title: Regulation of K-Ras4B Membrane Binding by Calmodulin.
abstract: K-Ras4B is a membrane-bound small GTPase with a prominent role in cancer development. It contains a polybasic farnesylated C-terminus that is required for the correct localization and clustering of K-Ras4B in distinct membrane domains. PDEδ and the Ca(2+)-binding protein calmodulin (CaM) are known to function as potential binding partners for farnesylated Ras proteins. However, they differ in the number of interaction sites with K-Ras4B, leading to different modes of interaction, and thus affect the subcellular distribution of K-Ras4B in different ways. Although it is clear that Ca(2+)-bound CaM can play a role in the dynamic spatial cycle of K-Ras4B in the cell, the exact molecular mechanism is only partially understood. In this biophysical study, we investigated the effect of Ca(2+)/CaM on the interaction of GDP- and GTP-loaded K-Ras4B with heterogeneous model biomembranes by using a combination of different spectroscopic and imaging techniques. The results show that Ca(2+)/CaM is able to extract K-Ras4B from negatively charged membranes in a nucleotide-independent manner. Moreover, the data demonstrate that the complex of Ca(2+)/CaM and K-Ras4B is stable in the presence of anionic membranes and shows no membrane binding. Finally, the influence of Ca(2+)/CaM on the interaction of K-Ras4B with membranes is compared with that of PDEδ, which was investigated in a previous study. Although both CaM and PDEδ exhibit a hydrophobic binding pocket for farnesyl, they have different effects on membrane binding of K-Ras4B and hence should be capable of regulating K-Ras4B plasma membrane localization in the cell.
authors.affiliation: Physical Chemistry I - Biophysical Chemistry, TU Dortmund University, Dortmund, Germany.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Sperlich
authors.affiliation: Chemical Biology, TU Dortmund University, Dortmund, Germany; Department of Chemical Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Shobhna
authors.initials: S
authors.lastname: Kapoor
authors.affiliation: Chemical Biology, TU Dortmund University, Dortmund, Germany; Department of Chemical Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
authors.forename: Herbert
authors.initials: H
authors.lastname: Waldmann
authors.affiliation: Physical Chemistry I - Biophysical Chemistry, TU Dortmund University, Dortmund, Germany. Electronic address: roland.winter@tu-dortmund.de.
authors.forename: Roland
authors.initials: R
authors.lastname: Winter
authors.affiliation: Physical Chemistry I - Biophysical Chemistry, TU Dortmund University, Dortmund, Germany. Electronic address: katrin.weise@tu-dortmund.de.
authors.forename: Katrin
authors.initials: K
authors.lastname: Weise
mesh_terms: D000595:Amino Acid Sequence; D002147:Calmodulin / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D016283:Proto-Oncogene Proteins p21(ras) / Q000737:chemistry / Q000378:metabolism*
chemical_list: D002147:Calmodulin; D016283:Proto-Oncogene Proteins p21(ras)