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Deoxyribonucleoside kinases belonging to the thymidine kinase 2 (TK2)-like group vary significantly in substrate specificity, kinetics and feed-back regulation

Knecht, Wolfgang LU ; Petersen, Gitte Ebert ; Munch-Petersen, Birgitte LU and Piskur, Jure LU (2002) In Journal of Molecular Biology 315(4). p.529-540
Abstract

In eukaryotic cells deoxyribonucleoside kinases belonging to three phylogenetic sub-families have been found: (i) thymidine kinase 1 (TK1)-like enzymes, which are strictly pyrimidine deoxyribonucleoside-specific kinases; (ii) TK2-like enzymes, which include pyrimidine deoxyribonucleoside kinases and a single multisubstrate kinase from Drosophila melanogaster (Dm-dNK); and (iii) deoxycytidine/deoxyguanosine kinase (dCK/dGK)-like enzymes, which are deoxycytidine and/or purine deoxyribonucleoside-specific kinases. We cloned and characterized two new deoxyribonucleoside kinases belonging to the TK2-like group from the insect Bombyx mori and the amphibian Xenopus laevis. The deoxyribonucleoside kinase from B. mori (Bm-dNK) turned out to be a... (More)

In eukaryotic cells deoxyribonucleoside kinases belonging to three phylogenetic sub-families have been found: (i) thymidine kinase 1 (TK1)-like enzymes, which are strictly pyrimidine deoxyribonucleoside-specific kinases; (ii) TK2-like enzymes, which include pyrimidine deoxyribonucleoside kinases and a single multisubstrate kinase from Drosophila melanogaster (Dm-dNK); and (iii) deoxycytidine/deoxyguanosine kinase (dCK/dGK)-like enzymes, which are deoxycytidine and/or purine deoxyribonucleoside-specific kinases. We cloned and characterized two new deoxyribonucleoside kinases belonging to the TK2-like group from the insect Bombyx mori and the amphibian Xenopus laevis. The deoxyribonucleoside kinase from B. mori (Bm-dNK) turned out to be a multisubstrate kinase like Dm-dNK. But uniquely for a deoxyribonucleoside kinase, Bm-dNK displayed positive cooperativity with all four natural deoxyribonucleoside substrates. The deoxyribonucleoside kinase from X. laevis (Xen-PyK) resembled closely the human and mouse TK2 enzymes displaying their characteristic Michaelis-Menten kinetic with deoxycytidine and negative cooperativity with its second natural substrate thymidine. Bm-dNK, Dm-dNK and Xen-PyK were shown to be homodimers. Significant differences in the feedback inhibition by deoxyribonucleoside triphosphates between these three enzymes were found. The insect multisubstrate deoxyribonucleoside kinases Bm-dNK and Dm-dNK were only inhibited by thymidine triphosphate, while Xen-PyK was inhibited by thymidine and deoxycytidine triphosphate in a complex pattern depending on the deoxyribonucleoside substrate. The broad substrate specificity and different feedback regulation of the multisubstrate insect deoxyribonucleoside kinases may indicate that these enzymes have a different functional role than the other members of the TK2-like group.

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type
Contribution to journal
publication status
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keywords
Amino Acid Sequence, Animals, Bombyx/enzymology, Chickens, Chromatography, Gel, Databases, Genetic, Deoxycytidine/metabolism, Drosophila melanogaster/enzymology, Feedback, Physiological, Humans, Inhibitory Concentration 50, Kinetics, Mice, Models, Biological, Models, Molecular, Molecular Sequence Data, Phosphotransferases (Alcohol Group Acceptor)/antagonists & inhibitors, Phylogeny, Protein Structure, Quaternary, Recombinant Fusion Proteins/antagonists & inhibitors, Sequence Alignment, Sequence Deletion/genetics, Substrate Specificity, Thymidine Kinase/chemistry, Xenopus laevis/genetics
in
Journal of Molecular Biology
volume
315
issue
4
pages
529 - 540
publisher
Elsevier
external identifiers
  • pmid:11812127
  • scopus:0036306450
ISSN
0022-2836
DOI
10.1006/jmbi.2001.5257
language
English
LU publication?
no
id
e75cddd2-6a5d-4305-9f78-e500b31d177b
date added to LUP
2020-07-22 14:25:16
date last changed
2024-01-02 15:00:48
@article{e75cddd2-6a5d-4305-9f78-e500b31d177b,
  abstract     = {{<p>In eukaryotic cells deoxyribonucleoside kinases belonging to three phylogenetic sub-families have been found: (i) thymidine kinase 1 (TK1)-like enzymes, which are strictly pyrimidine deoxyribonucleoside-specific kinases; (ii) TK2-like enzymes, which include pyrimidine deoxyribonucleoside kinases and a single multisubstrate kinase from Drosophila melanogaster (Dm-dNK); and (iii) deoxycytidine/deoxyguanosine kinase (dCK/dGK)-like enzymes, which are deoxycytidine and/or purine deoxyribonucleoside-specific kinases. We cloned and characterized two new deoxyribonucleoside kinases belonging to the TK2-like group from the insect Bombyx mori and the amphibian Xenopus laevis. The deoxyribonucleoside kinase from B. mori (Bm-dNK) turned out to be a multisubstrate kinase like Dm-dNK. But uniquely for a deoxyribonucleoside kinase, Bm-dNK displayed positive cooperativity with all four natural deoxyribonucleoside substrates. The deoxyribonucleoside kinase from X. laevis (Xen-PyK) resembled closely the human and mouse TK2 enzymes displaying their characteristic Michaelis-Menten kinetic with deoxycytidine and negative cooperativity with its second natural substrate thymidine. Bm-dNK, Dm-dNK and Xen-PyK were shown to be homodimers. Significant differences in the feedback inhibition by deoxyribonucleoside triphosphates between these three enzymes were found. The insect multisubstrate deoxyribonucleoside kinases Bm-dNK and Dm-dNK were only inhibited by thymidine triphosphate, while Xen-PyK was inhibited by thymidine and deoxycytidine triphosphate in a complex pattern depending on the deoxyribonucleoside substrate. The broad substrate specificity and different feedback regulation of the multisubstrate insect deoxyribonucleoside kinases may indicate that these enzymes have a different functional role than the other members of the TK2-like group.</p>}},
  author       = {{Knecht, Wolfgang and Petersen, Gitte Ebert and Munch-Petersen, Birgitte and Piskur, Jure}},
  issn         = {{0022-2836}},
  keywords     = {{Amino Acid Sequence; Animals; Bombyx/enzymology; Chickens; Chromatography, Gel; Databases, Genetic; Deoxycytidine/metabolism; Drosophila melanogaster/enzymology; Feedback, Physiological; Humans; Inhibitory Concentration 50; Kinetics; Mice; Models, Biological; Models, Molecular; Molecular Sequence Data; Phosphotransferases (Alcohol Group Acceptor)/antagonists & inhibitors; Phylogeny; Protein Structure, Quaternary; Recombinant Fusion Proteins/antagonists & inhibitors; Sequence Alignment; Sequence Deletion/genetics; Substrate Specificity; Thymidine Kinase/chemistry; Xenopus laevis/genetics}},
  language     = {{eng}},
  month        = {{01}},
  number       = {{4}},
  pages        = {{529--540}},
  publisher    = {{Elsevier}},
  series       = {{Journal of Molecular Biology}},
  title        = {{Deoxyribonucleoside kinases belonging to the thymidine kinase 2 (TK2)-like group vary significantly in substrate specificity, kinetics and feed-back regulation}},
  url          = {{http://dx.doi.org/10.1006/jmbi.2001.5257}},
  doi          = {{10.1006/jmbi.2001.5257}},
  volume       = {{315}},
  year         = {{2002}},
}