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Journal of Biomolecular NMR

, Volume 31, Issue 4, pp 321–330 | Cite as

A set of HA-detected experiments for measuring scalar and residual dipolar couplings

  • Peter Würtz
  • Kai Fredriksson
  • Perttu Permi
Article

Abstract

A new set of HCACO based three-dimensional NMR experiments for measuring residual dipolar couplings in proteins is presented. Using spin-state selection and editing in three dimensions, the experiments allow accurate measurement of intraresidual \(^{1}D_{\rm C^{\alpha} H^{\alpha}}\), \(^{1}D_{\rm C^{\prime}C^\alpha}\) and \(^{2} D_{\rm C^{\prime}H^\alpha}\) scalar and residual dipolar couplings of 15␣N/13C labeled proteins in D2O and dilute liquid crystals with minimal spectral crowding. The presented experiments are especially suitable for small or medium sized proline-rich proteins, or proteins that require high pH solvent conditions, making 1HN detected experiments unattractive. In addition, the tetrahedral coordination of Cα is superior to the planar peptide bond for determination of local alignments in partially structured polypeptides. For the efficient use of spectrometer time and to avoid complications arising from the varying magnitude of the alignment tensor during relatively long experiments, the \(^{1}D_{\rm C^{\alpha}H^\alpha}\) and \(^{2}D_{\rm C^{\prime}H^\alpha}\) couplings can also be measured simultaneously in an E.COSY like manner with high accuracy. The pulse sequences are balanced for cross-correlation effects and minimized for relaxation losses. The pulse sequences are tested with a sample of 15N/13C human ubiquitin. We find internuclear vector directions determined from the dipolar couplings to have an excellent correlation with those of ubiquitin’s refined solution structure.

Key words

dipolar couplings HCACO NMR proteins scalar couplings spin-state-selection 

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Copyright information

© Springer 2005

Authors and Affiliations

  1. 1.NMR Laboratory, Program in Structural Biology and Biophysics, Institute of BiotechnologyUniversity of HelsinkiHelsinkiFinland

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