title: The RAS-Effector Interface: Isoform-Specific Differences in the Effector Binding Regions.
abstract: RAS effectors specifically interact with the GTP-bound form of RAS in response to extracellular signals and link them to downstream signaling pathways. The molecular nature of effector interaction by RAS is well-studied but yet still incompletely understood in a comprehensive and systematic way. Here, structure-function relationships in the interaction between different RAS proteins and various effectors were investigated in detail by combining our in vitro data with in silico data. Equilibrium dissociation constants were determined for the binding of HRAS, KRAS, NRAS, RRAS1 and RRAS2 to both the RAS binding (RB) domain of CRAF and PI3Kα, and the RAS association (RA) domain of RASSF5, RALGDS and PLCε, respectively, using fluorescence polarization. An interaction matrix, constructed on the basis of available crystal structures, allowed identification of hotspots as critical determinants for RAS-effector interaction. New insights provided by this study are the dissection of the identified hotspots in five distinct regions (R1 to R5) in spite of high sequence variability not only between, but also within, RB/RA domain-containing effectors proteins. Finally, we propose that intermolecular β-sheet interaction in R1 is a central recognition region while R3 may determine specific contacts of RAS versus RRAS isoforms with effectors.
authors.affiliation: Institute of Biochemistry and Molecular Biology II, Medical Faculty of the Heinrich-Heine University, Düsseldorf, Germany.
authors.forename: Hossein
authors.initials: H
authors.lastname: Nakhaeizadeh
authors.affiliation: Institute of Biochemistry and Molecular Biology II, Medical Faculty of the Heinrich-Heine University, Düsseldorf, Germany.
authors.forename: Ehsan
authors.initials: E
authors.lastname: Amin
authors.affiliation: Institute of Biochemistry and Molecular Biology II, Medical Faculty of the Heinrich-Heine University, Düsseldorf, Germany.
authors.forename: Saeideh
authors.initials: S
authors.lastname: Nakhaei-Rad
authors.affiliation: Institute of Biochemistry and Molecular Biology II, Medical Faculty of the Heinrich-Heine University, Düsseldorf, Germany.
authors.forename: Radovan
authors.initials: R
authors.lastname: Dvorsky
authors.affiliation: Institute of Biochemistry and Molecular Biology II, Medical Faculty of the Heinrich-Heine University, Düsseldorf, Germany.
authors.forename: Mohammad Reza
authors.initials: MR
authors.lastname: Ahmadian
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D051017:Apoptosis Regulatory Proteins; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D058534:Class I Phosphatidylinositol 3-Kinases; D020558:GTP Phosphohydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D020559:Monomeric GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D000072417:Protein Domains; D017433:Protein Structure, Secondary; D016283:Proto-Oncogene Proteins p21(ras) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D020726:ral Guanine Nucleotide Exchange Factor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D058534:Class I Phosphatidylinositol 3-Kinases; D020558:GTP Phosphohydrolases; D008565:Membrane Proteins; D020559:Monomeric GTP-Binding Proteins; D019869:Phosphatidylinositol 3-Kinases; D016283:Proto-Oncogene Proteins p21(ras); D020726:ral Guanine Nucleotide Exchange Factor; C117307:KRAS protein, human; C454042:RASSF5 protein, human; C484760:PIK3CA protein, human; C087360:RRAS2 protein, human; C579846:NRAS protein, human; C501009:HRAS protein, human