Journal of Biomolecular NMR

, Volume 40, Issue 2, pp 95–106

Refined solution structure of the 82-kDa enzyme malate synthase G from joint NMR and synchrotron SAXS restraints

  • Alexander Grishaev
  • Vitali Tugarinov
  • Lewis E. Kay
  • Jill Trewhella
  • Ad Bax
Article

Abstract

Determination of the accurate three-dimensional structure of large proteins by NMR remains challenging due to a loss in the density of experimental restraints resulting from the often prerequisite perdeuteration. Solution small-angle scattering, which carries long-range translational information, presents an opportunity to enhance the structural accuracy of derived models when used in combination with global orientational NMR restraints such as residual dipolar couplings (RDCs) and residual chemical shift anisotropies (RCSAs). We have quantified the improvements in accuracy that can be obtained using this strategy for the 82 kDa enzyme Malate Synthase G (MSG), currently the largest single chain protein solved by solution NMR. Joint refinement against NMR and scattering data leads to an improvement in structural accuracy as evidenced by a decrease from ∼4.5 to ∼3.3 Å of the backbone rmsd between the derived model and the high-resolution X-ray structure, PDB code 1D8C. This improvement results primarily from medium-angle scattering data, which encode the overall molecular shape, rather than the lowest angle data that principally determine the radius of gyration and the maximum particle dimension. The effect of the higher angle data, which are dominated by internal density fluctuations, while beneficial, is also found to be relatively small. Our results demonstrate that joint NMR/SAXS refinement can yield significantly improved accuracy in solution structure determination and will be especially well suited for the study of systems with limited NMR restraints such as large proteins, oligonucleotides, or their complexes.

Keywords

NMR Protein structure SAXS RDC Malate synthase G 

Abbreviations

MSG

Malate synthase G

SAXS

Small-angle solution X-ray scattering

RDC

Residual dipolar coupling

RCSA

Residual chemical shift anisotropy

NOE

Nuclear Overhauser enhancement

SVD

Singular value decomposition

RG

Gyration radius

dmax

Maximum particle dimension

Supplementary material

10858_2007_9211_MOESM1_ESM.pdf (223 kb)
(PDF 222 kb)

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Copyright information

© Springer Science+Business Media B.V. 2007

Authors and Affiliations

  • Alexander Grishaev
    • 1
  • Vitali Tugarinov
    • 2
  • Lewis E. Kay
    • 2
  • Jill Trewhella
    • 3
  • Ad Bax
    • 1
  1. 1.Laboratory of Chemical PhysicsNIDDK, National Institutes of HealthBethesdaUSA
  2. 2.Departments of Medical Genetics, Biochemistry and ChemistryUniversity of TorontoTorontoCanada
  3. 3.School of Molecular and Microbial BioscienceUniversity of SydneySydneyAustralia

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