1887

Journal of General Virology header logo

Volume 78, Issue 12

Biosynthesis, intracellular transport and enzymatic activity of an avian influenza A virus neuraminidase: role of unpaired cysteines and individual oligosaccharides Free

Abstract

Intracellular transport, glycosylation, tetrameriza- tion and enzymatic activity of the neuraminidase (NA) of fowl plague virus (FPV) were analysed in vertebrate cells after expression from a vaccinia virus vector. Tetramerization occurred with a halftime of 15 min, whereas passage through the medial Golgi apparatus and transport to the plasma membrane occurred with half-times of 2 and 3 h, respectively, suggesting a step in NA maturation beyond tetramerization that limits the rate of transport to the medial Golgi. NA transport rates were about fourfold slower than those of haemagglutinin (HA). Slow transport and processing of FPV NA was not altered by coexpression of FPV HA, nor was the transport rate of HA influenced by NA. The slow transport kinetics of NA were also observed in FPV- infected CV-1 cells. As deduced from the coding sequence, FPV NA has the shortest stalk of all naturally occurring NAs described to date and contains only three potential -glycosylation sites, which are all located in the globular head domain. Elimination of each of the three -glycosylation sites revealed that the two oligosaccharides at positions 124 and 66 are of the complex type, whereas the one at Asn-213 remains in mannose- rich form. The glycosylation mutants showed also that oligosaccharides at positions 124 and 213 of FPV NA modulate enzymatic activity. Transport of NA is not influenced by single elimination of any of the three oligosaccharide attachment sites. Mutational analysis of the three Cys residues not involved in intrachain disulfide pairing revealed that Cys-49 in the stalk of the NA molecule is responsible for the formation of disulfide-linked dimers. Analysis of cysteine mutants of FPV NA also demonstrated that disulfide-linked dimers are not absolutely necessary for the formation of enzymatically active tetramers but may stabilize the quaternary structure of NA.

  • Published Online:
© Society for General Microbiology Ltd 1997
Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-78-12-3233
1997-12-01
2024-04-25
Loading full text...

Full text loading...

/deliver/fulltext/jgv/78/12/9400974.html?itemId=/content/journal/jgv/10.1099/0022-1317-78-12-3233&mimeType=html&fmt=ahah

References

  1. Allen A. K., Skehel J. J., Yuferov V. 1977; The amino acid and carbohydrate composition of the neuraminidase of B/Lee/40 influenza virus. Journal of General Virology 37:625–628
     [Google Scholar]
  2. Alvarez E., Girones N., Davis R. J. 1989; Intermolecular disulfide bonds are not required for the expression of the dimeric state and functional activity of the transferrin receptor. EMBO Journal 8:31–40
     [Google Scholar]
  3. Aymard-Henry M., Coleman M. T., Dowdle W. R., Laver W. G., Schild G. C., Webster R. G. 1973; Influenza virus neuraminidase and neuraminidase inhibition test procedures. Bulletin of the World Health Organization 48:199–202
     [Google Scholar]
  4. Becker S., Huppertz S., Klenk H.-D., Feldmann H. 1994; The nucleoprotein of Marburg virus is phosphorylated. Journal of General Virology 75:809–818
     [Google Scholar]
  5. Bilsel P., Castrucci M., Kawaoka Y. 1993; Mutations in the cytoplasmic tail of influenza A virus neuraminidase affect incorporation into virions. Journal of Virology 67:6762–6767
     [Google Scholar]
  6. Blok J., Air G. M. 1982; Variation in the membrane-insertion and ‘stalk’ sequences in eight subtypes of influenza type A virus neuraminidase. Biochemistry 21:4001–4007
     [Google Scholar]
  7. Blumberg B. M., Giorgi C., Roux L., Raju R., Dowling P., Chollet A., Kolakofsky D. 1985; Sequence determination of the Sendai virus HN gene and its comparison to the influenza glycoproteins. Cell 41:269–278
     [Google Scholar]
  8. Bos T. J., Davis A. R., Nayak D. P. 1984; NH2-terminal hydrophobic region of influenza virus neuraminidase provides the signal function in translocation. Proceedings of the National Academy of Sciences, USA 81:2327–2331
     [Google Scholar]
  9. Bucher D. J., Palese P. 1975; The biologically active proteins of influenza virus. Neuraminidase. In Influenza Virus and Influenza pp. 83–123 Kilbourne E. D. Edited by New York: Academic Press;
     [Google Scholar]
  10. Burnet F. M. 1948; Mucins and mucoids in relation to influenza virus action. IV. Inhibition by purified mucoid of infection and haemaggluti- nation with the virus strain WSE. Australian Journal of Experimental Biology and Medical Science 26:387–391
     [Google Scholar]
  11. Castrucci M. R., Kawaoka Y. 1993; Biologic importance of neuraminidase stalk length in influenza A virus. Journal of Virology 67:759–764
     [Google Scholar]
  12. Chakrabarti S., Brechling K., Moss B. 1985; Vaccinia virus expression vector: coexpression of beta-galactosidase provides visual screening of recombinant virus plaques. Molecular and Cellular Biology 5:3403–3409
     [Google Scholar]
  13. Colman P. M., Varghese J. N., Laver W. G. 1983; Structure of the catalytic and antigenic sites in influenza virus neuraminidase. Nature 303:41–44
     [Google Scholar]
  14. Doms R. W., Lamb R. A., Rose J. K., Helenius A. 1993; Folding and assembly of viral membrane proteins. Virology 193:545–562
     [Google Scholar]
  15. Earl P. L., Moss B., Doms R. W. 1991; Folding, interaction with GRP78-BiP, assembly, and transport of the human immunodeficiency virus type 1 envelope protein. Journal of Virology 65:2047–2055
     [Google Scholar]
  16. Enami M., Enami K. 1996; Influenza virus hemagglutinin and neuraminidase glycoproteins stimulate the membrane association of the matrix protein. Journal of Virology 70:6653–6657
     [Google Scholar]
  17. Garcia-Sastre A., Palese P. 1995; The cytoplasmic tail of the neuraminidase protein of influenza A virus does not play an important role in the packaging of this protein into viral envelopes. Virus Research 37:37–47
     [Google Scholar]
  18. Hausmann J., Kretzschmar E., Garten W., Klenk H.-D. 1995; N1 neuraminidase of influenza virus A/FPV/Rostock/34 has haemadsorbing activity. Journal of General Virology 76:1719–1728
     [Google Scholar]
  19. Hogue B. G., Nayak D. P. 1992; Synthesis and processing of the influenza virus neuraminidase, a type II transmembrane glycoprotein. Virology 188:510–517
     [Google Scholar]
  20. Holsinger L. J., Lamb R. A. 1991; Influenza virus M2 integral membrane protein is a homotetramer stabilized by formation of disulfide bonds. Virology 183:32–43
     [Google Scholar]
  21. Jin H., Leser G. P., Zhang I., Lamb R. A. 1997; Influenza virus hemagglutinin and neuraminidase cytoplasmic tails control particle shape. EMBO Journal 16:1236–1247
     [Google Scholar]
  22. Klenk H.-D., Rott R., Becht H. 1972; On the structure of the influenza virus envelope. Virology 47:579–591
     [Google Scholar]
  23. Kundu A., Jabbar M. A., Nayak D. P. 1991; Cell surface transport, oligomerization, and endocytosis of chimeric type II glycoproteins : role of cytoplasmic and anchor domains. Molecular and Cellular Biology 11:2675–2685
     [Google Scholar]
  24. Laver W. G. 1978; Crystallization and peptide maps of neuraminidase ‘heads’ from H2N2 and H3N2 influenza virus strains. Virology 86:78–87
     [Google Scholar]
  25. Laver W. G., Colman P. M., Webster R. G., Hinshaw V. S., Air G. M. 1984; Influenza virus neuraminidase with hemagglutinin activity. Virology 137:314–323
     [Google Scholar]
  26. Lentz M. R., Air G. M., Webster R. G. 1987; Site-directed mutation of the active site of influenza neuraminidase and implications for the catalytic mechanism. Biochemistry 26:5351–5358
     [Google Scholar]
  27. Li S., Schulman J. L., Itamura S., Palese P. 1993; Glycosylation of neuraminidase determines the neurovirulence of influenza A/WSN/33 virus. Journal of Virology 67:6667–6673
     [Google Scholar]
  28. Liu C., Air G. M. 1993; Selection and characterisation of a neuraminidase-minus mutant of influenza virus and its rescue by cloned neuraminidase genes. Virology 194:403–407
     [Google Scholar]
  29. Luo G., Chung J., Palese P. 1993; Alterations of the stalk of the influenza virus neuraminidase: deletions and insertions. Virus Research 29:141–153
     [Google Scholar]
  30. McGinnes L. W., Morrison T. G. 1994; The role of the individual cysteine residues in the formation of the mature, antigenic HN protein of Newcastle disease virus. Virology 200:470–483
     [Google Scholar]
  31. Mackett M., Smith G. L., Moss B. 1984; General method for production and selection of infectious vaccinia virus recombinants expressing foreign genes. Journal of Virology 49:857–864
     [Google Scholar]
  32. Mitnaul L. J., Castrucci M. R., Murti K. G., Kawaoka Y. 1996; The cytoplasmic tail of influenza A virus neuraminidase (NA) affects NA incorporation into virions, virion morphology, and virulence in mice but is not essential for virus replication. Journal of Virology 70:873–879
     [Google Scholar]
  33. Ng D. T. W., Randall R. E., Lamb R. A. 1989; Intracellular maturation and transport of the SV5 type II glycoprotein hemagglutinin- neuraminidase: specific and transient association with GRP78-BiP in the endoplasmic reticulum and extensive internalization from the cell surface. Journal of Cell Biology 109:3273–3289
     [Google Scholar]
  34. Ohuchi M., Feldmann A., Ohuchi R., Klenk H.-D. 1995; Neuraminidase is essential for fowl plague hemagglutinin to show hemagglutinating activity. Virology 212:77–83
     [Google Scholar]
  35. Palese P., Tobita K., Ueda M., Compans R. W. 1974; Characterisation of temperature-sensitive influenza virus mutants defective in neuraminidase. Virology 61:397–410
     [Google Scholar]
  36. Roberts C., Garten W., Klenk H.-D. 1993; The role of conserved glycosylation in the maturation and transport of the influenza hemag-glutinin. Journal of Virology 67:3048–3060
     [Google Scholar]
  37. Roux L. 1990; Selective and transient association of Sendai virus HN glycoprotein with BiP. Virology 175:161–166
     [Google Scholar]
  38. Saito T., Taylor G., Webster R. G. 1995; Steps in maturation of influenza A virus neuraminidase. Journal of Virology 69:5011–5017
     [Google Scholar]
  39. Sanger F., Nicklen S., Coulson A. R. 1977; DNA sequencing with chain-terminating inhibitors. Proceedings of the National Academy of Sciences, USA 74:5463–5467
     [Google Scholar]
  40. Sivasubramanian N., Nayak D. B. 1987; Mutational analysis of the signal-anchor domain of influenza virus neuraminidase. Proceedings of the National Academy of Sciences, USA 84:1–5
     [Google Scholar]
  41. Sommer M., Courtney R. J. 1991; Differential rates of processing and transport of herpes simplex virus type 1 glycoproteins gB and gC. Journal of Virology 65:520–525
     [Google Scholar]
  42. Taylor G., Garman E., Webster R., Saito T., Laver G. 1993; Crystallization and preliminary X-ray studies of influenza A virus neuraminidase of subtypes N5, N6, N8 and N9. Journal of Molecular Biology 230:345–348
     [Google Scholar]
  43. Taylor J. W., Ott J., Eckstein F. 1985; The rapid generation of oligonucleotide-directed mutations at high frequency using phosphoro- thiorate modified DNA. Nucleic Acids Research 13:8765–8785
     [Google Scholar]
  44. Varghese J. N., Laver W. G., Colman P. M. 1983; Structure of the influenza virus glycoprotein antigen neuraminidase at 2·9 Å resolution. Nature 393:35–40
     [Google Scholar]
  45. Ward C. W., Murray J. M., Roxburgh C. M., Jackson D. C. 1983; Chemical and antigenic characterisation of the carbohydrate side chains of an Asian (N2) influenza virus neuraminidase. Virology 126:370–375
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-78-12-3233
Loading
Biosynthesis, intracellular transport and enzymatic activity of an avian influenza A virus neuraminidase: role of unpaired cysteines and individual oligosaccharides
J. Gen. Virol. 78, 3233 (1997); https://doi.org/10.1099/0022-1317-78-12-3233
/content/journal/jgv/10.1099/0022-1317-78-12-3233
/content/journal/jgv/10.1099/0022-1317-78-12-3233
Loading

Data & Media loading...

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error