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1H, 15N, 13C backbone resonance assignment of human Alkbh5

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Abstract

N6-methyladenosine (m6A) is the most abundant and reversible post-transcriptional modification in eukaryotic mRNA and long non-coding RNA (lncRNA). The central role of m6A in various physiological processes has generated considerable biological and pharmacological interest. Alkbh5 (AlkB homologue 5) belongs to the AlkB family and is a non-heme Fe(II)/α-ketoglutarate-dependent dioxygenase that selectively catalyzes the oxidative demethylation of m6A. Herein, we report the backbone 1H, 15N, 13C chemical shift assignment of a fully active, 26 kDa construct of human Alkbh5. Experiments were acquired at 25 °C by heteronuclear multidimensional NMR spectroscopy. Collectively, 92% of all backbone resonances were assigned, with 195 out of a possible 212 residues assigned in the 1H–15N TROSY spectrum. Using the program TALOS+, a secondary structure prediction was generated from the assigned backbone resonance that is consistent with the previously reported X-ray structure of the enzyme. The reported assignment will permit investigations of the protein structural dynamics anticipated to provide crucial insight regarding fundamental aspects in the recognition and enzyme regulation processes.

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Acknowledgements

This work was supported by startup funding from Iowa State University (V.V.)

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Authors and Affiliations

  1. Department of Chemistry, Iowa State University, Hach Hall, 2438 Pammel Drive, Ames, IA, 50011, USA

    Jeffrey A. Purslow & Vincenzo Venditti

  2. Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, IA, 50011, USA

    Vincenzo Venditti

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  1. Jeffrey A. Purslow
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Purslow, J.A., Venditti, V. 1H, 15N, 13C backbone resonance assignment of human Alkbh5. Biomol NMR Assign 12, 297–301 (2018). https://doi.org/10.1007/s12104-018-9826-3

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