Enzymatic production of non-natural nucleoside-5′-monophosphates by a thermostable uracil phosphoribosyltransferase

Del Arco, JonAcosta, JavierPereira, Humberto M.Perona Requena, AlmudenaLokanath, Neratur K.Kunishima, NaokiFernández Lucas, Jesús2024-11-052024-11-052018del Arco, J., Acosta, J., Pereira, H. M., Perona, A., Lokanath, N. K., Kunishima, N., & Fernández-Lucas, J. (2018). Enzymatic Production of Non-Natural Nucleoside-5′-Monophosphates by a Thermostable Uracil Phosphoribosyltransferase. ChemCatChem, 10(2), 439-448. https://doi.org/10.1002/CCTC.2017012231867-388010.1002/cctc.201701223https://hdl.handle.net/20.500.14352/110029The use of enzymes as biocatalysts applied to synthesis ofmodified nucleoside-5’-monophosphates (NMPs) is an interest-ing alternative to traditional multistep chemical methodswhich offers several advantages, such as stereo-, regio-, andenantioselectivity, simple downstream processing, and mild re-action conditions. Herein we report the recombinant expres-sion, production, and purification of uracil phosphoribosyl-transferase from Thermus themophilus HB8 (TtUPRT). The struc-ture of TtUPRT has been determined by protein crystallogra-phy, and its substrate specificity and biochemical characteris-tics have been analyzed, providing new structural insights intothe substrate-binding mode. Biochemical characterization ofthe recombinant protein indicates that the enzyme is a homo-tetramer, with activity and stability across a broad range oftemperatures (50–80 8C), pH (5.5–9) and ionic strength (0–500 mm NaCl). Surprisingly, TtUPRT is able to recognize several5 and 6-substituted pyrimidines as substrates. These experi-mental results suggest TtUPRT could be a valuable biocatalystfor the synthesis of modified NMPs.engEnzymatic production of non-natural nucleoside-5′-monophosphates by a thermostable uracil phosphoribosyltransferasejournal article1867-3899https://doi.org/10.1002/cctc.201701223https://chemistry-europe.onlinelibrary.wiley.com/doi/epdf/10.1002/cctc.201701223restricted access577.113.3577.15577.2579.6BiocatalysisBiological activityEnzymesNucleotidesStructure elucidationBioquímica (Biología)Biología molecular (Química)Microbiología (Biología)2403 Bioquímica2415 Biología Molecular2414 Microbiología2302.09 Enzimología2302.23 Ácidos Nucleicos