| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
1 Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, 1402 South Grand Avenue, St Louis, MO 63104, USA
2 Center for Macromolecular Crystallography, University of Alabama at Birmingham, 79-THT, MCLM-244, 1918 University Boulevard, Birmingham, AL 35294-0005, USA
Correspondence
Thomas J. Chambers
thomas_chambers2{at}merck.com
A series of 46 charged-to-alanine mutations in the yellow fever virus NS2B–NS3 protease, previously characterized in cell-free and transient cellular expression systems, was tested for their effects on virus recovery. Four distinct plaque phenotypes were observed in cell culture: parental plaque-size (13 mutants), reduced plaque-size (17 mutants), small plaque-size (8 mutants) and no plaque-formation (8 mutants). No mutants displayed any temperature sensitivity based on recovery of virus after RNA transfection at 32 versus 37 °C. Most small plaque-mutants were defective in growth efficiency compared with parental virus. However not all small plaque-mutants had defective 2B/3 cleavage, with some showing selective defects at other non-structural protein cleavage sites. Revertant viruses were recovered for six mutations that caused reduced plaque sizes. Same-site and second-site mutations occurred in NS2B, and one second-site mutation occurred in the NS3 protease domain. Some reversion mutations ameliorated defects in cleavage activity and plaque size caused by the original mutation. These data indicate that certain mutations that reduce NS2B–NS3 protease cleavage activity cause growth restriction of yellow fever virus in cell culture. However, for at least two mutations, processing defects other than impaired cleavage activity at the 2B/3 site may account for the mutant phenotype. The existence of reversion mutations primarily in NS2B rather than NS3, suggests that the protease domain is less tolerant of structural perturbation compared with the NS2B protein.
This article has been cited by other articles:
|
|
 |
|
  Z. Zuo, O. W. Liew, G. Chen, P. C. J. Chong, S. H. Lee, K. Chen, H. Jiang, C. M. Puah, and W. Zhu Mechanism of NS2B-Mediated Activation of NS3pro in Dengue Virus: Molecular Dynamics Simulations and Bioassays J. Virol., January 15, 2009; 83(2): 1060 - 1070. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. J. Chappell, M. J. Stoermer, D. P. Fairlie, and P. R. Young Mutagenesis of the West Nile virus NS2B cofactor domain reveals two regions essential for protease activity J. Gen. Virol., April 1, 2008; 89(4): 1010 - 1014. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Radichev, S. A. Shiryaev, A. E. Aleshin, B. I. Ratnikov, J. W. Smith, R. C. Liddington, and A. Y. Strongin Structure-based mutagenesis identifies important novel determinants of the NS2B cofactor of the West Nile virus two-component NS2B-NS3 proteinase J. Gen. Virol., March 1, 2008; 89(3): 636 - 641. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Lohr, J. E. Knox, W. Y. Phong, N. L. Ma, Z. Yin, A. Sampath, S. J. Patel, W.-L. Wang, W.-L. Chan, K. R. R. Rao, et al. Yellow fever virus NS3 protease: peptide-inhibition studies J. Gen. Virol., August 1, 2007; 88(8): 2223 - 2227. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. F. Donaldson, R. L. Graham, A. C. Sims, M. R. Denison, and R. S. Baric Analysis of Murine Hepatitis Virus Strain A59 Temperature-Sensitive Mutant TS-LA6 Suggests that nsp10 Plays a Critical Role in Polyprotein Processing J. Virol., July 1, 2007; 81(13): 7086 - 7098. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. F. Donaldson, A. C. Sims, R. L. Graham, M. R. Denison, and R. S. Baric Murine Hepatitis Virus Replicase Protein nsp10 Is a Critical Regulator of Viral RNA Synthesis J. Virol., June 15, 2007; 81(12): 6356 - 6368. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. A. Shiryaev, B. I. Ratnikov, A. E. Aleshin, I. A. Kozlov, N. A. Nelson, M. Lebl, J. W. Smith, R. C. Liddington, and A. Y. Strongin Switching the Substrate Specificity of the Two-Component NS2B-NS3 Flavivirus Proteinase by Structure-Based Mutagenesis J. Virol., May 1, 2007; 81(9): 4501 - 4509. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. A. Rumyantsev, B. R. Murphy, and A. G. Pletnev A Tick-Borne Langat Virus Mutant That Is Temperature Sensitive and Host Range Restricted in Neuroblastoma Cells and Lacks Neuroinvasiveness for Immunodeficient Mice J. Virol., February 1, 2006; 80(3): 1427 - 1439. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| INT J SYST EVOL MICROBIOL | MICROBIOLOGY | J GEN VIROL |
| J MED MICROBIOL | ALL SGM JOURNALS | |
