction in the amount of virus that is released to infect other cells (13). 

The binding site is located at the tip of the subunit within the jelly roll structure (Figure 5.23). The sialic acid moiety of the hemagglutinin inhibitors binds in the center of a broad pocket on the surface of the barrel (Figure 5.24). In addition to this groove there is a hydrophobic channel that can accomodate large hydrophobic substituents at the C2 position of sialic acid (Figures 5.22 and 5.24). 

In addition to binding to sialic acid residues of the carbohydrate side chains of cellular proteins that the virus exploits as receptors, hemagglutinin has a second function in the infection of host cells. Viruses, bound to the plasma membrane via their membrane receptors, are taken into the cells by endocytosis. Proton pumps in the membrane of endocytic vesicles that now contain the bound viruses cause an accumulation of protons and a consequent lowering of the pH inside the vesicles. The acidic pH (below pH 6) allows hemagglutinin to fulfill its second role, namely, to act as a membrane fusogen by inducing the fusion of the viral envelope membrane with the membrane of the endosome. This expels the viral RNA into the cytoplasm, where it can begin to replicate. 

This fusogenic activity of influenza hemagglutinin is frequently exploited in the laboratory. If, for example, the virus is bound to cells at a temperature too low for endocytosis and then the pH of the external medium is lowered, the hemagglutinin causes direct fusion of the viral envelope with the plasma membrane infection is achieved without endocytosis. Similarly, artificial vesicles with hemagglutinin in their membrane and other molecules in their lumen can be caused to fuse with cells by first allowing the vesicles to bind to the plasma membrane via the hemagglutinin and then lowering the pH of the medium. In this way the contents of the vesicles are delivered to the recipient cell s cytoplasm. 

Figure 5.23 The globular head of the hemagglutinin subunit Is a distorted jelly roll stmcture (a). P strand 1 contains a long Insertion, and P strand 8 contains a bulge in the corresponding position. Each of these two strands is therefore subdivided Into shorter P strands. The loop region between P strands 3 and 4 contains a short a helix, which forms one side of the receptor binding site (yellow circle). A schematic diagram (b) Illustrates the organization of the p strands into a jelly roll motif.

Figure 5.24 Space-filling model (green) of the sialic acid binding domain of hemagglutinin with a bound inhibitor (red) Illustrating the different binding grooves. The sialic acid moiety of the Inhibitor binds in the central groove. A large hydrophobic substituent, Ri, at the Cz position of sialic acid binds in a hydrophobic channel that runs from the central groove to the bottom of the domain. (Adapted from S.J. Watowich et al.. Structure 2 719-731, 1994.)...

Figure 5.27 Schematic representation of a model for the conformational change of hemagglutinin that at low pH brings the fusion peptide to the same end of the molecule as the receptor binding site. The fusion peptide (purple) is at the end of heUx A about 100 A away from the receptor binding site in the high pH form. In the low pH fragment this region of helix A has moved about 100 A towards the area where the receptor binding sites are expected to be in the intact hemagglutinin molecule. (Adapted from D. Stuart, Nature 371 19-20, 1994.)...

The influenza virus inhibitors, zanamivir, and oseltamivir, act outside the cell after virus particles have been formed. The dtugs have been designed to fit into the active site of the viral envelope enzyme neuraminidase, which is required to cleave sialic acid off the surface of the producing cells. When its activity is blocked, new virus particles stay attached to the cell surface through binding of the virus protein hemagglutinin to sialic acid and are prevented from spreading to other cells. 

Skehel JJ, Wiley DC (2000) Receptor binding and membrane fusion in virus entry the influenza hemagglutinin. Ann Rev Biochem 69 531-569... 

Influenza virus particles are spheroidal and approximately 100 nm in diameter. The outer-membrane envelope contains 500 copies of hemagglutinin (HA) trimers and 100 copies of neuraminidase tetramers. The hemagglutinin constitutes the receptor sites for a-sialoside ligands. X-ray analyses show that the three sialic acid binding pockets reside 46 A apart, each trimer being separated on the virion surface by about 65-110 A [42],... 

There is a conserved part on neuraminidase, and this does not mutate or bind to sialic acid. X-ray crystallography revealed that this conserved part is a cleft with four parts. Drug molecules were designed to fit into this cleft and jam the neuraminidase, so that it is not available to cleave the sialic acid. When the sialic acid remains intact on the hemagglutinin, the virus is unable to attach to new cells and propagate the infection. 

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