title: Crowding effects on the temperature and pressure dependent structure, stability and folding kinetics of Staphylococcal Nuclease.
abstract: FT-IR spectroscopic, small-angle X-ray scattering and calorimetric measurements have been applied to explore the effect of the macromolecular crowder agent Ficoll on the temperature- and pressure-dependent stability diagram and folding reaction of the protein Staphylococcal Nuclease (SNase). Additionally, we compare the experimental data with approximate theoretical predictions. We found that temperature- and pressure-induced equilibrium unfolding of SNase is markedly shifted to higher temperatures and pressures in 30 wt% Ficoll solutions. The structure of the unfolded state ensemble does not seem to be strongly influenced in the presence of the crowder. Self-crowding effects have been found to become important at SNase concentrations above 10 wt% only. Our kinetic results show that the folding rate of SNase decreases markedly in the presence of Ficoll. These results indicate that besides the commonly encountered excluded volume effect, other factors need to be considered when assessing confinement effects on protein folding kinetics. Among those, crowder-induced viscosity changes seem to be prominent.
authors.affiliation: TU Dortmund University, Department of Chemistry and Chemical Biology, Physical Chemistry I - Biophysical Chemistry, D-44221 Dortmund, Germany. roland.winter@tu-dortmund.de.
authors.forename: M
authors.initials: M
authors.lastname: Erlkamp
authors.forename: S
authors.initials: S
authors.lastname: Grobelny
authors.forename: R
authors.initials: R
authors.lastname: Winter
mesh_terms: D002152:Calorimetry, Differential Scanning; D005362:Ficoll / Q000737:chemistry; D007700:Kinetics; D008836:Micrococcal Nuclease / Q000737:chemistry* / Q000378:metabolism; D011312:Pressure; D011489:Protein Denaturation; D017510:Protein Folding; D053838:Scattering, Small Angle; D017550:Spectroscopy, Fourier Transform Infrared; D013696:Temperature; D013816:Thermodynamics; D014961:X-Ray Diffraction
chemical_list: D005362:Ficoll; D008836:Micrococcal Nuclease