Immobilized Drosophila melanogaster Deoxyribonucleoside Kinase (DmdNK) as a High Performing Biocatalyst for the Synthesis of Purine Arabinonucleotides

Immacolata Serra, Silvia Conti, Jure Piskur, Anders Ranegaard Clausen, Birgitte Munch-Petersen, Marco Terreni, Daniela Ubiali

Research output: Contribution to journalArticlepeer-review

22 Citations (SciVal)

Abstract

Fruit fly (Drosophila melanogaster) deoxyribonucleoside kinase (DmdNK; EC: 2.7.1.145) was characterized for its substrate specificity towards natural and non-natural nucleosides, confirming its potential in the enzymatic synthesis of modified nucleotides. DmdNK was adsorbed on a solid ion exchange support (bearing primary amino groups) achieving an expressed activity >98%. Upon cross-linking with aldehyde dextran, expressed activity was 30-40%. Both biocatalysts (adsorbed or cross-linked) were stable at pH10 and room temperature for 24h (about 70% of retained activity). The cross-linked DmdNK preparation was used for the preparative synthesis of arabinosyladenine monophosphate (araA-MP) and fludarabine monophosphate (FaraA-MP). Upon optimization of the reaction conditions (50mM ammonium acetate, substrate/ATP ratio=1:1.25, 2mM MgCl2, 37 degrees C, pH8) immobilized DmdNK afforded the title nucleotides with high conversion (>90%), whereas with the soluble enzyme lower conversions were achieved (78-87%). Arabinosyladenine monophosphate was isolated in 95% yield and high purity (96.5%).
Original languageEnglish
Pages (from-to)563-570
JournalAdvanced Synthesis & Catalysis
Volume356
Issue number2-3
DOIs
Publication statusPublished - 2014

Subject classification (UKÄ)

  • Biological Sciences

Keywords

  • biocatalysis
  • deoxyribonucleoside kinase
  • immobilization
  • nucleotides
  • phosphorylation

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