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Jure Piskur - One of the best experts on this subject based on the ideXlab platform.

  • Plants salvage Deoxyribonucleosides in mitochondria.
    Nucleosides Nucleotides & Nucleic Acids, 2014
    Co-Authors: Anders R Clausen, Birgitte Munch-petersen, Zeeshan Mutahir, Jure Piskur
    Abstract:

    Deoxyribonucleoside kinases phosphorylate Deoxyribonucleosides into the corresponding 5′-monophosphate Deoxyribonucleosides to supply the cell with nucleic acid precursors. In mitochondrial fractions of the model plant Arabidopsis thaliana, we detected deoxyadenosine and thymidine kinase activities, while the cytosol fraction contained six-fold lower activity and chloroplasts contained no measurable activities. In addition, a mitochondrial fraction isolated from the potato Solanum tuberosum contained thymidine kinase and deoxyadenosine kinase activities. We conclude that an active salvage of Deoxyribonucleosides in plants takes place in their mitochondria. In general, the observed localization of the plant dNK activities in the mitochondrion suggests that plants have a different organization of the deoxyribonucleoside salvage compared to mammals.

  • two thymidine kinases and one multisubstrate deoxyribonucleoside kinase salvage dna precursors in arabidopsis thaliana
    FEBS Journal, 2012
    Co-Authors: Anders R Clausen, Wolfgang Knecht, Michael P B Sandrini, Birgitte Munchpetersen, Lenart Girandon, Ashfaq Ali, Elzbieta Rozpedowska, Erik Andreasson, Jure Piskur
    Abstract:

    Deoxyribonucleotides are the building blocks of DNA and can be synthesized via de novo and salvage pathways. Deoxyribonucleoside kinases (dNKs) salvage Deoxyribonucleosides by transfer of a phosphate group to the 5' of a deoxyribonucleoside. This salvage pathway is well characterized in mammals but in contrast little is known about how plants salvage Deoxyribonucleosides. We show that during salvage, Deoxyribonucleosides can be phosphorylated by extracts of Arabidopsis thaliana into corresponding mono-phosphate compounds with a surprising preference for purines over pyrimidines. Deoxyribonucleoside kinase activities were present in all tissues during all growth stages. In the A. thaliana genome we identified two types of genes that could encode enzymes which are involved in the salvage of Deoxyribonucleosides. Thymidine kinase activity was encoded by two thymidine kinase 1-like genes (AtTK1a and AtTK1b) and deoxyadenosine, deoxyguanosine and deoxycytidine kinase activities were encoded by a single AtdNK gene. T-DNA insertion lines of AtTK1a and AtTK1b mutant genes had normal growth, but AtTK1a AtTK1b double mutants died at an early stage, which indicates that AtTK1a and AtTK1b catalyze redundant reactions. Our results point out a crucial role for salvage of thymidine during early plant development. © 2012 The Authors Journal compilation © 2012 FEBS. (Less)

  • animal deoxyribonucleoside kinases forward and retrograde evolution of their substrate specificity
    FEBS Letters, 2004
    Co-Authors: Jure Piskur, Wolfgang Knecht, Michael P B Sandrini, Birgitte Munchpetersen
    Abstract:

    Deoxyribonucleoside kinases, which catalyse the phosphorylation of Deoxyribonucleosides, are present in several copies in most multicellular organisms and therefore represent an excellent model to study gene duplication and specialisation of the duplicated copies through partitioning of substrate specificity. Recent studies suggest that in the animal lineage one of the progenitor kinases, the so-called dCK/dGK/TK2-like gene, was duplicated prior to separation of the insect and mammalian lineages. Thereafter, insects lost all but one kinase, dNK (EC 2.7.1.145), which subsequently, through remodelling of a limited number of amino acid residues, gained a broad substrate specificity.

  • mosquito has a single multisubstrate deoxyribonucleoside kinase characterized by unique substrate specificity
    Nucleic Acids Research, 2003
    Co-Authors: Wolfgang Knecht, Gitte Petersen, Michael P B Sandrini, Leif Sondergaard, Birgitte Munchpetersen, Jure Piskur
    Abstract:

    In mammals four deoxyribonucleoside kinases, with a relatively restricted specificity, catalyze the phosphorylation of the four natural Deoxyribonucleosides. When cultured mosquito cells, originating from the malaria vector Anopheles gambiae, were examined for deoxyribonucleoside kinase activities, only a single enzyme was isolated. Subsequently, the corresponding gene was cloned and over-expressed. While the mosquito kinase (Ag-dNK) phosphorylated all four natural Deoxyribonucleosides, it displayed an unexpectedly higher relative efficiency for the phosphorylation of purine versus pyrimidine Deoxyribonucleosides than the fruit fly multisubstrate deoxyribonucleoside kinase (EC 2.7.1.145). In addition, Ag-dNK could also phosphorylate some medically interesting nucleoside analogs, like stavudine (D4T), 2-chloro-deoxyadenosine (CdA) and 5-bromo-vinyl-deoxyuridine (BVDU). Although the biological significance of multisubstrate deoxyribonucleoside kinases and their diversity among insects remains unclear, the observed variation provides a whole range of applications, as species specific and highly selective targets for insecticides, they have a potential to be used in the enzymatic production of various (di-)(deoxy-)ribonucleoside monophosphates, and as suicide genes in gene therapy.

  • a few amino acid substitutions can convert deoxyribonucleoside kinase specificity from pyrimidines to purines
    The EMBO Journal, 2002
    Co-Authors: Wolfgang Knecht, Kenth Johansson, Hans Eklund, Michael P B Sandrini, Birgitte Munchpetersen, Jure Piskur
    Abstract:

    In mammals, the four native Deoxyribonucleosides are phosphorylated to the corresponding monophosphates by four deoxyribonucleoside kinases, which have specialized substrate specificities. These four enzymes are likely to originate from a common progenitor kinase. Insects appear to have only one multisubstrate deoxyribonucleoside kinase (dNK, EC 2.7.1.145), which prefers pyrimidine nucleosides, but can also phosphorylate purine substrates. When the structures of the human deoxyguanosine kinase (dGK, EC 2.7.1.113) and the dNK from Drosophila melanogaster were compared, a limited number of amino acid residues were identified and proposed to be responsible for the substrate specificity. Three of these key residues in Drosophila dNK were then mutagenized and the mutant enzymes were characterized regarding their ability to phosphorylate native Deoxyribonucleosides and nucleoside analogs. The mutations converted the dNK substrate specificity from predominantly pyrimidine specific into purine specific. A similar scenario could have been followed during the evolution of kinases. Upon gene duplication of the progenitor kinase, only a limited number of single amino acid changes has taken place in each copy and resulted in substrate-specialized enzymes.

Birgitte Munchpetersen - One of the best experts on this subject based on the ideXlab platform.

  • isolation of a novel protein p12 from adult drosophila melanogaster that inhibits deoxyribonucleoside and protein kinase activities and activates 3 5 exonuclease activity
    Nucleosides Nucleotides & Nucleic Acids, 2016
    Co-Authors: Louise Slot Christiansen, Leif Sondergaard, Gabriella Christina Van Zanten, Dvora Berenstein, Marianne Lauridsen, Soren Kjaerulff, Birgitte Munchpetersen
    Abstract:

    ABSTRACTWe have previously found that Drosophila melanogaster only has one deoxyribonucleoside kinase, Dm-dNK, however, capable to phosphorylate all four natural Deoxyribonucleosides. Dm-dNK was originally isolated from an embryonic cell line. We wanted to study the expression of Dm-dNK during development from embryonic cells to adult flies and found declining Dm-dNK activity during development and no activity in adult flies. Surprisingly, the extract from adult flies exhibited a strong inhibitory effect on deoxyribonucloside kinase activity. The dNK-inhibitor was precipitable with ammonium sulfate, and was purified to a high degree by gel-filtration as indicated by LC-MS/MS analysis. Since the inhibitor eluted from G-200 gel-filtration with a size of 10–13 kDa, we named it P12. We tested the purified fraction for specificity towards various enzymes and found that both mammalian and bacterial dNKs were inhibited, whereas there was no effect on hexokinase and pyruvate kinases and acidic phosphatase. Howeve...

  • non viral deoxyribonucleoside kinases diversity and practical use
    Journal of Genetics and Genomics, 2015
    Co-Authors: Louise Slot Christiansen, Birgitte Munchpetersen, Wolfgang Knecht
    Abstract:

    Deoxyribonucleoside kinases (dNKs) phosphorylate Deoxyribonucleosides to their corresponding monophosphate compounds. dNks also phosphorylate deoxyribonucleoside analogues that are used in the treatment of cancer or viral infections. The study of the mammalian dNKs has therefore always been of great medical interest. However, during the last 20 years, research on dNKs has gone into non-mammalian organisms. In this review, we focus on non-viral dNKs, in particular their diversity and their practical applications. The diversity of this enzyme family in different organisms has proven to be valuable in studying the evolution of enzymes. Some of these newly discovered enzymes have been useful in numerous practical applications in medicine and biotechnology, and have contributed to our understanding of the structural basis of nucleoside and nucleoside analogue activation.

  • two thymidine kinases and one multisubstrate deoxyribonucleoside kinase salvage dna precursors in arabidopsis thaliana
    FEBS Journal, 2012
    Co-Authors: Anders R Clausen, Wolfgang Knecht, Michael P B Sandrini, Birgitte Munchpetersen, Lenart Girandon, Ashfaq Ali, Elzbieta Rozpedowska, Erik Andreasson, Jure Piskur
    Abstract:

    Deoxyribonucleotides are the building blocks of DNA and can be synthesized via de novo and salvage pathways. Deoxyribonucleoside kinases (dNKs) salvage Deoxyribonucleosides by transfer of a phosphate group to the 5' of a deoxyribonucleoside. This salvage pathway is well characterized in mammals but in contrast little is known about how plants salvage Deoxyribonucleosides. We show that during salvage, Deoxyribonucleosides can be phosphorylated by extracts of Arabidopsis thaliana into corresponding mono-phosphate compounds with a surprising preference for purines over pyrimidines. Deoxyribonucleoside kinase activities were present in all tissues during all growth stages. In the A. thaliana genome we identified two types of genes that could encode enzymes which are involved in the salvage of Deoxyribonucleosides. Thymidine kinase activity was encoded by two thymidine kinase 1-like genes (AtTK1a and AtTK1b) and deoxyadenosine, deoxyguanosine and deoxycytidine kinase activities were encoded by a single AtdNK gene. T-DNA insertion lines of AtTK1a and AtTK1b mutant genes had normal growth, but AtTK1a AtTK1b double mutants died at an early stage, which indicates that AtTK1a and AtTK1b catalyze redundant reactions. Our results point out a crucial role for salvage of thymidine during early plant development. © 2012 The Authors Journal compilation © 2012 FEBS. (Less)

  • animal deoxyribonucleoside kinases forward and retrograde evolution of their substrate specificity
    FEBS Letters, 2004
    Co-Authors: Jure Piskur, Wolfgang Knecht, Michael P B Sandrini, Birgitte Munchpetersen
    Abstract:

    Deoxyribonucleoside kinases, which catalyse the phosphorylation of Deoxyribonucleosides, are present in several copies in most multicellular organisms and therefore represent an excellent model to study gene duplication and specialisation of the duplicated copies through partitioning of substrate specificity. Recent studies suggest that in the animal lineage one of the progenitor kinases, the so-called dCK/dGK/TK2-like gene, was duplicated prior to separation of the insect and mammalian lineages. Thereafter, insects lost all but one kinase, dNK (EC 2.7.1.145), which subsequently, through remodelling of a limited number of amino acid residues, gained a broad substrate specificity.

  • mosquito has a single multisubstrate deoxyribonucleoside kinase characterized by unique substrate specificity
    Nucleic Acids Research, 2003
    Co-Authors: Wolfgang Knecht, Gitte Petersen, Michael P B Sandrini, Leif Sondergaard, Birgitte Munchpetersen, Jure Piskur
    Abstract:

    In mammals four deoxyribonucleoside kinases, with a relatively restricted specificity, catalyze the phosphorylation of the four natural Deoxyribonucleosides. When cultured mosquito cells, originating from the malaria vector Anopheles gambiae, were examined for deoxyribonucleoside kinase activities, only a single enzyme was isolated. Subsequently, the corresponding gene was cloned and over-expressed. While the mosquito kinase (Ag-dNK) phosphorylated all four natural Deoxyribonucleosides, it displayed an unexpectedly higher relative efficiency for the phosphorylation of purine versus pyrimidine Deoxyribonucleosides than the fruit fly multisubstrate deoxyribonucleoside kinase (EC 2.7.1.145). In addition, Ag-dNK could also phosphorylate some medically interesting nucleoside analogs, like stavudine (D4T), 2-chloro-deoxyadenosine (CdA) and 5-bromo-vinyl-deoxyuridine (BVDU). Although the biological significance of multisubstrate deoxyribonucleoside kinases and their diversity among insects remains unclear, the observed variation provides a whole range of applications, as species specific and highly selective targets for insecticides, they have a potential to be used in the enzymatic production of various (di-)(deoxy-)ribonucleoside monophosphates, and as suicide genes in gene therapy.

Anna Karlsson - One of the best experts on this subject based on the ideXlab platform.

  • progressive loss of mitochondrial dna in thymidine kinase 2 deficient mice
    Human Molecular Genetics, 2008
    Co-Authors: Xiaoshan Zhou, Magnus Johansson, Nicola Solaroli, Mia Bjerke, James B Stewart, Bjorn Rozell, Anna Karlsson
    Abstract:

    Deficient enzymatic activity of the mitochondrial deoxyribonucleoside kinases deoxyguanosine kinase (DGUOK) or thymidine kinase 2 (TK2) cause mitochondrial DNA (mtDNA)-depletion syndromes in humans. Here we report the generation of a Tk2-deficient mouse strain and show that the mice develop essentially normally for the first week but from then on exhibit growth retardation and die within 2-4 weeks of life. Several organs including skeletal muscle, heart, liver and spleen showed progressive loss of mtDNA without increased mtDNA mutations or structural alterations. There were no major histological changes in skeletal muscle, but heart muscle showed disorganized and damaged muscle fibers. Electron microscopy showed mitochondria with distorted cristae. The Tk2-deficient mice exhibited pronounced hypothermia and showed loss of hypodermal fat and abnormal brown adipose tissue. We conclude that Tk2 has a major role in supplying deoxyribonucleotides for mtDNA replication and that other pathways of deoxyribonucleotide synthesis cannot compensate for loss of this enzyme.

  • the drosophila melanogaster ump cmp kinase cdna encodes an n terminal mitochondrial import signal
    Biochemical and Biophysical Research Communications, 2003
    Co-Authors: Sophie Curbo, Magnus Johansson, Marjan Amiri, Fariba Foroogh, Anna Karlsson
    Abstract:

    Drosophila melanogaster cells express a multi-substrate deoxyribonucleoside kinase that phosphorylates both purine and pyrimidine Deoxyribonucleosides. The subsequent phosphorylation step is catalyzed by nucleoside monophosphate kinases. We have cloned and characterized the D. melanogaster UMP-CMP kinase (Dm.UMP-CMP kinase) to further study the nucleotide metabolizing enzymes in these cells. The kinase, encoded by the dak1 gene, was approximately 60% similar to the human UMP-CMP kinase and predominantly phosphorylated CMP, dCMP, and UMP. Expression analysis showed that the Dm.UMP-CMP kinase mRNA was constitutively expressed throughout the Drosophila development. The open-reading frame of the Dm.UMP-CMP kinase cDNA was extended in the 5'-region compared to UMP-CMP kinases of other species. The extended region encoded an N-terminal sequence with properties characteristic of a mitochondrial import signal. Expression of the enzyme in fusion with the green fluorescent protein verified that the N-terminal signal targeted the enzyme to mitochondria. This is the first time a mitochondrial pyrimidine nucleoside monophosphate kinase has been cloned from any organism and we discuss the implication of this finding for deoxyribonucleoside salvage in both Drosophila and other organisms.

  • substrate specificity and phosphorylation of antiviral and anticancer nucleoside analogues by human deoxyribonucleoside kinases and ribonucleoside kinases
    Pharmacology & Therapeutics, 2003
    Co-Authors: An R. Van Rompay, Magnus Johansson, Anna Karlsson
    Abstract:

    Structural analogues of nucleosides, nucleoside analogues (NA), are used in the treatment of cancer and viral infections. Antiviral NAs inhibit replication of the viral genome, whereas anticancer NAs inhibit cellular DNA replication and repair. NAs are inactive prodrugs that are dependent on intracellular phosphorylation to their pharmacologically active triphosphate form. The deoxyribonucleoside kinases (dNK) and ribonucleoside kinases (rNK) catalyze the first phosphorylation step, converting Deoxyribonucleosides and ribonucleosides to their corresponding monophosphate form. The dNKs have been studied intensively, whereas the rNKs have not been as thoroughly investigated. This overview is focused on the substrate specificity, tissue distribution, and subcellular location of the mammalian dNKs and rNKs and their role in the activation of NAs.

  • cloning and characterization of the multisubstrate deoxyribonucleoside kinase of drosophila melanogaster
    Journal of Biological Chemistry, 1999
    Co-Authors: Magnus Johansson, An R Van Rompay, B Degreve, Jan Balzarini, Anna Karlsson
    Abstract:

    A Drosophila melanogaster deoxyribonucleoside kinase (Dm-dNK) was reported to phosphorylate all four natural Deoxyribonucleosides as well as several nucleoside analogs (Munch-Petersen, B., Piskur, J., and Sondergaard, L. (1998) J. Biol. Chem. 273, 3926–3931). The broad substrate specificity of this enzyme together with a high catalytic rate makes it unique among the nucleoside kinases. We have in the present study cloned the Dm-dNK cDNA, expressed the 29-kDa protein in Escherichia coli, and characterized the recombinant enzyme for the phosphorylation of nucleosides and clinically important nucleoside analogs. The recombinant enzyme preferentially phosphorylated the pyrimidine nucleosides dThd, dCyd, and dUrd, but phosphorylation of the purine nucleosides dAdo and dGuo was also efficiently catalyzed. Dm-dNK is closely related to human and herpes simplex virus deoxyribonucleoside kinases. The highest level of sequence similarity was noted with human mitochondrial thymidine kinase 2, and these enzymes also share many substrates. The cDNA cloning and characterization of Dm-dNK will be the basis for studies on the use of this multisubstrate nucleoside kinase as a suicide gene in combined gene/chemotherapy of cancer.

Wolfgang Knecht - One of the best experts on this subject based on the ideXlab platform.

  • non viral deoxyribonucleoside kinases diversity and practical use
    Journal of Genetics and Genomics, 2015
    Co-Authors: Louise Slot Christiansen, Birgitte Munchpetersen, Wolfgang Knecht
    Abstract:

    Deoxyribonucleoside kinases (dNKs) phosphorylate Deoxyribonucleosides to their corresponding monophosphate compounds. dNks also phosphorylate deoxyribonucleoside analogues that are used in the treatment of cancer or viral infections. The study of the mammalian dNKs has therefore always been of great medical interest. However, during the last 20 years, research on dNKs has gone into non-mammalian organisms. In this review, we focus on non-viral dNKs, in particular their diversity and their practical applications. The diversity of this enzyme family in different organisms has proven to be valuable in studying the evolution of enzymes. Some of these newly discovered enzymes have been useful in numerous practical applications in medicine and biotechnology, and have contributed to our understanding of the structural basis of nucleoside and nucleoside analogue activation.

  • two thymidine kinases and one multisubstrate deoxyribonucleoside kinase salvage dna precursors in arabidopsis thaliana
    FEBS Journal, 2012
    Co-Authors: Anders R Clausen, Wolfgang Knecht, Michael P B Sandrini, Birgitte Munchpetersen, Lenart Girandon, Ashfaq Ali, Elzbieta Rozpedowska, Erik Andreasson, Jure Piskur
    Abstract:

    Deoxyribonucleotides are the building blocks of DNA and can be synthesized via de novo and salvage pathways. Deoxyribonucleoside kinases (dNKs) salvage Deoxyribonucleosides by transfer of a phosphate group to the 5' of a deoxyribonucleoside. This salvage pathway is well characterized in mammals but in contrast little is known about how plants salvage Deoxyribonucleosides. We show that during salvage, Deoxyribonucleosides can be phosphorylated by extracts of Arabidopsis thaliana into corresponding mono-phosphate compounds with a surprising preference for purines over pyrimidines. Deoxyribonucleoside kinase activities were present in all tissues during all growth stages. In the A. thaliana genome we identified two types of genes that could encode enzymes which are involved in the salvage of Deoxyribonucleosides. Thymidine kinase activity was encoded by two thymidine kinase 1-like genes (AtTK1a and AtTK1b) and deoxyadenosine, deoxyguanosine and deoxycytidine kinase activities were encoded by a single AtdNK gene. T-DNA insertion lines of AtTK1a and AtTK1b mutant genes had normal growth, but AtTK1a AtTK1b double mutants died at an early stage, which indicates that AtTK1a and AtTK1b catalyze redundant reactions. Our results point out a crucial role for salvage of thymidine during early plant development. © 2012 The Authors Journal compilation © 2012 FEBS. (Less)

  • animal deoxyribonucleoside kinases forward and retrograde evolution of their substrate specificity
    FEBS Letters, 2004
    Co-Authors: Jure Piskur, Wolfgang Knecht, Michael P B Sandrini, Birgitte Munchpetersen
    Abstract:

    Deoxyribonucleoside kinases, which catalyse the phosphorylation of Deoxyribonucleosides, are present in several copies in most multicellular organisms and therefore represent an excellent model to study gene duplication and specialisation of the duplicated copies through partitioning of substrate specificity. Recent studies suggest that in the animal lineage one of the progenitor kinases, the so-called dCK/dGK/TK2-like gene, was duplicated prior to separation of the insect and mammalian lineages. Thereafter, insects lost all but one kinase, dNK (EC 2.7.1.145), which subsequently, through remodelling of a limited number of amino acid residues, gained a broad substrate specificity.

  • mosquito has a single multisubstrate deoxyribonucleoside kinase characterized by unique substrate specificity
    Nucleic Acids Research, 2003
    Co-Authors: Wolfgang Knecht, Gitte Petersen, Michael P B Sandrini, Leif Sondergaard, Birgitte Munchpetersen, Jure Piskur
    Abstract:

    In mammals four deoxyribonucleoside kinases, with a relatively restricted specificity, catalyze the phosphorylation of the four natural Deoxyribonucleosides. When cultured mosquito cells, originating from the malaria vector Anopheles gambiae, were examined for deoxyribonucleoside kinase activities, only a single enzyme was isolated. Subsequently, the corresponding gene was cloned and over-expressed. While the mosquito kinase (Ag-dNK) phosphorylated all four natural Deoxyribonucleosides, it displayed an unexpectedly higher relative efficiency for the phosphorylation of purine versus pyrimidine Deoxyribonucleosides than the fruit fly multisubstrate deoxyribonucleoside kinase (EC 2.7.1.145). In addition, Ag-dNK could also phosphorylate some medically interesting nucleoside analogs, like stavudine (D4T), 2-chloro-deoxyadenosine (CdA) and 5-bromo-vinyl-deoxyuridine (BVDU). Although the biological significance of multisubstrate deoxyribonucleoside kinases and their diversity among insects remains unclear, the observed variation provides a whole range of applications, as species specific and highly selective targets for insecticides, they have a potential to be used in the enzymatic production of various (di-)(deoxy-)ribonucleoside monophosphates, and as suicide genes in gene therapy.

  • a few amino acid substitutions can convert deoxyribonucleoside kinase specificity from pyrimidines to purines
    The EMBO Journal, 2002
    Co-Authors: Wolfgang Knecht, Kenth Johansson, Hans Eklund, Michael P B Sandrini, Birgitte Munchpetersen, Jure Piskur
    Abstract:

    In mammals, the four native Deoxyribonucleosides are phosphorylated to the corresponding monophosphates by four deoxyribonucleoside kinases, which have specialized substrate specificities. These four enzymes are likely to originate from a common progenitor kinase. Insects appear to have only one multisubstrate deoxyribonucleoside kinase (dNK, EC 2.7.1.145), which prefers pyrimidine nucleosides, but can also phosphorylate purine substrates. When the structures of the human deoxyguanosine kinase (dGK, EC 2.7.1.113) and the dNK from Drosophila melanogaster were compared, a limited number of amino acid residues were identified and proposed to be responsible for the substrate specificity. Three of these key residues in Drosophila dNK were then mutagenized and the mutant enzymes were characterized regarding their ability to phosphorylate native Deoxyribonucleosides and nucleoside analogs. The mutations converted the dNK substrate specificity from predominantly pyrimidine specific into purine specific. A similar scenario could have been followed during the evolution of kinases. Upon gene duplication of the progenitor kinase, only a limited number of single amino acid changes has taken place in each copy and resulted in substrate-specialized enzymes.

Staffan Eriksson - One of the best experts on this subject based on the ideXlab platform.

  • structure and function of cellular deoxyribonucleoside kinases
    Cellular and Molecular Life Sciences, 2002
    Co-Authors: Staffan Eriksson, Kenth Johansson, Birgitte Munchpetersen, Hans Eklund
    Abstract:

    Deoxyribonucleoside kinases phosphorylate Deoxyribonucleosides, a crucial reaction in biosynthesis of DNA precursors through the salvage pathway. Their medical interest stems from their activation of a number of anticancer and antiviral drugs such as 2-chloro-2'-deoxyadenosine, azidothymidine and acyclovir. Here we review what is presently known about each of the mammalian kinases as well as some other members of the deoxyribonucleoside kinase family. A description of the biochemical properties of the enzymes is followed by an overview of the structural studies made on this family of enzymes, including the catalytic mechanism as well as the mechanism for feedback inhibition. A presentation of homology models of other proteins in the family is made and, finally, the determinants of substrate and substrate analog specificities are described.

  • Structural basis for substrate specificities of cellular deoxyribonucleoside kinases.
    Nature structural biology, 2001
    Co-Authors: Kenth Johansson, Staffan Eriksson, S. Ramaswamy, Catarina Ljungcrantz, Wolfgang Knecht, Jure Piskur, Birgitte Munch-petersen, Hans Eklund
    Abstract:

    Deoxyribonucleoside kinases phosphorylate Deoxyribonucleosides and activate a number of medically important nucleoside analogs. Here we report the structure of the Drosophila deoxyribonucleoside kinase with deoxycytidine bound at the nucleoside binding site and that of the human deoxyguanosine kinase with ATP at the nucleoside substrate binding site. Compared to the human kinase, the Drosophila kinase has a wider substrate cleft, which may be responsible for the broad substrate specificity of this enzyme. The human deoxyguanosine kinase is highly specific for purine substrates; this is apparently due to the presence of Arg 118, which provides favorable hydrogen bonding interactions with the substrate. The two new structures provide an explanation for the substrate specificity of cellular deoxyribonucleoside kinases.

  • stereoisomeric selectivity of human deoxyribonucleoside kinases
    Biochemistry, 1999
    Co-Authors: Jianghai Wang, Devapriya Choudhury, Jyoti Chattopadhyaya, Staffan Eriksson
    Abstract:

    Deoxynucleoside kinases catalyze the 5'-phosphorylation of 2'-Deoxyribonucleosides with nucleoside triphosphates as phosphate donors. One of the cellular kinases, deoxycytidine kinase (dCK), has been shown to phosphorylate several L-nucleosides that are efficient antiviral agents. In this study we investigated the potentials of stereoisomers of the natural deoxyribonucleoside to serve as substrates for the recombinant cellular deoxynucleoside kinases. The cytosolic thymidine kinase exhibited a strict selectivity and phosphorylated only ‚-D-Thd, while the mitochondrial thymidine kinase (TK2) and deoxyguanosine kinase (dGK) as well as dCK all had broad substrate specificities. TK2 phosphorylated Thd and dCyd stereoisomers in the order: ‚-D- g ‚-L- .R -D- g R-L-isomer. dCK activated both enantiomers of ‚-dCyd, ‚-dGuo, and ‚-dAdo with similar efficiencies, and R-D-dCyd also served as a substrate. dGK phosphorylated the ‚-dGuo enantiomers with no preference for the ribose configuration; R-L-dGuo was also phosphorylated, and ‚-L-dAdo and ‚-L-dCyd were substrates but showed reduced efficiencies. The anomers of the 2',3'-dideoxy-D-nucleosides (ddNs) were tested, and TK2 and dCK retained their low selectivities. Unexpectedly, R-dideoxycytidine (ddC) was a 3-fold better substrate for dCK than ‚-ddC. Similarly, R-dideoxythymidine (ddT) was a better substrate for TK2 than ‚-ddT. dGK did not accept any D-ddNs. Thus, TK2, dCK, and dGK, similar to herpes simplex virus type 1 thymidine kinase (HSV-1 TK), showed relaxed stereoselectivities, and these results substantiate the functional similarities within this enzyme family. Docking simulations with the Thd isomers and the active site of HSV-1 TK showed that the viral enzyme may in some respects serve as a model for studying the substrate specificities of the cellular enzymes.

  • Mammalian Deoxyribonucleoside Kinases
    Pharmacology & therapeutics, 1995
    Co-Authors: Elias S.j. Arnér, Staffan Eriksson
    Abstract:

    The mammalian deoxyribonucleoside kinases are deoxycytidine kinase, thymidine kinase 1 and 2 and deoxyguanosine kinase. These enzymes phosphorylate Deoxyribonucleosides and thereby provide an alternative to de novo synthesis of DNA precursors. Their activities are essential for the activation of several chemotherapeutically important nucleoside analogues. In recent years, these enzymes have been thoroughly characterised with regard to structure, substrate specificity and patterns of expression. In this review, these results are reviewed and furthermore, the physiologic metabolic role of the anabolic enzymes is discussed in relation to catabolic pathways. The significance of this information for the development of therapeutic protocols and choice of animal model systems is discussed. Finally, alternative pathways for nucleoside analogue phosphorylation are surveyed, such as the phosphotransfer capacity of 5'-nucleotidase.

  • selective assays for thymidine kinase 1 and 2 and deoxycytidine kinase and their activities in extracts from human cells and tissues
    Biochemical and Biophysical Research Communications, 1992
    Co-Authors: Elias S.j. Arnér, Tatjana Spasokoukotskaja, Staffan Eriksson
    Abstract:

    Abstract Human cells salvage pyrimidine Deoxyribonucleosides via 5′-phosphorylation which is also the route of activation of many chemotherapeutically used nucleoside analogs. Key enzymes in this metabolism are the cytosolic thymidine kinase (TK1), the mitochondrial thymidine kinase (TK2) and the cytosolic deoxycytidine kinase (dCK). These enzymes are expressed differently in different tissues and cell cycle phases, and they display overlapping substrate specificities. Thymidine is phosphorylated by both thymidine kinases, and deoxycytidine is phosphorylated by both dCK and TK2. The enzymes also phosphorylate nucleoside analogs with very different efficiencies. Here we present specific radiochemical assays for the three kinase activities utilizing analogs as substrates that are by more than 90 percent phosphorylated solely by one of the kinases; i.e. 3′-azido-2′,3′-dideoxythymidine (AZT) as substrate for TK1, 1-β-D-arabinofura-anosylthymidine (AraT) for TK2 and 2-chlorodeoxyadenosine (CdA) for dCK. We determined the fraction of the total deoxycytidine and thymidine phosphorylating activity that was provided by each of the three enzymes in different human cells and tissues, such as resting and proliferating lymphocytes, lymphocytic cells of leukemia patients (chronic lymphocytic, chronic myeloic and hairy cell leukemia), muscle, brain and gastrointestinal tissue. The detailed knowledge of the pyrimidine deoxyribonucleoside kinase activities and substrate specificities are of importance for studies on chemotherapeutically active nucleoside analogs, and the assays and data presented here should be valuable tools in that research.