Browsing by Author "Hammouda, B"

Now showing 1 - 2 of 2
  • No Thumbnail Available
    Item
    Effects of macromolecular crowding on an intrinsically disordered protein characterized by small-angle neutron scattering with contrast matching
    (Elsevier (Cell Press), 2011-02-16) Johansen, D; Jeffries, CM; Hammouda, B; Trewhella, J; Goldenberg, DP
    Small-angle neutron scattering was used to examine the effects of molecular crowding on an intrinsically disordered protein, the N protein of bacteriophage lambda, in the presence of high concentrations of a small globular protein, bovine pancreatic trypsin inhibitor (BPTI). The N protein was labeled with deuterium, and the D2O concentration of the solvent was adjusted to eliminate the scattering contrast between the solvent and unlabeled BPTI, leaving only the scattering signal from the unfolded protein. The scattering profile observed in the absence of BPTI closely matched that predicted for an ensemble of random conformations. With BPTI added to a concentration of 65 mg/mL, there was a clear change in the scattering profile representing an increase in the mass fractal dimension of the unfolded protein, from 1.7 to 1.9, as expected if crowding favors more compact conformations. The crowding protein also inhibited aggregation of the unfolded protein. At 130 mg/mL BPTI, however, the fractal dimension was not significantly different from that measured at the lower concentration, contrary to the predictions of models that treat the unfolded conformations as convex particles. These results are reminiscent of the behavior of polymers in concentrated melts, suggesting that these synthetic mixtures may provide useful insights into the properties of unfolded proteins under crowding conditions. © 2011, Cell Press
  • No Thumbnail Available
    Item
    Structure of the KinA-Sda complex suggests an allosteric mechanism of histidine kinase inhibition
    (Elsevier, 2007-04-27) Whitten, AE; Jacques, DA; Hammouda, B; Hanley, TL; King, GF; Guss, JM; Trewhella, J; Langley, DB
    The Bacillus subtilis histidine kinase KinA controls activation of the transcription factor governing sporulation, SpoOA. The decision to sporulate involves KinA phosphorylating itself on a conserved histidine residue, after which the phosphate moiety is relayed via two other proteins to SpoOA. The DNA-damage checkpoint inhibitor Sda halts this pathway by binding KinA and blocking the autokinase reaction. We have performed small-angle X-ray scattering and neutron contrast variation studies on the complex formed by KinA and Sda. The data show that two Sda molecules bind to the base of the DHp dimerization domain of the KinA dimer. In this position Sda does riot appear to be able to sterically block the catalytic domain from accessing its target histidine, as previously proposed, but rather may effect an allosteric mode of inhibition involving transmission of the inhibitory signal via the four-helix bundle that forms the DHp domain. © 2007, Elsevier Ltd.