Browsing by Author "Grishaev, A"

Now showing 1 - 3 of 3
  • No Thumbnail Available
    Item
    Refined solution structure of the 82-kDa enzyme malate synthase G from joint NMR and synchrotron SAXS restraints
    (Springer, 2008-02) Grishaev, A; Tugarinov, V; Kay, LE; Trewhella, J; Bax, A
    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. © 2008, Springer.
  • Thumbnail Image
    Item
    A round‐robin approach provides a detailed assessment of biomolecular small‐angle scattering data reproducibility and yields consensus curves for benchmarking
    (International Union of Crystallography (IUCr), 2022-11) Trewhella, J; Vachette, P; Bierma, J; Blanchet, Cl; Brookes, E; Chakravarthy, S; Chatzimagas, L; Cleveland, TE; Cowieson, NP; Crossett, B; Duff, AP; Franke, D; Gabel, F; Gillilan, RE; Graewert, MA; Grishaev, A; Guss, JM; Hammel, M; Hopkins, JB; Huang, Q; Hub, JS; Hura, GL; Irving, TC; Jeffries, CM; Jeong, C; Kirby, N; Krueger, S; Martel, A; Matsui, T; Li, N; Pérez, J; Porcar, L; Prangé, T; Rajkovic, I; Rocco, M; Rosenberg, DJ; Ryan, TM; Seifert, S; Sekiguchi, H; Svergun, D; Teixeira, S; Thureau, A; Weiss, TM; Whitten, AE; Wood, K; Zuo, X
    Through an expansive international effort that involved data collection on 12 small-angle X-ray scattering (SAXS) and four small-angle neutron scattering (SANS) instruments, 171 SAXS and 76 SANS measurements for five proteins (ribonuclease A, lysozyme, xylanase, urate oxidase and xylose isomerase) were acquired. From these data, the solvent-subtracted protein scattering profiles were shown to be reproducible, with the caveat that an additive constant adjustment was required to account for small errors in solvent subtraction. Further, the major features of the obtained consensus SAXS data over the q measurement range 0–1 Å−1 are consistent with theoretical prediction. The inherently lower statistical precision for SANS limited the reliably measured q-range to <0.5 Å−1, but within the limits of experimental uncertainties the major features of the consensus SANS data were also consistent with prediction for all five proteins measured in H2O and in D2O. Thus, a foundation set of consensus SAS profiles has been obtained for benchmarking scattering-profile prediction from atomic coordinates. Additionally, two sets of SAXS data measured at different facilities to q > 2.2 Å−1 showed good mutual agreement, affirming that this region has interpretable features for structural modelling. SAS measurements with inline size-exclusion chromatography (SEC) proved to be generally superior for eliminating sample heterogeneity, but with unavoidable sample dilution during column elution, while batch SAS data collected at higher concentrations and for longer times provided superior statistical precision. Careful merging of data measured using inline SEC and batch modes, or low- and high-concentration data from batch measurements, was successful in eliminating small amounts of aggregate or interparticle interference from the scattering while providing improved statistical precision overall for the benchmarking data set. © The Authors published by International Union of Crystallography. Open Access CC-By licence 4.0.
  • No Thumbnail Available
    Item
    Synaptic arrangement of the neuroligin/β-neurexin complex revealed by x-ray and neutron scattering
    (Elsevier (Cell Press), 2007-06) Comoletti, D; Grishaev, A; Whitten, AE; Tsignelny, I; Taylor, P; Trewhella, J
    Neuroligins are postsynaptic cell-adhesion proteins that associate with their presynaptic partners, the neurexins. Using small-angle X-ray scattering, we determined the shapes of the extracellular region of several neuroligin isoforms in solution. We conclude that the neuroligins dimerize via the characteristic four-helix bundle observed in cholinesterases, and that the connecting sequence between the globular lobes of the dimer and the cell membrane is elongated, projecting away from the dimer interface. X-ray scattering and neutron contrast variation data show that two neurexin monomers, separated by 107 A, bind at symmetric locations on opposite sides of the long axis of the neuroligin dimer. Using these data, we developed structural models that delineate the spatial arrangements of different neuroligin domains and their partnering molecules. As mutations of neurexin and neuroligin genes appear to be linked to autism, these models provide a structural framework for understanding altered recognition by these proteins in neurodevelopmental disorders. © 2007, Cell Press