1887

Abstract

The large non-structural polyprotein (206 kDa) of turnip yellow mosaic tymovirus (TYMV) undergoes auto-cleavage, producing N- and C-terminal proteins. Here we show that the viral proteinase responsible for this event is active when produced in , as monitored in Western blots by examining the generation of the C-terminal cleavage product after induction by IPTG. The outer boundaries and critical amino acids of the proteinase domain were characterized by deletion analysis and site-directed mutagenesis. A miniproteinase of 273 residues resulting from combined N- and C- terminal deletions still performed efficient cleavage. Sequence analysis of the bacterially-purified C-terminal cleavage product indicated that cleavage occurs between Ala and Thr of the non-structural protein.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-76-11-2853
1995-11-01
2021-10-18
Loading full text...

Full text loading...

/deliver/fulltext/jgv/76/11/JV0760112853.html?itemId=/content/journal/jgv/10.1099/0022-1317-76-11-2853&mimeType=html&fmt=ahah

References

  1. Baker S. C., Yokomori K., Dong S., Carlisle R., Gorbalenya A. E., Koonin E. V., Lai M. M. C. 1993; Identification of the catalytic sites of a papain-like cysteine proteinase of murine corona virus. Journal of Virology 67:6056–6063
    [Google Scholar]
  2. Boyer J.-C., Drugeon G., Seron K., Morch-Devignes M.-D., Agnes F., Haenni A.-L. 1993; In vitro transcripts of turnip yellow mosaic virus encompassing a long 3′ extension or produced from a full-length cDNA clone harbouring a 2 kb-long PCR- amplified segment are infectious. Research in Virology 144:339–348
    [Google Scholar]
  3. Bransom K. L., Dreher T. W. 1994; Identification of the essential cysteine and histidine residues of the turnip yellow mosaic virus protease. Virology 198:148–154
    [Google Scholar]
  4. Bransom K. L., Weiland J. J., Dreher T. W. 1991; Proteolytic maturation of the 206-kDa nonstructural protein encoded by turnip yellow mosaic virus RNA. Virology 184:351–358
    [Google Scholar]
  5. Carrington J. C., Herndon K. L. 1992; Characterization of the potyviral HC-Pro autoproteolytic cleavage site. Virology 187:308–315
    [Google Scholar]
  6. Choi G. H., Pawlyk D. M., Nuss D. L. 1991; The autocatalytic protease p29 encoded by a hypovirulence-associated virus of the chestnut blight fungus resembles the potyvirus-encoded protease HC-Pro. Virology 183:747–752
    [Google Scholar]
  7. Das T., Banerjee A. K. 1993; Expression of the vesicular stomatitis virus nucleocapsid protein gene in Escherichia coli: analysis of its biological activity in vitro . Virology 193:340–347
    [Google Scholar]
  8. Dong S., Baker S. C. 1994; Determinants of the p28 cleavage site recognized by the first papain-like cysteine proteinase of murine coronavirus. Virology 204:541–549
    [Google Scholar]
  9. Gorbalenya A. E., Koonin E. V. 1989; Viral proteins containing the purine NTP-binding sequence pattern. Nucleic Acids Research 17:8413–8440
    [Google Scholar]
  10. Kadare G., Drugeon G., Savithri H. S., Haenni A.-L. 1992; Comparison of strategies of expression of five tymovirus RNAs by in vitro translation studies. Journal of General Virology 73:493–498
    [Google Scholar]
  11. Kamer G., Argos P. 1984; Primary structural comparison of RNA-dependent polymerases from plant, animal and bacterial viruses. Nucleic Acids Research 12:7269–7282
    [Google Scholar]
  12. Lawrence D. M., Rozanov M. H., Hillman B. I. 1995; Autocatalytic processing of the 223-kDa protein of blueberry scorch carlavirus by a papain-like proteinase. Virology 207:127–135
    [Google Scholar]
  13. Lemm J. A., Rumenapf T., Strauss E. G., Strauss J. H., Rice C. M. 1994; Polypeptide requirements for assembly of functional Sindbis virus replication complexes: a model for the temporal regulation of minus- and plus-strand RNA synthesis. EMBO Journal 13:2925–2934
    [Google Scholar]
  14. Morch M.-D., Drugeon G., Szafranski P., Haenni A.-L. 1989; Proteolytic origin of the 150-kilodalton protein encoded by turnip yellow mosaic virus genomic RNA. Journal of Virology 63:5153–5158
    [Google Scholar]
  15. Rozanov M. N., Koonin E. V., Gorbalenya A. E. 1992; Conservation of the putative methyltransferase domain: a hallmark of the ‘Sindbis-like’ supergroup of positive-strand RNA viruses. Journal of General Virology 73:2129–2134
    [Google Scholar]
  16. Rozanov M. N., Drugeon G., Haenni A. -L. 1995; Papain-like proteinase of turnip yellow mosaic virus: a prototype of a new viral proteinase group. Archives of Virology 140:273–288
    [Google Scholar]
  17. Shapira R., Nuss D. L. 1991; Gene expression by a hypovirulence- associated virus of the chestnut blight fungus involves two papainlike proteinase activities. Essential residues and cleavage site requirements for p48 autoproteolysis. Journal of Biological Chemistry 266:19419–19425
    [Google Scholar]
  18. Snijder E. J., Wassenaar A. L. M., Spaan W. J. M. 1992; The 5′ end of the equine arteritis virus replicase gene encodes a papainlike cysteine protease. Journal of Virology 66:7040–7048
    [Google Scholar]
  19. Strauss J. H., Strauss E. G. 1990; Alphavirus proteinases. Seminars in Virology 1:347–356
    [Google Scholar]
  20. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. 1990; Use of T7 RNA polymerase to direct expression of cloned genes. Methods in Eniymology 185:60–89
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-76-11-2853
Loading
/content/journal/jgv/10.1099/0022-1317-76-11-2853
Loading

Data & Media loading...

Most cited this month 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