title: Synthesis of fluorophosphate nucleotide analogues and their characterization as tools for ¹⁹F NMR studies.
abstract: To broaden the scope of existing methods based on (19)F nucleotide labeling, we developed a new method for the synthesis of fluorophosphate (oligo)nucleotide analogues containing an O to F substitution at the terminal position of the (oligo)phosphate moiety and evaluated them as tools for (19)F NMR studies. Using three efficient and comprehensive synthetic approaches based on phosphorimidazolide chemistry and tetra-n-butylammonium fluoride, fluoromonophosphate, or fluorophosphate imidazolide as fluorine sources, we prepared over 30 fluorophosphate-containing nucleotides, varying in nucleobase type (A, G, C, U, m(7)G), phosphate chain length (from mono to tetra), and presence of additional phosphate modifications (thio, borano, imido, methylene). Using fluorophosphate imidazolide as fluorophosphorylating reagent for 5'-phosphorylated oligos we also synthesized oligonucleotide 5'-(2-fluorodiphosphates), which are potentially useful as (19)F NMR hybridization probes. The compounds were characterized by (19)F NMR and evaluated as (19)F NMR molecular probes. We found that fluorophosphate nucleotide analogues can be used to monitor activity of enzymes with various specificities and metal ion requirements, including human DcpS enzyme, a therapeutic target for spinal muscular atrophy. The compounds can also serve as reporter ligands for protein binding studies, as exemplified by studying interaction of fluorophosphate mRNA cap analogues with eukaryotic translation initiation factor (eIF4E).
authors: †Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland.
authors: Marek R
authors: MR
authors: Baranowski
authors: †Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland.|§Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097 Warsaw, Poland.
authors: Anna
authors: A
authors: Nowicka
authors: ‡Chemistry Research Laboratory, University of Oxford, 12 Mansfield Road, OX1 3TA Oxford, United Kingdom.
authors: Anna M
authors: AM
authors: Rydzik
authors: †Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland.
authors: Marcin
authors: M
authors: Warminski
authors: †Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland.
authors: Renata
authors: R
authors: Kasprzyk
authors: §Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097 Warsaw, Poland.
authors: Blazej A
authors: BA
authors: Wojtczak
authors: ∥Laboratory of Biological NMR, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawinskiego 5a, 02-106 Warsaw, Poland.
authors: Jacek
authors: J
authors: Wojcik
authors: ‡Chemistry Research Laboratory, University of Oxford, 12 Mansfield Road, OX1 3TA Oxford, United Kingdom.
authors: Timothy D W
authors: TD
authors: Claridge
authors: †Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland.
authors: Joanna
authors: J
authors: Kowalska
authors: §Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097 Warsaw, Poland.
authors: Jacek
authors: J
authors: Jemielity
mesh_terms: D004722:Endoribonucleases / Q000737:chemistry* / Q000494:pharmacology; D039561:Eukaryotic Initiation Factor-4E / Q000737:chemistry*; D005459:Fluorides / Q000138:chemical synthesis* / Q000737:chemistry; D005462:Fluorine Radioisotopes / Q000737:chemistry*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D009134:Muscular Atrophy, Spinal / Q000201:enzymology / Q000378:metabolism; D009711:Nucleotides / Q000138:chemical synthesis* / Q000737:chemistry; D009841:Oligonucleotides / Q000138:chemical synthesis* / Q000737:chemistry; D010710:Phosphates / Q000138:chemical synthesis* / Q000737:chemistry; D011485:Protein Binding; D000644:Quaternary Ammonium Compounds / Q000737:chemistry*; D012333:RNA, Messenger / Q000138:chemical synthesis / Q000737:chemistry*; D013113:Spin Labels / Q000138:chemical synthesis*
chemical_list: D039561:Eukaryotic Initiation Factor-4E; D005462:Fluorine Radioisotopes; D009711:Nucleotides; D009841:Oligonucleotides; D010710:Phosphates; D000644:Quaternary Ammonium Compounds; D012333:RNA, Messenger; D013113:Spin Labels; C012980:fluorophosphate; C009405:tetrabutylammonium; D004722:Endoribonucleases; C485332:DcpS protein, human; D005459:Fluorides