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Nucleoside analogues are activated by bacterial deoxyribonucleoside kinases in a species-specific manner

Sandrini, Michael P. B.; Clausen, Anders Ranegaard LU ; On, Stephen L. W.; Aarestrup, Frank M.; Munch-Petersen, Birgitte and Piskur, Jure LU (2007) In Journal of Antimicrobial Chemotherapy 60(3). p.510-520
Abstract
Objectives: To investigate the bactericidal activity of antiviral and anticancer nucleoside analogues against a variety of pathogenic bacteria and characterize the activating enzymes, deoxyribonucleoside kinases (dNKs). Methods: Several FDA-approved nucleoside analogue drugs were screened for their potential bactericidal activity against several clinical bacterial isolates and type strains. We identified and subcloned the genes coding for putative deoxyribonucleoside kinases in Escherichia coli, Pasteurella multocida, Salmonella enterica, Yersinia enterocolitica, Bacillus cereus, Clostridium perfringens and Listeria monocytogenes. These genes were tested for their ability to increase the susceptibility of a dNK-deficient E. coli strain to... (More)
Objectives: To investigate the bactericidal activity of antiviral and anticancer nucleoside analogues against a variety of pathogenic bacteria and characterize the activating enzymes, deoxyribonucleoside kinases (dNKs). Methods: Several FDA-approved nucleoside analogue drugs were screened for their potential bactericidal activity against several clinical bacterial isolates and type strains. We identified and subcloned the genes coding for putative deoxyribonucleoside kinases in Escherichia coli, Pasteurella multocida, Salmonella enterica, Yersinia enterocolitica, Bacillus cereus, Clostridium perfringens and Listeria monocytogenes. These genes were tested for their ability to increase the susceptibility of a dNK-deficient E. coli strain to various analogues. We overexpressed, purified and characterized the substrate specificity and kinetic properties of the recombinant enzymes from S. enterica and B. cereus. Results: The tested Gram-negative bacteria were susceptible to 3 '-azido-3 '-deoxythymidine (AZT) in the concentration range 0.032-31.6 mu M except for a single E. coli isolate and two Pseudomonas aeruginosa isolates which were resistant to the tested AZT concentrations. Purified recombinant S. enterica thymidine kinase phosphorylated AZT efficiently with a K-m of 73.3 mM and k(cat)/K-m of 6.6 x 10(4) s(-1)M(-1) and is the activator of this drug in vivo. 2 ',2 '-Difluoro-2 '-deoxycytidine ( gemcitabine) was a potent antibiotic against Gram-positive bacteria in the concentration range between 0.001 and 1.0 mu M. The B. cereus deoxyadenosine kinase had a Km for gemcitabine of 33.5 mM and kcat/Km of 5.1 x 10(3) s(-1) M-1 and activates gemcitabine in vivo. S. enterica and B. cereus are now amongst the first bacteria with a completely characterized set of dNK enzymes. Conclusions: Bacterial dNKs efficiently activate nucleoside analogues in a species-specific manner. Therefore, nucleoside analogues have a potential to be employed as antibiotics in the fight against emerging multiresistant bacteria. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
thymidine kinase, antibacterial, multidrug-resistant, salvage
in
Journal of Antimicrobial Chemotherapy
volume
60
issue
3
pages
510 - 520
publisher
Oxford University Press
external identifiers
  • wos:000249882200009
  • scopus:34548133685
ISSN
1460-2091
DOI
10.1093/jac/dkm240
language
English
LU publication?
yes
id
113cf091-beb4-43b9-8b5f-326796fac681 (old id 655701)
date added to LUP
2007-12-18 13:47:32
date last changed
2017-08-20 03:33:40
@article{113cf091-beb4-43b9-8b5f-326796fac681,
  abstract     = {Objectives: To investigate the bactericidal activity of antiviral and anticancer nucleoside analogues against a variety of pathogenic bacteria and characterize the activating enzymes, deoxyribonucleoside kinases (dNKs). Methods: Several FDA-approved nucleoside analogue drugs were screened for their potential bactericidal activity against several clinical bacterial isolates and type strains. We identified and subcloned the genes coding for putative deoxyribonucleoside kinases in Escherichia coli, Pasteurella multocida, Salmonella enterica, Yersinia enterocolitica, Bacillus cereus, Clostridium perfringens and Listeria monocytogenes. These genes were tested for their ability to increase the susceptibility of a dNK-deficient E. coli strain to various analogues. We overexpressed, purified and characterized the substrate specificity and kinetic properties of the recombinant enzymes from S. enterica and B. cereus. Results: The tested Gram-negative bacteria were susceptible to 3 '-azido-3 '-deoxythymidine (AZT) in the concentration range 0.032-31.6 mu M except for a single E. coli isolate and two Pseudomonas aeruginosa isolates which were resistant to the tested AZT concentrations. Purified recombinant S. enterica thymidine kinase phosphorylated AZT efficiently with a K-m of 73.3 mM and k(cat)/K-m of 6.6 x 10(4) s(-1)M(-1) and is the activator of this drug in vivo. 2 ',2 '-Difluoro-2 '-deoxycytidine ( gemcitabine) was a potent antibiotic against Gram-positive bacteria in the concentration range between 0.001 and 1.0 mu M. The B. cereus deoxyadenosine kinase had a Km for gemcitabine of 33.5 mM and kcat/Km of 5.1 x 10(3) s(-1) M-1 and activates gemcitabine in vivo. S. enterica and B. cereus are now amongst the first bacteria with a completely characterized set of dNK enzymes. Conclusions: Bacterial dNKs efficiently activate nucleoside analogues in a species-specific manner. Therefore, nucleoside analogues have a potential to be employed as antibiotics in the fight against emerging multiresistant bacteria.},
  author       = {Sandrini, Michael P. B. and Clausen, Anders Ranegaard and On, Stephen L. W. and Aarestrup, Frank M. and Munch-Petersen, Birgitte and Piskur, Jure},
  issn         = {1460-2091},
  keyword      = {thymidine kinase,antibacterial,multidrug-resistant,salvage},
  language     = {eng},
  number       = {3},
  pages        = {510--520},
  publisher    = {Oxford University Press},
  series       = {Journal of Antimicrobial Chemotherapy},
  title        = {Nucleoside analogues are activated by bacterial deoxyribonucleoside kinases in a species-specific manner},
  url          = {http://dx.doi.org/10.1093/jac/dkm240},
  volume       = {60},
  year         = {2007},
}