title: Translational Dynamics of Lipidated Ras Proteins in the Presence of Crowding Agents and Compatible Osmolytes.
abstract: Ras proteins are small GTPases and are involved in transmitting signals that control cell growth, differentiation, and proliferation. Since the cell cytoplasm is crowded with different macromolecules, understanding the translational dynamics of Ras proteins in crowded environments is crucial to yielding deeper insight into their reactivity and function. Herein, the translational dynamics of lipidated N-Ras and K-Ras4B is studied in the bulk and in the presence of a macromolecular crowder (Ficoll) and the compatible osmolyte and microcrowder sucrose by fluorescence correlation spectroscopy. The results reveal that N-Ras forms dimers due to the presence of its lipid moiety in the hypervariable region, whereas K-Ras4B remains in its monomeric form in the bulk. Addition of a macromolecular crowding agent gradually favors clustering of the Ras proteins. In 20 wt % Ficoll N-Ras forms trimers and K-Ras4B dimers. Concentrations of sucrose up to 10 wt % foster formation of N-Ras trimers and K-Ras dimers as well. The results can be rationalized in terms of the excluded-volume effect, which enhances the association of the proteins, and, for the higher concentrations, by limited-hydration conditions. The results of this study shed new light on the association state of these proteins in a crowded environment. This is of particular interest for the Ras proteins, because their solution state-monomeric or clustered-influences their membrane-partitioning behavior and their interplay with cytosolic interaction partners.
authors.affiliation: Department of Chemistry and Chemical Biology, Biophysical Chemistry, TU Dortmund University, Otto-Hahn-Strasse 4a, 44227, Dortmund, Germany.
authors.forename: Satyajit
authors.initials: S
authors.lastname: Patra
authors.affiliation: Department of Chemistry and Chemical Biology, Biophysical Chemistry, TU Dortmund University, Otto-Hahn-Strasse 4a, 44227, Dortmund, Germany.
authors.forename: Nelli
authors.initials: N
authors.lastname: Erwin
authors.affiliation: Department of Chemistry and Chemical Biology, Biophysical Chemistry, TU Dortmund University, Otto-Hahn-Strasse 4a, 44227, Dortmund, Germany. roland.winter@tu-dortmund.de.
authors.forename: Roland
authors.initials: R
authors.lastname: Winter
mesh_terms: D004058:Diffusion; D005362:Ficoll / Q000737:chemistry; D057446:Hydrodynamics; D018613:Microscopy, Confocal; D017368:Protein Prenylation; D013050:Spectrometry, Fluorescence; D013395:Sucrose / Q000737:chemistry; D018631:ras Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: D005362:Ficoll; D013395:Sucrose; D018631:ras Proteins
keywords: excluded-volume effect; fluorescence correlation spectroscopy; molecular crowding; proteins; translational dynamics