Journal of Biomolecular NMR

, Volume 47, Issue 3, pp 183–194

MQ-HNCO-TROSY for the measurement of scalar and residual dipolar couplings in larger proteins: application to a 557-residue IgFLNa16-21

  • Sampo Mäntylahti
  • Outi Koskela
  • Pengju Jiang
  • Perttu Permi
Article

Abstract

We describe a novel pulse sequence, MQ-HNCO-TROSY, for the measurement of scalar and residual dipolar couplings between amide proton and nitrogen in larger proteins. The experiment utilizes the whole 2TN polarization transfer delay for labeling of 15N chemical shift in a constant time manner, which efficiently doubles the attainable resolution in 15N dimension with respect to the conventional HNCO-TROSY experiment. In addition, the accordion principle is employed for measuring (J + D)NHs, and the multiplet components are selected with the generalized version of the TROSY scheme introduced by Nietlispach (J Biomol NMR 31:161–166, 2005). Therefore, cross peak overlap is diminished while the time period during which the 15N spin is susceptible to fast transverse relaxation associated with the anti-TROSY transition is minimized per attainable resolution unit. The proposed MQ-HNCO-TROSY scheme was employed for measuring RDCs in high molecular weight protein IgFLNa16-21 of 557 residues, resulting in 431 experimental RDCs. Correlations between experimental and back-calculated RDCs in individual domains gave relatively low Q-factors (0.19–0.39), indicative of sufficient accuracy that can be obtained with the proposed MQ-HNCO-TROSY experiment in high molecular weight proteins.

Keywords

Dipolar couplings HNCO NMR Proteins Scalar couplings TROSY 

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Copyright information

© Springer Science+Business Media B.V. 2010

Authors and Affiliations

  • Sampo Mäntylahti
    • 1
  • Outi Koskela
    • 2
  • Pengju Jiang
    • 3
  • Perttu Permi
    • 1
  1. 1.NMR Laboratory, Program in Structural Biology and Biophysics, Institute of Biotechnology/NMR LaboratoryUniversity of HelsinkiHelsinkiFinland
  2. 2.Laboratory of Organic Chemistry, Department of ChemistryUniversity of HelsinkiHelsinkiFinland
  3. 3.Biochemistry DepartmentUniversity of OxfordOxfordUK

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