Mutagenic and Inhibitory Effects of Ribavirin on Hepatitis C Virus RNA Polymerase
Crotty et al. recently proposed the primary antiviral action of ribavirin to be that of a potent RNA mutagen [Crotty, S., Maag, D., Arnold, J. J., Zhong, W., Lau, J. Y., Hong, Z., Andino, R., and Cameron, C. E. (2000) Nat. Med. 6, 1375−1379]. Here we investigate the effect of ribavirin triphosphate...
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Published in | Biochemistry (Easton) Vol. 42; no. 35; pp. 10462 - 10471 |
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Main Authors | , , |
Format | Journal Article |
Language | English |
Published |
United States
American Chemical Society
09.09.2003
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Subjects | |
Online Access | Get full text |
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Summary: | Crotty et al. recently proposed the primary antiviral action of ribavirin to be that of a potent RNA mutagen [Crotty, S., Maag, D., Arnold, J. J., Zhong, W., Lau, J. Y., Hong, Z., Andino, R., and Cameron, C. E. (2000) Nat. Med. 6, 1375−1379]. Here we investigate the effect of ribavirin triphosphate (RTP) on RNA synthesis catalyzed by a full-length hepatitis C virus (HCV) RNA polymerase in vitro. HCV polymerase can use RTP as a nucleotide substrate in a template-dependent manner, incorporating it opposite a pyrimidine (C or U) template residue, but not a purine (A or G). Kinetic analysis revealed that incorporation of ribavirin monophosphate (RMP) across from C is 3 times more efficient catalytically than that across from U, as determined by the k cat/K m parameter. The efficiency of RMP incorporation, however, is 50−100 fold lower than that of the natural NMP. RMP incorporation does not lead to termination of RNA chain synthesis, as evidenced by the ability of the polymerase to extend its RNA product many nucleotides beyond the site of RMP incorporation. However, multiple-RMP incorporation at low GTP concentrations induced the formation of stalled elongation complexes, particularly at the template region containing consecutive C residues. Most, but not all, such elongation blocks can be relieved by the re-addition of GTP. When ribavirin is present in the RNA template, pyrimidine (but neither purine nor ribavirin) monophosphate is incorporated opposite ribavirin, but at an exceedingly low catalytic efficiency (200−3000-fold lower) compared to the efficiencies of those templated by A or G. Consequently, the level of RNA synthesis on a ribavirin-containing template is significantly reduced. These findings suggest that ribavirin not only is mutagenic but also interferes with HCV polymerase-mediated RNA synthesis. |
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Bibliography: | istex:C9DFD99D44D41FF3C03591B4417E8CDF57E76D57 ark:/67375/TPS-VXDVM0FV-K This work was partially supported by National Institutes of Health Grant AI 47348. M.M.C.L. is an Investigator of the Howard Hughes Medical Institute. ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
ISSN: | 0006-2960 1520-4995 |
DOI: | 10.1021/bi0344681 |