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

Abstract Fruit fly ( Drosophila melanogaster ) deoxyribonucleoside kinase ( Dm dNK; 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. Dm dNK 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 pH 10 and room temperature for 24 h (about 70% of retained activity). The cross‐linked Dm dNK preparation was used for the preparative synthesis of arabinosyladenine monophosphate (araA‐MP) and fludarabine monophosphate (FaraA‐MP). Upon optimization of the reaction conditions (50 mM ammonium acetate, substrate/ATP ratio=1:1.25, 2 mM MgCl 2 , 37 °C, pH 8) immobilized Dm dNK 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%). magnified image