1887

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Volume 92, Issue 10

Identification of an antibody-binding epitope on the rotavirus A non-structural protein NSP2 using phage display analysis Free

Abstract

The non-structural protein 2 (NSP2) of rotavirus has important roles in rotavirus replication associated with RNA binding, hydrolysis of NTPs and RNA, and helix destabilizing properties. A cell-culture assay using an NSP2-specific mAb and polyclonal antiserum to block virus replication showed a 73 and 96 % reduction in the amount of virus produced during replication, respectively. Phage display technology was used to identify the antibody-binding region on the NSP2 protein with the motif T-(Y/F)-Ø-Ø-Ø-X-K-Ø-G, where Ø is a hydrophilic residue and X is any amino acid. This region was mapped to the three-dimensional NSP2 crystal structure to visualize the epitope. Analysis revealed identity to a region on NSP2 that mapped to a site exposed on the surface of the protein, which could possibly interfere with a functionally important region of the protein. Antibody binding to this region could disrupt the essential roles of NSP2, such as the formation of viroplasms with NSP5 or the interaction with viral RNA, thereby indicating a possible mechanism for the observed inhibition of virus replication. Genetic analysis of the putative binding region of NSP2 revealed a high level of conservation, suggesting that the region is under strict control.

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© 2011 SGM
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2011-10-01
2024-04-16
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References

  1. Adda C. G., Tilley L., Anders R. F., Foley M. 1999; Isolation of peptides that mimic epitopes on a malarial antigen from random peptide libraries displayed on phage. Infect Immun 67:4679–4688[PubMed]
     [Google Scholar]
  2. Afrikanova I., Fabbretti E., Miozzo M. C., Burrone O. R. 1998; Rotavirus NSP5 phosphorylation is up-regulated by interaction with NSP2. J Gen Virol 79:2679–2686[PubMed]
     [Google Scholar]
  3. Berois M., Sapin C., Erk I., Poncet D., Cohen J. 2003; Rotavirus nonstructural protein NSP5 interacts with major core protein VP2. J Virol 77:1757–1763 [View Article][PubMed]
     [Google Scholar]
  4. Casey J. L., Coley A. M., Anders R. F., Murphy V. J., Humberstone K. S., Thomas A. W., Foley M. 2004; Antibodies to malaria peptide mimics inhibit Plasmodium falciparum invasion of erythrocytes. Infect Immun 72:1126–1134 [View Article][PubMed]
     [Google Scholar]
  5. Chen D., Gombold J. L., Ramig R. F. 1990; Intracellular RNA synthesis directed by temperature-sensitive mutants of simian rotavirus SA11. Virology 178:143–151 [View Article][PubMed]
     [Google Scholar]
  6. Coley A. M., Campanale N. V., Casey J. L., Hodder A. N., Crewther P. E., Anders R. F., Tilley L. M., Foley M. 2001; Rapid and precise epitope mapping of monoclonal antibodies against Plasmodium falciparum AMA1 by combined phage display of fragments and random peptides. Protein Eng 14:691–698 [View Article][PubMed]
     [Google Scholar]
  7. Colomina J., Gil M. T., Codoñer P., Buesa J. 1998; Viral proteins VP2, VP6, and NSP2 are strongly precipitated by serum and fecal antibodies from children with rotavirus symptomatic infection. J Med Virol 56:58–65 [View Article][PubMed]
     [Google Scholar]
  8. Contin R., Arnoldi F., Campagna M., Burrone O. R. 2010; Rotavirus NSP5 orchestrates recruitment of viroplasmic proteins. J Gen Virol 91:1782–1793 [View Article][PubMed]
     [Google Scholar]
  9. Cuadras M. A., Arias C. F., López S. 1997; Rotaviruses induce an early membrane permeabilization of MA104 cells and do not require a low intracellular Ca2+ concentration to initiate their replication cycle. J Virol 71:9065–9074[PubMed]
     [Google Scholar]
  10. DeLano W. L. 2002; The PyMOL molecular graphics system. DeLano Scientific, Palo Alto, CA, USA. http://www.pymol.org
  11. Denisova E., Dowling W., LaMonica R., Shaw R., Scarlata S., Ruggeri F., Mackow E. R. 1999; Rotavirus capsid protein VP5* permeabilizes membranes. J Virol 73:3147–3153[PubMed]
     [Google Scholar]
  12. Desselberger U., Huppertz H. I. 2011; Immune responses to rotavirus infection and vaccination and associated correlates of protection. J Infect Dis 203:188–195 [View Article][PubMed]
     [Google Scholar]
  13. Eichwald C., Rodriguez J. F., Burrone O. R. 2004; Characterization of rotavirus NSP2/NSP5 interactions and the dynamics of viroplasm formation. J Gen Virol 85:625–634 [View Article][PubMed]
     [Google Scholar]
  14. Fabbretti E., Afrikanova I., Vascotto F., Burrone O. R. 1999; Two non-structural rotavirus proteins, NSP2 and NSP5, form viroplasm-like structures in vivo . J Gen Virol 80:333–339[PubMed]
     [Google Scholar]
  15. Feng N., Lawton J. A., Gilbert J., Kuklin N., Vo P., Prasad B. V., Greenberg H. B. 2002; Inhibition of rotavirus replication by a non-neutralizing, rotavirus VP6-specific IgA mAb. J Clin Invest 109:1203–1213[PubMed] [CrossRef]
     [Google Scholar]
  16. Franco M. A., Angel J., Greenberg H. B. 2006; Immunity and correlates of protection for rotavirus vaccines. Vaccine 24:2718–2731 [View Article][PubMed]
     [Google Scholar]
  17. Fujioka H., Emancipator S. N., Aikawa M., Huang D. S., Blatnik F., Karban T., DeFife K., Mazanec M. B. 1998; Immunocytochemical colocalization of specific immunoglobulin A with Sendai virus protein in infected polarized epithelium. J Exp Med 188:1223–1229 [View Article][PubMed]
     [Google Scholar]
  18. Gorrell R. J., Bishop R. F. 1997; Production of reassortant viruses containing human rotavirus VP4 and SA11 VP7 for measuring neutralizing antibody following natural infection. Clin Diagn Lab Immunol 4:509–514[PubMed]
     [Google Scholar]
  19. Harris K. S., Casey J. L., Coley A. M., Masciantonio R., Sabo J. K., Keizer D. W., Lee E. F., McMahon A., Norton R. S. et al. other authors 2005; Binding hot spot for invasion inhibitory molecules on Plasmodium falciparum apical membrane antigen 1. Infect Immun 73:6981–6989 [View Article][PubMed]
     [Google Scholar]
  20. Huang Y. T., Wright A., Gao X., Kulick L., Yan H., Lamm M. E. 2005; Intraepithelial cell neutralization of HIV-1 replication by IgA. J Immunol 174:4828–4835[PubMed] [CrossRef]
     [Google Scholar]
  21. Jayaram H., Taraporewala Z., Patton J. T., Prasad B. V. 2002; Rotavirus protein involved in genome replication and packaging exhibits a HIT-like fold. Nature 417:311–315 [View Article][PubMed]
     [Google Scholar]
  22. Jiang X., Jayaram H., Kumar M., Ludtke S. J., Estes M. K., Prasad B. V. V. 2006; Cryoelectron microscopy structures of rotavirus NSP2–NSP5 and NSP2–RNA complexes: implications for genome replication. J Virol 80:10829–10835 [View Article][PubMed]
     [Google Scholar]
  23. Kaetzel C. S. 2005; The polymeric immunoglobulin receptor: bridging innate and adaptive immune responses at mucosal surfaces. Immunol Rev 206:83–99 [View Article][PubMed]
     [Google Scholar]
  24. Kirkwood C. D., Boniface K., Richardson S., Taraporewala Z. F., Patton J. T., Bishop R. F. 2008; Non-structural protein NSP2 induces heterotypic antibody responses during primary rotavirus infection and reinfection in children. J Med Virol 80:1090–1098 [View Article][PubMed]
     [Google Scholar]
  25. Kumar M., Jayaram H., Vasquez-Del Carpio R., Jiang X., Taraporewala Z. F., Jacobson R. H., Patton J. T., Prasad B. V. V. 2007; Crystallographic and biochemical analysis of rotavirus NSP2 with nucleotides reveals a nucleoside diphosphate kinase-like activity. J Virol 81:12272–12284 [View Article][PubMed]
     [Google Scholar]
  26. Matthijnssens J., Ciarlet M., Rahman M., Attoui H., Bányai K., Estes M. K., Gentsch J. R., Iturriza-Gómara M., Kirkwood C. D. et al. other authors 2008; Recommendations for the classification of group A rotaviruses using all 11 genomic RNA segments. Arch Virol 153:1621–1629 [View Article][PubMed]
     [Google Scholar]
  27. Mazanec M. B., Kaetzel C. S., Lamm M. E., Fletcher D., Nedrud J. G. 1992; Intracellular neutralization of virus by immunoglobulin A antibodies. Proc Natl Acad Sci U S A 89:6901–6905 [View Article][PubMed]
     [Google Scholar]
  28. Mohan K. V. K., Muller J., Som I., Atreya C. D. 2003; The N- and C-terminal regions of rotavirus NSP5 are the critical determinants for the formation of viroplasm-like structures independent of NSP2. J Virol 77:12184–12192 [View Article][PubMed]
     [Google Scholar]
  29. Mostov K. E. 1994; Transepithelial transport of immunoglobulins. Annu Rev Immunol 12:63–84 [View Article][PubMed]
     [Google Scholar]
  30. Parashar U. D., Gibson C. J., Bresse J. S., Glass R. I. 2006; Rotavirus and severe childhood diarrhea. Emerg Infect Dis 12:304–306[PubMed] [CrossRef]
     [Google Scholar]
  31. Poncet D., Lindenbaum P., L’Haridon R., Cohen J. 1997; In vivo and in vitro phosphorylation of rotavirus NSP5 correlates with its localization in viroplasms. J Virol 71:34–41[PubMed]
     [Google Scholar]
  32. Ray P. G., Kelkar S. D. 2004; Prevalence of neutralizing antibodies against different rotavirus serotypes in children with severe rotavirus-induced diarrhea and their mothers. Clin Diagn Lab Immunol 11:186–194[PubMed]
     [Google Scholar]
  33. Ruggeri F. M., Johansen K., Basile G., Kraehenbuhl J.-P., Svensson L. 1998; Antirotavirus immunoglobulin A neutralizes virus in vitro after transcytosis through epithelial cells and protects infant mice from diarrhea. J Virol 72:2708–2714[PubMed]
     [Google Scholar]
  34. Schuck P., Taraporewala Z., McPhie P., Patton J. T. 2001; Rotavirus nonstructural protein NSP2 self-assembles into octamers that undergo ligand-induced conformational changes. J Biol Chem 276:9679–9687 [View Article][PubMed]
     [Google Scholar]
  35. Schwartz-Cornil I., Benureau Y., Greenberg H., Hendrickson B. A., Cohen J. 2002; Heterologous protection induced by the inner capsid proteins of rotavirus requires transcytosis of mucosal immunoglobulins. J Virol 76:8110–8117 [View Article][PubMed]
     [Google Scholar]
  36. Scott J. K., Smith G. P. 1990; Searching for peptide ligands with an epitope library. Science 249:386–390 [View Article][PubMed]
     [Google Scholar]
  37. Sen A., Agresti D., Mackow E. R. 2006; Hyperphosphorylation of the rotavirus NSP5 protein is independent of serine 67 or NSP2, and the intrinsic insolubility of NSP5 is regulated by cellular phosphatases. J Virol 80:1807–1816 [View Article][PubMed]
     [Google Scholar]
  38. Svensson L., Sheshberadaran H., Vene S., Norrby E., Grandien M., Wadell G. 1987a; Serum antibody responses to individual viral polypeptides in human rotavirus infections. J Gen Virol 68:643–651 [View Article][PubMed]
     [Google Scholar]
  39. Svensson L., Sheshberadaran H., Vesikari T., Norrby E., Wadell G. 1987b; Immune response to rotavirus polypeptides after vaccination with heterologous rotavirus vaccines (RIT 4237, RRV-1). J Gen Virol 68:1993–1999 [View Article][PubMed]
     [Google Scholar]
  40. Tamura K., Peterson D., Peterson N., Stecher G., Nei M., Kumar S. 2011; mega5: Molecular Evolutionary Genetics Analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol (Epub ahead of print) [View Article][PubMed]
     [Google Scholar]
  41. Taraporewala Z. F., Patton J. T. 2001; Identification and characterization of the helix-destabilizing activity of rotavirus nonstructural protein NSP2. J Virol 75:4519–4527 [View Article][PubMed]
     [Google Scholar]
  42. Taraporewala Z., Chen D., Patton J. T. 1999; Multimers formed by the rotavirus nonstructural protein NSP2 bind to RNA and have nucleoside triphosphatase activity. J Virol 73:9934–9943[PubMed]
     [Google Scholar]
  43. Taraporewala Z. F., Schuck P., Ramig R. F., Silvestri L., Patton J. T. 2002; Analysis of a temperature-sensitive mutant rotavirus indicates that NSP2 octamers are the functional form of the protein. J Virol 76:7082–7093 [View Article][PubMed]
     [Google Scholar]
  44. Taraporewala Z. F., Jiang X., Vasquez-Del Carpio R., Jayaram H., Prasad B. V., Patton J. T. 2006; Structure–function analysis of rotavirus NSP2 octamer by using a novel complementation system. J Virol 80:7984–7994 [View Article][PubMed]
     [Google Scholar]
  45. Vascotto F., Campagna M., Visintin M., Cattaneo A., Burrone O. R. 2004; Effects of intrabodies specific for rotavirus NSP5 during the virus replicative cycle. J Gen Virol 85:3285–3290 [CrossRef]
     [Google Scholar]
  46. Vasquez-Del Carpio R., González-Nilo F. D., Jayaram H., Spencer E., Prasad B. V., Patton J. T., Taraporewala Z. F. 2004; Role of the histidine triad-like motif in nucleotide hydrolysis by the rotavirus RNA-packaging protein NSP2. J Biol Chem 279:10624–10633 [View Article][PubMed]
     [Google Scholar]
  47. Vasquez-Del Carpio R., Gonzalez-Nilo F. D., Riadi G., Taraporewala Z. F., Patton J. T. 2006; Histidine triad-like motif of the rotavirus NSP2 octamer mediates both RTPase and NTPase activities. J Mol Biol 362:539–554 [View Article][PubMed]
     [Google Scholar]
  48. Vende P., Tortorici M. A., Taraporewala Z. F., Patton J. T. 2003; Rotavirus NSP2 interferes with the core lattice protein VP2 in initiation of minus-strand synthesis. Virology 313:261–273 [View Article][PubMed]
     [Google Scholar]
  49. Ward R. L. 2008; Rotavirus vaccines: how they work or don’t work. Expert Rev Mol Med 10:e5 [View Article][PubMed]
     [Google Scholar]
  50. Ward R. L. 2009; Mechanisms of protection against rotavirus infection and disease. Pediatr Infect Dis J 28:Suppl.S57–S59 [View Article][PubMed]
     [Google Scholar]
  51. Ward R. L., Knowlton D. R., Greenberg H. B., Schiff G. M., Bernstein D. I. 1990; Serum-neutralizing antibody to VP4 and VP7 proteins in infants following vaccination with WC3 bovine rotavirus. J Virol 64:2687–2691[PubMed]
     [Google Scholar]
  52. Ward R. L., McNeal M. M., Sander D. S., Greenberg H. B., Bernstein D. I. 1993; Immunodominance of the VP4 neutralization protein of rotavirus in protective natural infections of young children. J Virol 67:464–468[PubMed]
     [Google Scholar]
  53. Wright A., Yan H., Lamm M. E., Huang Y. T. 2006; Immunoglobulin A antibodies against internal HIV-1 proteins neutralize HIV-1 replication inside epithelial cells. Virology 356:165–170 [View Article][PubMed]
     [Google Scholar]
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Identification of an antibody-binding epitope on the rotavirus A non-structural protein NSP2 using phage display analysis
J. Gen. Virol. 92, 2374 (2011); https://doi.org/10.1099/vir.0.032599-0
/content/journal/jgv/10.1099/vir.0.032599-0
/content/journal/jgv/10.1099/vir.0.032599-0
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