Cell membrane Neuraminidase

All enveloped human vimses acquire their phospholipid coating by budding through cellular membranes. The maturation and release of enveloped influenza particles is illustrated in Fig. 3.8. The capsid protein subunits are transported flom the ribosomes to the nucleus, where they combine with new viral RNA molecules and are assembled into the helical capsids. The haemagglutinin and neuraminidase proteins that project fiom the envelope of the normal particles migrate to the cytoplasmic membrane where they displace the normal cell membrane proteins. The assembled nucleocapsids finally pass out from the nucleus, and as they impinge on the altered cytoplasmic membrane they cause it to bulge and bud off completed enveloped particles flxm the cell. Vims particles are released in this way over a period of hours before the cell eventually dies. 

Figure 4. Accessibility of GM3 in parental and WgaR cell membranes to action of neuraminidase...

One approach to this problem would be to isolate the intact lectin-receptor molecules from the cell membranes and characterize them. Work in this direction has been initiated in our laboratory, and a method for the isolation of the peanut agglutinin receptor from membranes of neuraminidase-treated human erythrocytes on a column of peanut agglutinin-polyacryl-hydrazido-Sepharose has been developed (21). The amino acid composition, D-glucosamine ancHg-galac-tosamine content, and the electrophoretic mobility on polyacrylamide gel electrophoresis in sodium... 

The existence of specific receptors for transferrin on the reticulocyte membrane was implied by the work of Jandl and associates, who observed that trypsin virtually abolished the ability of reticulocytes to take up iron from transferrin without affecting other metabolic functions of the cells (8). Whether the effect of the enzyme was to cleave the receptor from the cell membrane or to degrade it proteolytically was not clear. Neuraminidase treatment also depressed iron uptake by reticulocytes, but to a much lesser degree than trypsin and only at much higher concentrations than needed to abolish the hemagglutinating effects of influenza virus. Subsequent work from Morgan s laboratory has confirmed these results and has shown further that binding of transferrin to the receptor protects it from proteolytic enzymes (70). 

Nicolson, G. L. (1973). Neuraminidase unmasking and failure of trypsin to unmask -D-galactose-like sites on erythrocyte, lymphoma, and normal and virus-transformed fibroblast cell membranes. J. Natl. Cancer Inst. 50, 1443-1451. 

After the virus penetrates the cell membrane, another viral protein, neuraminidase (sialidase), cleaves the glycosidic bonds to the sialic acid residues, freeing the virus to infect the cell. Inhibitors of this enTiyme such as oseltamivir (Tamiflu) and zanamivir (Relenza) are important anti-influenza agents. 

Animal V. The first stage of infection is adsorption of the vims to the exterior surface of the animal cell membrane. Adsorption occurs by interaction between a virus-coded protein on the surface of the virion and a receptor moleeule on the cell membrane. Most cell receptors are glycoproteins. Moreover, they are normal membrane glycoproteins with specific functions unrelated to virus infection. The interaction is specific. Thus, the binding protein of influenza virus interacts with the o2 3 linked terminal sialic add residue of the host cell membrane glycopFOtein treatment of a cell with sialidase (neuraminidase) renders it resistant to infection, and glycoproteins with al 6 linked siaUc acid do not serve as receptors. Under natural conditions, the presence of appropriate receptor molecules on the cell membrane is a precondition for virus infection, i.e. in the absence of receptors, the cell is not permissive for virus infection... 

Influenza vims A has two surface glycoproteins that offer potential for the development of competitive multivalent sugar clusters featuring 5-A -acetyl neuraminic acid (Neu5Ac) units [35]. Haemagglutinin (HA) binds to sialic acid receptors and fuses the cell membranes of the vims and host and neuraminidase (NA), an enzyme, cleaves sialic acid residues and is involved in release of new vims by the host cell. Whilst small molecule inhibitors of NA, such as oseltamivir (Tamiflu) and zanamavir (Relenza), are now available, no therapy based on inhibition of binding of HA has reached the clinic. 

The resistance of cells to lysis by antibodies and complement can be removed by treating the cells with neuraminidase. Glycoproteins have been shown to participate in binding Ca + ions to, and in transporting Ca + ions across, mitochondrial membranes. Removal of sialic acid residues from the surface of cultured heart cells increased the ability of the cells to exchange Ca " ions, but that of K+ ions was unaffected. Glycoproteins on the surface of human KB cells are involved in binding adenovirus. ... 

Receptor sites for serotonin have been found wherever physiological reaction to it takes place. Neuraminidase action is followed by a loss of the inhibition of the activity of serotonin by lysergic acid (Gielen, 1968) and a decrease of the number of active receptor sites a similar decrease was observed in the presence of inhibitors of sialic acid biosynthesis (Wesemann and Zilliken, 1968). The selective destruction of the receptor sites by neuraminidase in the presence of EDTA and the reactivations by addition of tissue lipids (Woolley and Gommi, 1964) led to the identification of gangliosides as receptors in the cell membrane. This was confirmed by reactivation with purified gangliosides (Woolley and Gommi, 1965). 

The role of sialic acid in the interaction between cells of different origin has been far less studied than in the interaction between identical cells. Cormack (1970) observed a decreased attachment of neuraminidase-treated Walker 25 tumor cells to the mesothelial membrane of rat, but the interpretation of this experiment performed in vivo is complicated by the resynthesis of sialic acid residues at the surface of the treated cell. Weiss and Cudney (1971) found no effect of neuraminidase on the immunolysis of mastocytoma P815 cells by sensitized spleen cells, and it is doubtful that sialic acid residues play a direct role in the interaction between different cells. It may play an indirect one, however, for example, by masking receptor sites at the surface of either human peripheral blood lymphocytes, or sheep red blood cells, as shown by the increased stability of the rosettes formed by these cells upon neuraminidase treatment (Galili and Schlesinger, 1974). 

Conflicting results were obtained in the study of the influence of sialic acid on the flow of ions through the cell membrane Click and Githens (1965) observed a sharp response of the K+ ions to the removal of sialic acid with neuraminidase in L1210 leukemia cells, whereas the transport of Na ions was only slightly inhibited the transport of ions was inhibited regardless of the direction of flow. In Ehrlich ascites cells, however, removal of sialic acid residues with neuraminidase did not alter the content of Na and ions in the cells. Only a very small reduction in the unidirectional fluxes of K ions was observed after neuraminidase treatment. These observations led Weiss and Levinson (1969) to conclude that anionic sites on the cell membrane were not of major importance in regulating the intracellular concentration of Na and K ions or the unidirectional, transmembrane flux of ions. 

Progeny vims particles then bud from patches of the infected cell s plasma membrane that contain both the viral hemagglutinin and neuraminidase. The viral envelopes therefore contain both viral membrane proteins but no cellular membrane proteins. 

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