Dissociation of the K-Ras4B/PDEδ complex upon contact with lipid membranes: membrane delivery instead of extraction.
K-Ras4B is a small GTPase whose selective membrane localization and clustering into microdomains are mediated by its polybasic farnesylated C-terminus. The importance of the subcellular distribution for the signaling activity of K-Ras4B became apparent from recent in vivo studies, showing that the delta subunit of cGMP phosphodiesterase (PDEδ), which possesses a hydrophobic prenyl-binding pocket, is able to function as a potential binding partner for farnesylated proteins, thereby leading to a modulation of the spatiotemporal organization of K-Ras. Even though PDEδ has been suggested to serve as a cytosolic carrier for Ras, the functional transport mechanism still remains largely elusive. In this study, the effect of PDEδ on the interaction of GDP- and GTP-loaded K-Ras4B with neutral and anionic model biomembranes has been investigated by a combination of different spectroscopic and imaging techniques. The results show that PDEδ is not able to extract K-Ras4B from membranes. Rather, the K-Ras4B/PDEδ complex formed in bulk solution turned out to be unstable in the presence of heterogeneous membranes, resulting in a release of farnesylated K-Ras4B upon membrane contact. With the additional observation of enhanced membrane affinity for the K-Ras4B/PDEδ complex, a molecular mechanism for the PDEδ-K-Ras4B-membrane interaction could be proposed. This includes an effective delivery of PDEδ-solubilized K-Ras4B to the plasma membrane, probably through cytoplasmic diffusion, the dissociation of the K-Ras4B/PDEδ complex upon plasma membrane contact, and finally the membrane binding of released farnesylated K-Ras4B that leads to K-Ras4B-enriched microdomain formation.
Physical Chemistry I-Biophysical Chemistry, Faculty of Chemistry, TU Dortmund University, Otto-Hahn-Strasse 6, D-44227 Dortmund, Germany.
Katrin
K
Weise
Shobhna
S
Kapoor
Alexander
A
Werkmüller
Simone
S
Möbitz
Gunther
G
Zimmermann
Gemma
G
Triola
Herbert
H
Waldmann
Roland
R
Winter
D054703:Cyclic Nucleotide Phosphodiesterases, Type 4 / Q000737:chemistry*; D011905:Genes, ras; D007700:Kinetics; D008563:Membrane Lipids / Q000737:chemistry*; D008958:Models, Molecular
D008563:Membrane Lipids; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4