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New Variants of Tomato Thymidine Kinase 1 Selected for Increased Sensitivity of E. coli KY895 towards Azidothymidine.

Slot Christiansen, Louise LU ; Egeblad, Louise LU ; Munch-Petersen, Birgitte LU ; Piskur, Jure LU and Knecht, Wolfgang LU (2015) In Cancers 7(2). p.966-980
Abstract
Nucleoside analogues (NA) are prodrugs that are phosphorylated by deoxyribonucleoside kinases (dNKs) as the first step towards a compound toxic to the cell. During the last 20 years, research around dNKs has gone into new organisms other than mammals and viruses. Newly discovered dNKs have been tested as enzymes for suicide gene therapy. The tomato thymidine kinase 1 (ToTK1) is a dNK that has been selected for its in vitro kinetic properties and then successfully been tested in vivo for the treatment of malignant glioma. We present the selection of two improved variants of ToTK1 generated by random protein engineering for suicide gene therapy with the NA azidothymidine (AZT).We describe their selection, recombinant production and a... (More)
Nucleoside analogues (NA) are prodrugs that are phosphorylated by deoxyribonucleoside kinases (dNKs) as the first step towards a compound toxic to the cell. During the last 20 years, research around dNKs has gone into new organisms other than mammals and viruses. Newly discovered dNKs have been tested as enzymes for suicide gene therapy. The tomato thymidine kinase 1 (ToTK1) is a dNK that has been selected for its in vitro kinetic properties and then successfully been tested in vivo for the treatment of malignant glioma. We present the selection of two improved variants of ToTK1 generated by random protein engineering for suicide gene therapy with the NA azidothymidine (AZT).We describe their selection, recombinant production and a subsequent kinetic and biochemical characterization. Their improved performance in killing of E. coli KY895 is accompanied by an increase in specificity for the NA AZT over the natural substrate thymidine as well as a decrease in inhibition by dTTP, the end product of the nucleoside salvage pathway for thymidine. The understanding of the enzymatic properties improving the variants efficacy is instrumental to further develop dNKs for use in suicide gene therapy. (Less)
Please use this url to cite or link to this publication:
author
; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Cancers
volume
7
issue
2
pages
966 - 980
publisher
MDPI AG
external identifiers
  • pmid:26061968
  • scopus:84934944172
  • pmid:26061968
  • wos:000209951100026
ISSN
2072-6694
DOI
10.3390/cancers7020819
language
English
LU publication?
yes
id
df2f08db-38e3-4bec-ba6a-04e1cc4bb14f (old id 7487063)
alternative location
http://www.ncbi.nlm.nih.gov/pubmed/26061968?dopt=Abstract
date added to LUP
2016-04-04 09:32:24
date last changed
2022-04-15 23:47:46
@article{df2f08db-38e3-4bec-ba6a-04e1cc4bb14f,
  abstract     = {{Nucleoside analogues (NA) are prodrugs that are phosphorylated by deoxyribonucleoside kinases (dNKs) as the first step towards a compound toxic to the cell. During the last 20 years, research around dNKs has gone into new organisms other than mammals and viruses. Newly discovered dNKs have been tested as enzymes for suicide gene therapy. The tomato thymidine kinase 1 (ToTK1) is a dNK that has been selected for its in vitro kinetic properties and then successfully been tested in vivo for the treatment of malignant glioma. We present the selection of two improved variants of ToTK1 generated by random protein engineering for suicide gene therapy with the NA azidothymidine (AZT).We describe their selection, recombinant production and a subsequent kinetic and biochemical characterization. Their improved performance in killing of E. coli KY895 is accompanied by an increase in specificity for the NA AZT over the natural substrate thymidine as well as a decrease in inhibition by dTTP, the end product of the nucleoside salvage pathway for thymidine. The understanding of the enzymatic properties improving the variants efficacy is instrumental to further develop dNKs for use in suicide gene therapy.}},
  author       = {{Slot Christiansen, Louise and Egeblad, Louise and Munch-Petersen, Birgitte and Piskur, Jure and Knecht, Wolfgang}},
  issn         = {{2072-6694}},
  language     = {{eng}},
  number       = {{2}},
  pages        = {{966--980}},
  publisher    = {{MDPI AG}},
  series       = {{Cancers}},
  title        = {{New Variants of Tomato Thymidine Kinase 1 Selected for Increased Sensitivity of E. coli KY895 towards Azidothymidine.}},
  url          = {{http://dx.doi.org/10.3390/cancers7020819}},
  doi          = {{10.3390/cancers7020819}},
  volume       = {{7}},
  year         = {{2015}},
}