title: Analysis of binding site hot spots on the surface of Ras GTPase.
abstract: We have recently discovered an allosteric switch in Ras, bringing an additional level of complexity to this GTPase whose mutants are involved in nearly 30% of cancers. Upon activation of the allosteric switch, there is a shift in helix 3/loop 7 associated with a disorder to order transition in the active site. Here, we use a combination of multiple solvent crystal structures and computational solvent mapping (FTMap) to determine binding site hot spots in the "off" and "on" allosteric states of the GTP-bound form of H-Ras. Thirteen sites are revealed, expanding possible target sites for ligand binding well beyond the active site. Comparison of FTMaps for the H and K isoforms reveals essentially identical hot spots. Furthermore, using NMR measurements of spin relaxation, we determined that K-Ras exhibits global conformational dynamics very similar to those we previously reported for H-Ras. We thus hypothesize that the global conformational rearrangement serves as a mechanism for allosteric coupling between the effector interface and remote hot spots in all Ras isoforms. At least with respect to the binding sites involving the G domain, H-Ras is an excellent model for K-Ras and probably N-Ras as well. Ras has so far been elusive as a target for drug design. The present work identifies various unexplored hot spots throughout the entire surface of Ras, extending the focus from the disordered active site to well-ordered locations that should be easier to target.
authors.affiliation: Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC 27695, USA.
authors.forename: Greg
authors.initials: G
authors.lastname: Buhrman
authors.forename: Casey
authors.initials: C
authors.lastname: O'Connor
authors.forename: Brandon
authors.initials: B
authors.lastname: Zerbe
authors.forename: Bradley M
authors.initials: BM
authors.lastname: Kearney
authors.forename: Raeanne
authors.initials: R
authors.lastname: Napoleon
authors.forename: Elizaveta A
authors.initials: EA
authors.lastname: Kovrigina
authors.forename: Sandor
authors.initials: S
authors.lastname: Vajda
authors.forename: Dima
authors.initials: D
authors.lastname: Kozakov
authors.forename: Evgenii L
authors.initials: EL
authors.lastname: Kovrigin
authors.forename: Carla
authors.initials: C
authors.lastname: Mattos
mesh_terms: D000495:Allosteric Site; D001665:Binding Sites; D018360:Crystallography, X-Ray; D006160:Guanosine Triphosphate / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D018631:ras Proteins / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D006160:Guanosine Triphosphate; D018631:ras Proteins