Enzymatic synthesis of ribo- and 2′-deoxyribonucleosides from glycofuranosyl phosphates: An approach to facilitate isotopic labeling

Zhang, Wenhui; Turney, T. A.; Surjancev, Ivana; Serianni, Anthony S.

doi:10.1016/j.carres.2017.07.006

Cited by 6 publications

(9 citation statements)

References 66 publications

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“…It is worth noting that Serianni and co-workers have also developed a complementary approach to enzymatically couple nucleobase and ribose sources using four enzymes (Table ). Their method uses hypoxanthine 84 and 1- O -acetyl-2,3,5-tri- O -benzoyl-α- d -ribofuranoside (ATBR) 85 in a Vorbrüggen reaction (detailed in Scheme ) to yield inosine 86 . Then purine nucleoside phosphorylase (PNPase) (EC 2.4.2.1) replaces the hypoxanthine moiety on the C1 position of 86 with a phosphate group to give α- d -ribofuranosyl-1-phosphate sodium salt (αR1P) 87 (Table ).…”

Section: Preparation Of 15n 2h 13c and 19f Isotope-labeled Rnamentioning

confidence: 99%

“…As described in Section and shown in Scheme , the chemo-enzymatic labeling method developed by Dayie and co-workers ,, permits the synthesis of a versatile assortment of rNTPs with atom-specific isotope labels (Table ). While there are other enzymatic methods to generate both atom-specific (e.g., the Gilles-Schramm-Williamson ,− or Serriani methods shown in Schemes and , respectively) and position-specific (e.g., PLOR and the Schwalbe method , ) labels, no other technique offers the versatility and simplicity that is afforded by the Dayie method. Our one-pot chemo-enzymatic approach can produce isotope-labeled purine and pyrimidine rNTPs in a few days and with high yield (75–95%) (Table ).…”

Section: Preparation Of 15n 2h 13c and 19f Isotope-labeled Rnamentioning

confidence: 99%

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Isotope Labels Combined with Solution NMR Spectroscopy Make Visible the Invisible Conformations of Small-to-Large RNAs

2022

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RNA is central to the proper function of cellular processes important for life on earth and implicated in various medical dysfunctions. Yet, RNA structural biology lags significantly behind that of proteins, limiting mechanistic understanding of RNA chemical biology. Fortunately, solution NMR spectroscopy can probe the structural dynamics of RNA in solution at atomic resolution, opening the door to their functional understanding. However, NMR analysis of RNA, with only four unique ribonucleotide building blocks, suffers from spectral crowding and broad linewidths, especially as RNAs grow in size. One effective strategy to overcome these challenges is to introduce NMR-active stable isotopes into RNA. However, traditional uniform labeling methods introduce scalar and dipolar couplings that complicate the implementation and analysis of NMR measurements. This challenge can be circumvented with selective isotope labeling. In this review, we outline the development of labeling technologies and their application to study biologically relevant RNAs and their complexes ranging in size from 5 to 300 kDa by NMR spectroscopy.

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Section: Preparation Of 15n 2h 13c and 19f Isotope-labeled Rnamentioning

confidence: 99%

Section: Preparation Of 15n 2h 13c and 19f Isotope-labeled Rnamentioning

confidence: 99%

Isotope Labels Combined with Solution NMR Spectroscopy Make Visible the Invisible Conformations of Small-to-Large RNAs

2022

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“…It is worth noting that there are other approaches to enzymatically coupling nucleobase and ribose sources. Indeed, the synthetic method of Serianni and co-workers begins with the chemical synthesis of inosine, followed by its phosphorylation by purine nucleoside phosphorylase (PNPase) (EC 2.4.2.1) to give α-D-ribofuranosyl 1-phosphate sodium salt (αR1P) [ 60 ]. The αR1P intermediate is then glycosylated enzymatically to the ribonucleoside which can easily be converted to the desired rNTP.…”

Section: Stable Isotope Labeling Of Rna Building Blocksmentioning

confidence: 99%

Isotope-Labeled RNA Building Blocks for NMR Structure and Dynamics Studies

et al. 2021

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RNA structural research lags behind that of proteins, preventing a robust understanding of RNA functions. NMR spectroscopy is an apt technique for probing the structures and dynamics of RNA molecules in solution at atomic resolution. Still, RNA analysis by NMR suffers from spectral overlap and line broadening, both of which worsen for larger RNAs. Incorporation of stable isotope labels into RNA has provided several solutions to these challenges. In this review, we summarize the benefits and limitations of various methods used to obtain isotope-labeled RNA building blocks and how they are used to prepare isotope-labeled RNA for NMR structure and dynamics studies.

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“…Although this synthetic strategy is well suited for purine nucleoside phosphoramidites, the 2′- O -tBDMS regioselectivity is not applicable to pyrimidines [74]. Virtually any labeled nucleobase can be readily coupled to ribose via commercially available purine nucleoside phosphorylase, as recently demonstrated [75]. Although labeled nucleobase is readily coupled to ribose using phosphorylases, the introduction of labeled ribose remains a challenge.…”

Section: Stable Isotope Labeling Of Rna Phosphoramiditesmentioning

confidence: 99%

Solid-Phase Chemical Synthesis of Stable Isotope-Labeled RNA to Aid Structure and Dynamics Studies by NMR Spectroscopy

2019

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RNA structure and dynamic studies by NMR spectroscopy suffer from chemical shift overlap and line broadening, both of which become worse as RNA size increases. Incorporation of stable isotope labels into RNA has provided several solutions to these limitations. Nevertheless, the only method to circumvent the problem of spectral overlap completely is the solid-phase chemical synthesis of RNA with labeled RNA phosphoramidites. In this review, we summarize the practical aspects of this methodology for NMR spectroscopy studies of RNA. These types of investigations lie at the intersection of chemistry and biophysics and highlight the need for collaborative efforts to tackle the integrative structural biology problems that exist in the RNA world. Finally, examples of RNA structure and dynamic studies using labeled phosphoramidites are highlighted.

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Enzymatic synthesis of ribo- and 2′-deoxyribonucleosides from glycofuranosyl phosphates: An approach to facilitate isotopic labeling

Cited by 6 publications

References 66 publications

Isotope Labels Combined with Solution NMR Spectroscopy Make Visible the Invisible Conformations of Small-to-Large RNAs

Isotope Labels Combined with Solution NMR Spectroscopy Make Visible the Invisible Conformations of Small-to-Large RNAs

Isotope-Labeled RNA Building Blocks for NMR Structure and Dynamics Studies

Solid-Phase Chemical Synthesis of Stable Isotope-Labeled RNA to Aid Structure and Dynamics Studies by NMR Spectroscopy

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