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<p>NMR spectroscopy of biomolecules provides atomic level information into their structure, dynamics and interactions with their binding partners. However, signal attenuation from line broadening caused by fast relaxation and signal overlap often limits the application of NMR to large macromolecular systems. Here we leverage the slow relaxation properties of <sup>13</sup>C nuclei attached to <sup>19</sup>F in aromatic <sup>19</sup>F–<sup>13</sup>C spin pairs as well as the spin–spin coupling between the fluorinated <sup>13</sup>C nucleus and the hydrogen atom at the <i>meta</i>-position to record two-dimensional <sup>1</sup>H–<sup>13</sup>C<sub>F</sub> correlation spectra with transverse relaxation-optimized spectroscopy selection on <sup>13</sup>C<sub>F</sub>. To accomplish this, we synthesized [4-<sup>19</sup>F<sup>13</sup>C<sup><i>ζ</i></sup>; 3,5-<sup>2</sup>H<sub>2</sub><sup><i>ε</i></sup>] Phe, engineered for optimal relaxation properties, and adapted a residue-specific route to incorporate this residue globally into proteins and a site-specific 4-<sup>19</sup>F Phe encoding strategy. This approach resulted in narrow linewidths for proteins ranging from 30 kDa to 180 kDa, enabling interaction studies with small-molecule ligands without requiring specialized <sup>19</sup>F-compatible probes.</p><p></p>

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Leveraging relaxation-optimized 1H–13CF correlations in 4-19F-phenylalanine as atomic beacons for probing structure and dynamics of large proteins

  • 期刊论文

  • 发表日期：

    2025年5月5日

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  • Andras Boeszoermenyi,
  • Denitsa L. Radeva,
  • Sebastian Schindler,
  • Veronica Valadares,
  • Krishna M. Padmanabha Das,
  • Abhinav Dubey,
  • Thibault Viennet,
  • Max Schmitt,
  • Peter Kast,
  • Vladimir M. Gelev,
  • Nikolay Stoyanov,
  • Nikola Burdzhiev,
  • Ognyan Petrov,
  • Scott Ficarro,
  • Jarred Marto,
  • Ezekiel A. Geffken,
  • Sirano Dhe-Paganon,
  • Hyuk-Soo Seo,
  • Nathan D. Alexander,
  • Richard B. Cooley,
  • Ryan A. Mehl,
  • Helena Kovacs,
  • Clemens Anklin,
  • Wolfgang Bermel,
  • Ilya Kuprov,
  • Koh Takeuchi,
  • Haribabu Arthanari

摘要

NMR spectroscopy of biomolecules provides atomic level information into their structure, dynamics and interactions with their binding partners. However, signal attenuation from line broadening caused by fast relaxation and signal overlap often limits the application of NMR to large macromolecular systems. Here we leverage the slow relaxation properties of 13C nuclei attached to 19F in aromatic 19F–13C spin pairs as well as the spin–spin coupling between the fluorinated 13C nucleus and the hydrogen atom at the meta-position to record two-dimensional 1H–13CF correlation spectra with transverse relaxation-optimized spectroscopy selection on 13CF. To accomplish this, we synthesized [4-19F13Cζ; 3,5-2H2ε] Phe, engineered for optimal relaxation properties, and adapted a residue-specific route to incorporate this residue globally into proteins and a site-specific 4-19F Phe encoding strategy. This approach resulted in narrow linewidths for proteins ranging from 30 kDa to 180 kDa, enabling interaction studies with small-molecule ligands without requiring specialized 19F-compatible probes.