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Journal of Virology, February 2006, p. 1440-1450, Vol. 80, No. 3
0022-538X/06/$08.00+0     doi:10.1128/JVI.80.3.1440-1450.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Role of Helix P of the Human Cytomegalovirus DNA Polymerase in Resistance and Hypersusceptibility to the Antiviral Drug Foscarnet

Egor P. Tchesnokov,1,2 Christian Gilbert,3 Guy Boivin,3 and Matthias Götte1,2,4*

McGill University AIDS Centre, Lady Davis Institute, Jewish General Hospital, Montréal, Québec, Canada H3T 1E2,1 Departments of Microbiology and Immunology,2 Medicine, McGill University, Montréal, Québec, Canada,4 Laval University and Research Center in Infectious Diseases of the CHUQ-CHUL, Ste.-Foy, Québec, Canada G1V 4G23

Received 3 May 2005/ Accepted 2 November 2005

Mutations in the human cytomegalovirus DNA polymerase (UL54) can not only decrease but also increase susceptibility to the pyrophosphate (PPi) analogue foscarnet. The proximity of L802M, which confers resistance, and K805Q, which confers hypersusceptibility, suggests a possible unifying mechanism that affects drug susceptibility in one direction or the other. We found that the polymerase activities of L802M- and K805Q-containing mutant enzymes were literally indistinguishable from that of wild-type UL54; however, susceptibility to foscarnet was decreased or increased, respectively. A comparison with the crystal structure model of the related RB69 polymerase suggests that L802 and K805 are located in the conserved {alpha}-helix P that is implicated in nucleotide binding. Although L802 and K805 do not appear to make direct contacts with the incoming nucleotide, it is conceivable that changes at these residues could exert their effects through the adjacent, highly conserved amino acids Q807 and/or K811. Our data show that a K811A substitution in UL54 causes reductions in rates of nucleotide incorporation. The activity of the Q807A mutant is only marginally affected, while this enzyme shows relatively high levels of resistance to foscarnet. Based on these data, we suggest that L802M exerts its effects through subtle structural changes in {alpha}-helix P that affect the precise positioning of Q807 and, in turn, its presumptive involvement in binding of foscarnet. In contrast, the removal of a positive charge associated with the K805Q change may facilitate access or increase affinity to the adjacent Q807.

* Corresponding author. Mailing address: McGill University, Department of Microbiology and Immunology, Room D-6, Duff Medical Building, 3775 University St., Montreal, Québec, Canada H3A 2B4. Phone: (514) 398-1365. Fax: (514) 398-7052. E-mail: matthias.gotte{at}mcgill.ca.

Journal of Virology, February 2006, p. 1440-1450, Vol. 80, No. 3
0022-538X/06/$08.00+0     doi:10.1128/JVI.80.3.1440-1450.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Tchesnokov, E. P., Obikhod, A., Schinazi, R. F., Gotte, M. (2009). Engineering of a Chimeric RB69 DNA Polymerase Sensitive to Drugs Targeting the Cytomegalovirus Enzyme. J. Biol. Chem. 284: 26439-26446 [Abstract] [Full Text]  
  • Marchand, B., Tchesnokov, E. P., Gotte, M. (2007). The Pyrophosphate Analogue Foscarnet Traps the Pre-translocational State of HIV-1 Reverse Transcriptase in a Brownian Ratchet Model of Polymerase Translocation. J. Biol. Chem. 282: 3337-3346 [Abstract] [Full Text]  
  • Scott, G. M., Weinberg, A., Rawlinson, W. D., Chou, S. (2007). Multidrug Resistance Conferred by Novel DNA Polymerase Mutations in Human Cytomegalovirus Isolates. Antimicrob. Agents Chemother. 51: 89-94 [Abstract] [Full Text]  
  • Cruchaga, C., Anso, E., Rouzaut, A., Martinez-Irujo, J. J. (2006). Selective Excision of Chain-terminating Nucleotides by HIV-1 Reverse Transcriptase with Phosphonoformate as Substrate. J. Biol. Chem. 281: 27744-27752 [Abstract] [Full Text]  


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