title: Effects of macromolecular crowding on an intrinsically disordered protein characterized by small-angle neutron scattering with contrast matching.
abstract: Small-angle neutron scattering was used to examine the effects of molecular crowding on an intrinsically disordered protein, the N protein of bacteriophage λ, 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 D(2)O 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.
authors.affiliation: Department of Biology, University of Utah, Salt Lake City, Utah, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Johansen
authors.forename: Cy M J
authors.initials: CM
authors.lastname: Jeffries
authors.forename: Boualem
authors.initials: B
authors.lastname: Hammouda
authors.forename: Jill
authors.initials: J
authors.lastname: Trewhella
authors.forename: David P
authors.initials: DP
authors.lastname: Goldenberg
mesh_terms: D000818:Animals; D007611:Aprotinin / Q000378:metabolism; D002417:Cattle; D003198:Computer Simulation; D006863:Hydrogen-Ion Concentration; D046911:Macromolecular Substances / Q000378:metabolism*; D008958:Models, Molecular; D033363:Neutron Diffraction; D058767:Protein Unfolding; D053838:Scattering, Small Angle; D012997:Solvents; D054334:Viral Regulatory and Accessory Proteins / Q000737:chemistry / Q000378:metabolism*; D014961:X-Ray Diffraction
chemical_list: D046911:Macromolecular Substances; C059417:N protein, Bacteriophage lambda; D012997:Solvents; D054334:Viral Regulatory and Accessory Proteins; D007611:Aprotinin