1887

Abstract

Mutations were generated in residues at the putative catalytic site of the haemagglutinin–neuraminidase (HN) protein of Newcastle disease virus Clone 30 strain (Arg498, Glu258, Tyr262, Tyr317 and Ser418) and their effects on its three associated activities were studied. Expression of the mutant proteins at the surface of HeLa cells was similar to that of the wild-type. Sialidase, receptor-binding and fusion-promotion activities were affected to different degrees for all mutants studied. Mutant Arg498Lys lost most of its sialidase activity, although it retained most of the receptor-binding activity, suggesting that, for the former activity, besides the presence of a basic residue, the proximity to the substrate molecule is also important, as Lys is shorter than Arg. Proximity also seems to be important in substrate recognition, since Tyr262Phe retained most of its sialidase activity while Tyr262Ser lost most of it. Also, Ser418Ala displayed most of the wild-type sialidase activity. However, a kinetic and thermodynamic study of the sialidase activity of the Tyr262Ser and Ser418Ala mutants was performed and revealed that the hydroxyl group of these residues also plays an important role in catalysis, since such activity was much less effective than that of the wild-type and these mutations modified their activation energy for sialidase catalysis. The discrepancy of the modifications in sialidase and receptor-binding activities in the mutants analysed does not account for the topological coincidence of the two sites. These results also suggest that the globular head of HN protein may play a role in fusion-promotion activity.

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2004-07-01
2024-03-29
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References

  1. Bousse T., Takimoto T., Gorman W. L., Takahashi T., Portner A. 1994; Regions on the hemagglutinin–neuraminidase proteins of human parainfluenza virus type-1 and Sendai virus important for membrane fusion. Virology 204:506–514 [CrossRef]
    [Google Scholar]
  2. Bousse T., Takimoto T., Portner A. 1995; A single amino acid change enhances the fusion promotion activity of human parainfluenza virus type 1 hemagglutinin–neuraminidase glycoprotein. Virology 209:654–657 [CrossRef]
    [Google Scholar]
  3. Connaris H., Takimoto T., Russell R., Crennell S., Moustafa I., Portner A., Taylor G. 2002; Probing the sialic acid binding site of the hemagglutinin–neuraminidase of Newcastle disease virus: identification of key amino acids involved in cell binding, catalysis, and fusion. J Virol 76:1816–1824 [CrossRef]
    [Google Scholar]
  4. Crennell S. J., Garman E. F., Laver W. G., Vimr E. R., Taylor G. L. 1993; Crystal structure of a bacterial sialidase (from Salmonella typhimurium LT2) shows the same fold as an influenza virus neuraminidase. Proc Natl Acad Sci U S A 90:9852–9856 [CrossRef]
    [Google Scholar]
  5. Crennell S., Garman E., Laver G., Vimr E., Taylor G. 1994; Crystal structure of Vibrio cholerae neuraminidase reveals dual lectin-like domains in addition to the catalytic domain. Structure 2:535–544 [CrossRef]
    [Google Scholar]
  6. Crennell S., Takimoto T., Portner A., Taylor G. 2000; Crystal structure of the multifunctional paramyxovirus hemagglutinin–neuraminidase. Nat Struct Biol 7:1068–1074 [CrossRef]
    [Google Scholar]
  7. Deng R., Wang Z., Mirza A. M., Iorio R. M. 1995; Localization of a domain on the paramyxovirus attachment protein required for the promotion of cellular fusion by its homologous fusion protein spike. Virology 209:457–469 [CrossRef]
    [Google Scholar]
  8. Ferreira L., Villar E., Muñoz-Barroso I. 2004; Conformational changes of Newcastle disease virus envelope glycoproteins triggered by gangliosides. Eur J Biochem 271:581–588 [CrossRef]
    [Google Scholar]
  9. Fuerst T. R., Niles E. G., Studier F. W., Moss B. 1986; Eukaryotic transient-expression system based on recombinant vaccinia virus that synthesizes bacteriophage T7 RNA polymerase. Proc Natl Acad Sci U S A 83:8122–8126 [CrossRef]
    [Google Scholar]
  10. García-Sastre A., Cobaleda C., Cabezas J. A., Villar E. 1991; On the inhibition mechanism of the sialidase activity from Newcastle disease virus. Biol Chem Hoppe Seyler 372:923–927 [CrossRef]
    [Google Scholar]
  11. Gaskell A., Crennell S., Taylor G. 1995; The three domains of a bacterial sialidase: a beta-propeller, an immunoglobulin module and a galactose-binding jelly-roll. Structure 3:1197–1205 [CrossRef]
    [Google Scholar]
  12. Gorman W. L., Gill D. S., Scroggs R. A., Portner A. 1990; The hemagglutinin–neuraminidase glycoproteins of human parainfluenza virus type 1 and Sendai virus have high structure–function similarity with limited antigenic cross-reactivity. Virology 175:211–221 [CrossRef]
    [Google Scholar]
  13. Iorio R. M., Field G. M., Sauvron J. M., Mirza A. M., Deng R., Mahon P. J., Langedijk J. P. 2001; Structural and functional relationship between the receptor recognition and neuraminidase activities of the Newcastle disease virus hemagglutinin–neuraminidase protein: receptor recognition is dependent on neuraminidase activity. J Virol 75:1918–1927 [CrossRef]
    [Google Scholar]
  14. Kurganov B. I., Lyubarev A. E., Sánchez-Ruiz J. M., Shnyrov V. L. 1997; Analysis of differential scanning calorimetry data for proteins. Criteria of validity of one-step mechanism of irreversible protein denaturation. Biophys Chem 69:125–135 [CrossRef]
    [Google Scholar]
  15. Lamb R. A., Kolakofsky D. 2001; Paramyxoviridae : the viruses and their replication. In Fundamental Virology pp  1305–1340 Edited by Fields B. N., Knippe D. M., Kato A. New York: Lippincott–Raven;
    [Google Scholar]
  16. Loladze V. V., Ermolenko D. N., Makhatadze G. I. 2001; Heat capacity changes upon burial of polar and nonpolar groups in proteins. Protein Sci 10:1343–1352
    [Google Scholar]
  17. Merz D. C., Prehm P., Scheid A., Choppin P. W. 1981; Inhibition of the neuraminidase of paramyxoviruses by halide ions: a possible means of modulating the two activities of the HN protein. Virology 112:296–305 [CrossRef]
    [Google Scholar]
  18. Mirza A. M., Deng R., Iorio R. M. 1994; Site-directed mutagenesis of a conserved hexapeptide in the paramyxovirus hemagglutinin–neuraminidase glycoprotein: effects on antigenic structure and function. J Virol 68:5093–5099
    [Google Scholar]
  19. Muñoz-Barroso I., Cobaleda C., Zhadan G., Shnyrov V., Villar E. 1997; Dynamic properties of Newcastle disease virus envelope and their relations with viral hemagglutinin–neuraminidase membrane glycoprotein. Biochim Biophys Acta 132717–31 [CrossRef]
    [Google Scholar]
  20. Nussbaum O., Broder C. C., Berger E. A. 1994; Fusogenic mechanisms of enveloped-virus glycoproteins analyzed by a novel recombinant vaccinia virus-based assay quantitating cell fusion-dependent reporter gene activation. J Virol 68:5411–5422
    [Google Scholar]
  21. Nussbaum O., Broder C. C., Moss B., Stern L. B.-L., Rozenblatt S., Berger E. A. 1995; Functional and structural interactions between measles virus hemagglutinin and CD46. J Virol 69:3341–3349
    [Google Scholar]
  22. Ray R., Compans R. W. 1986; Monoclonal antibodies reveal extensive antigenic differences between the hemagglutinin–neuraminidase glycoproteins of human and bovine parainfluenza 3 viruses. Virology 148:232–236 [CrossRef]
    [Google Scholar]
  23. Sagrera A., Cobaleda C., Muñoz-Barroso I., Shnyrov V., Villar E. 1995; Modulation of the neuraminidase activity of the HN protein from Newcastle disease virus by substrate binding and conformational change: kinetic and thermal denaturation studies. Biochem Mol Biol Int 37:717–727
    [Google Scholar]
  24. Sagrera A., Cobaleda C., Gonzalez de Buitrago J. M., Garcia-Sastre A., Villar E. 2001; Membrane glycoproteins of Newcastle disease virus: nucleotide sequence of the hemagglutinin–neuraminidase cloned gene and structure/function relationship of predicted amino acid sequence. Glycoconj J 18:283–289 [CrossRef]
    [Google Scholar]
  25. Scheid A., Choppin P. W. 1974; Identification of biological activities of paramyxovirus glycoproteins. Activation of cell fusion, hemolysis, and infectivity of proteolytic cleavage of an inactive precursor protein of Sendai virus. Virology 57:475–490 [CrossRef]
    [Google Scholar]
  26. Sergel T., McGinnes L. W., Peeples M. E., Morrison T. G. 1993; The attachment function of the Newcastle disease virus hemagglutinin–neuraminidase protein can be separated from fusion promotion by mutation. Virology 193:717–726 [CrossRef]
    [Google Scholar]
  27. Stone-Hulslander J., Morrison T. G. 1999; Mutational analysis of heptad repeats in the membrane-proximal region of Newcastle disease virus HN protein. J Virol 73:3630–3637
    [Google Scholar]
  28. Takimoto T., Taylor G. L., Connaris H. C., Crennell S. J., Portner A. 2002; Role of the hemagglutinin–neuraminidase protein in the mechanism of paramyxovirus–cell membrane fusion. J Virol 76:13028–13033 [CrossRef]
    [Google Scholar]
  29. Tanabayashi K., Compans R. W. 1996; Functional interaction of paramyxovirus glycoproteins: identification of a domain in Sendai virus HN which promotes cell fusion. J Virol 70:6112–6118
    [Google Scholar]
  30. Taylor K. B. 2002; Effects of substrate inhibition. In Enzyme Kinetics and Mechanisms pp  106–121 Dordrecht: Kluwer Academic;
    [Google Scholar]
  31. Thompson S. D., Portner A. 1987; Localization of functional sites on the hemagglutinin–neuraminidase glycoprotein of Sendai virus by sequence analysis of antigenic and temperature-sensitive mutants. Virology 160:1–8 [CrossRef]
    [Google Scholar]
  32. Tsurudome M., Kawano M., Yuasa T., Tabata N., Nishio M., Komada H., Ito Y. 1995; Identification of regions on the hemagglutinin–neuraminidase protein of human parainfluenza virus type 2 important for promoting cell fusion. Virology 213:190–203 [CrossRef]
    [Google Scholar]
  33. Tsurudome M., Ito M., Nishio M., Kawano M., Okamoto K., Kusagawa S., Komada H., Ito Y. 1998; Identification of regions on the fusion protein of human parainfluenza virus type 2 which are required for hemagglutinin–neuraminidase protein to promote cell fusion. J Gen Virol 79:279–289
    [Google Scholar]
  34. Varghese J. N., Laver W. G., Colman P. M. 1983; Structure of the influenza virus glycoprotein antigen neuraminidase at 2·9 Å resolution. Nature 303:35–40 [CrossRef]
    [Google Scholar]
  35. Watson J. N., Dookhun V., Borgford T. J., Bennet A. J. 2003; Mutagenesis of the conserved active-site tyrosine changes a retaining sialidase into an inverting sialidase. Biochemistry 42:12682–12690 [CrossRef]
    [Google Scholar]
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