Skip to content
1887

Journal of General Virology header logo Journal of General Virology

Volume 72, Issue 5

Specificity of the polioviral proteinase 3C towards genetically engineered cleavage sites in the viral capsid No Access

Abstract

In a study of the cleavage specificity of poliovirus proteinase 3C, two mutant polioviruses were constructed to include putative 3C cleavage sites in the BC loop of VP1. The BC loop of VP1 in the wild-type virus is the neutralization antigenic site IA, consisting of a continuous chain of nine amino acids (ASTTNKDKL). The first mutant, W1-1D-BC1, has four altered amino acids in the BC loop (ASTKL); the second mutant, W1-1D-BC2, has an insertion of nine amino acids in the BC loop (ASTNKDKL). W1-1D-BC1 and W1-1D-BC2 were viable, grew to high titre and produced plaques of normal size. W1-1D-BC1 virions were resistant to proteolytic cleavage of the BC loop as well as upon incubation with a large excess of 3C , although a synthetic decapeptide (PASTQGPGKL) containing the amino acids of the BC loop in W1-1D-BC1 was cleaved by 3C. In contrast, W1-1D-BC2 yielded virus the VP1 of which was cleaved partially and completely when incubated with 3C . Our results showed that an insertion of nine amino acids into the antigenic loop of poliovirus, representing a synthetic 3C cleavage site, renders the loop susceptible to cleavage by proteinase 3C, but that this cleavage is restricted if the loop is the length of that in the native virion. This result implies that, in this case, structural restrictions override sequence determinants for cleavage of the BC loop by 3C.

  • Received:
  • Accepted:
  • Published Online:
© The Journal of General Virology 1991
Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-72-5-1159
1991-05-01
2025-01-25
Loading full text...

Full text loading...

References

  1. Arnold E., Luo M., Vriend G., Rossmann M. G., Palmenberg A. C., Parks G. D., Nicklin M. J. H., Wimmer E. 1987; Implications of the picornavirus capsid structure for polyprotein processing. Proceedings of the National Academy of Sciences, U,. S,. A 84:21–25
     [Google Scholar]
  2. Blair W. S., Hwang S., Ypma-Wong M. F., Semler B. L. 1990; A mutant poliovirus containing a novel proteolytic cleavage site in VP3 is altered in viral maturation. Journal of Virology 64:1784–1793
     [Google Scholar]
  3. Colbère-Garrapin F., Christodoulou C., Crainic R., Garapin A. C., Candrea A. 1988; Addition of a foreign oligopeptide to the major capsid protein of poliovirus. Proceedings of the National Academy of Sciences, U,. S,. A 85:8668–8672
     [Google Scholar]
  4. Fricks C. E., Icenogle J. P., Hogle J. M. 1985; Trypsin sensitivity of the Sabin strain of type 1 poliovirus:cleavage sites in virions and related particles. Journal of Virology 54:856–859
     [Google Scholar]
  5. Hanecak R., Semler B. L., Anderson C. W., Wimmer E. 1982; Proteolytic processing of poliovirus polypeptide P3-7c inhibits cleavage at glutamine-glycine pairs. Proceedings of the National Academy of Sciences, U,. S,. A 79:3973–3977
     [Google Scholar]
  6. Harber J. J., Bradely J., Anderson C. W., Wimmer E. 1991; Catalysis of poliovirus VP1 maturation cleavage is not mediated by serine 10 of VP2. Journal of Virology 65:326–334
     [Google Scholar]
  7. Hellen C. U. T., Kräusslich H. G., Wimmer E. 1989; Proteolytic processing of polyproteins in the replication of RNA viruses. Biochemistry 28:9881–9890
     [Google Scholar]
  8. Hogle J. M., Chow M., Filman D. J. 1985; Three-dimensional structure of poliovirus at 2.9 A resolution. Science 229:1358–1368
     [Google Scholar]
  9. Jore J., De Geus B., Jackson R. J., Pouwels P. H., Enger-Valk B.E. 1988; Poliovirus protein 3CD is the active protease for processing of the precursor protein P1 in vitro . Journal of General Virology 69:1627–1636
     [Google Scholar]
  10. Kuhn R. J., Tada H., Ypma-Wong M. F., Semler B. L., Wimmer E. 1988; Mutational analysis of the genome-linked protein VPg of poliovirus. Journal of Virology 62:4207–4215
     [Google Scholar]
  11. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, London 227:680–685
     [Google Scholar]
  12. Larsen G. R., Anderson C. W., Dorner A. J., Semler B. L., Wimmer E. 1982; Cleavage sites within the poliovirus capsid protein precursor. Journal of Virology 41:340–344
     [Google Scholar]
  13. Murray M. G., Kuhn R. J., Arita M., Kawamuta N., Nomoto A., Wimmer E. 1988; Poliovirus type 1/type 3 antigenic hybrid virus constructed in vitro elicits type 1 and type 3 neutralization antibodies in rabbits and monkeys. Proceedings of the National Academy of Sciences, U,. S,. A. 85:3203–3207
     [Google Scholar]
  14. Nicklin M. J. H., Toyoda H., Murray M. G., Wimmer E. 1986; Proteolytic processing in the replication of polio and related viruses. Bio/Technology 4:33–42
     [Google Scholar]
  15. Nicklin M. J. H., Kräusslich H. G., Toyoda H., Dunn J. J., Wimmer E. 1987; Poliovirus polypeptide precursors: expression in vitro and processing by 3C and 2A proteinases. Proceedings of the National Academy of Sciences, U,. S,. A. 84:4002–4006
     [Google Scholar]
  16. Nicklin M. J. H., Harris K. S., Pallai P. V., Wimmer E. 1988; Poliovirus proteinase 3C: large-scale expression, purification, and specific cleavage activity on natural and synthetic substrates in vitro . Journal of Virology 62:4586–4593
     [Google Scholar]
  17. Pallai P. V., Burkhardt F., Skoog M., Schreiner K., Bax P., Cohen K. A., Hansen G., Palladino D. E. H., Harris K. S., Nicklin M. J. H., Wimmer E. 1989; Cleavage of synthetic peptides by purified poliovirus 3C proteinase. Journal of Biological Chemistry 264:9738–9741
     [Google Scholar]
  18. Palmenberg A. C., Kirby E. M., Janda M. R., Drake N. L., Duke G. M., Potratz K. F., Collett M. S. 1984; The nucleotide and deduced amino acid sequences of the encephalomyo-carditis viral polyprotein coding region. Nucleic Acids Research 12:2969–2985
     [Google Scholar]
  19. Roivanien M., Hovi T. 1988; Cleavage of VP1 and modification of antigenic site I of type 2 polioviruses by intestinal trypsin. Journal of Virology 62:3536–3539
     [Google Scholar]
  20. Rueckert R. R., Wimmer E. 1984; Systematic nomenclature of picomavirus proteins. Journal of Virology 50:957–959
     [Google Scholar]
  21. Toyoda H., Nicklin M. J. H., Murray M. G., Anderson C. W., Dunn J. J., Studier F. W., Wimmer E. 1986; A second virus-encoded proteinase involved in proteolytic processing of poliovirus polyprotein. Cell 45:761–770
     [Google Scholar]
  22. Van Der Werf S., Bradley J., Wimmer E., Studier F. W., Dunn J. J. 1986; Synthesis of infectious poliovirus RNA by purified T7 RNA polymerase. Proceedings of the National Academy of Sciences, U,. S,. A 82:2330–2334
     [Google Scholar]
  23. Wimmer E., Emini E. A., Diamond D. C. 1986; Mapping neutralization domains of viruses. In Concepts of Viral Pathogenesis II pp 159–173 Edited by Notkins A. L., Oldstone M. B. A. New York: Springer-Verlag;
     [Google Scholar]
  24. Ypma-Wong M. F., Semler B. L. 1987; Processing determinants required for in vitro cleavage of the poliovirus P1 precursor to capsid proteins. Journal of Virology 61:3181–3189
     [Google Scholar]
  25. Ypma-Wong M. F., Dewalt P. G., Johnson V. H., Lamb J. G., Semler B. L. 1988a; Protein 3CD is the major poliovirus proteinase responsible for cleavage of the P1 capsid precursor. Virology 166:265–270
     [Google Scholar]
  26. Ypma-Wong M. F., Filman D. J., Hogle J. M., Semler B. L. 1988b; Structural domains of the poliovirus polyprotein are major determinants for proteolytic cleavage at Gln-Gly pairs. Journal of Biological Chemistry 263:17846–17856
     [Google Scholar]
/content/journal/jgv/10.1099/0022-1317-72-5-1159
Loading
Specificity of the polioviral proteinase 3C towards genetically engineered cleavage sites in the viral capsid
J. Gen. Virol. 72, 1159 (1991); https://doi.org/10.1099/0022-1317-72-5-1159
/content/journal/jgv/10.1099/0022-1317-72-5-1159
/content/journal/jgv/10.1099/0022-1317-72-5-1159
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