title: Dynamic and thermodynamic response of the Ras protein Cdc42Hs upon association with the effector domain of PAK3.
abstract: Human cell division cycle protein 42 (Cdc42Hs) is a small, Rho-type guanosine triphosphatase involved in multiple cellular processes through its interactions with downstream effectors. The binding domain of one such effector, the actin cytoskeleton-regulating p21-activated kinase 3, is known as PBD46. Nitrogen-15 backbone and carbon-13 methyl NMR relaxation was measured to investigate the dynamical changes in activated GMPPCP·Cdc42Hs upon PBD46 binding. Changes in internal motion of the Cdc42Hs, as revealed by methyl axis order parameters, were observed not only near the Cdc42Hs-PBD46 interface but also in remote sites on the Cdc42Hs molecule. The binding-induced changes in side-chain dynamics propagate along the long axis of Cdc42Hs away from the site of PBD46 binding with sharp distance dependence. Overall, the binding of the PBD46 effector domain on the dynamics of methyl-bearing side chains of Cdc42Hs results in a modest rigidification, which is estimated to correspond to an unfavorable change in conformational entropy of approximately -10kcalmol(-1) at 298K. A cluster of methyl probes closest to the nucleotide-binding pocket of Cdc42Hs becomes more rigid upon binding of PBD46 and is proposed to slow the catalytic hydrolysis of the γ phosphate moiety. An additional cluster of methyl probes surrounding the guanine ring becomes more flexible on binding of PBD46, presumably facilitating nucleotide exchange mediated by a guanosine exchange factor. In addition, the Rho insert helix, which is located at a site remote from the PBD46 binding interface, shows a significant dynamic response to PBD46 binding.
authors.affiliation: Graduate Group in Biochemistry and Molecular Biophysics, Johnson Research Foundation and Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6059, USA.
authors.forename: Veronica R
authors.initials: VR
authors.lastname: Moorman
authors.affiliation: Graduate Group in Biochemistry and Molecular Biophysics, Johnson Research Foundation and Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6059, USA.
authors.forename: Kathleen G
authors.initials: KG
authors.lastname: Valentine
authors.affiliation: Graduate Group in Biochemistry and Molecular Biophysics, Johnson Research Foundation and Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6059, USA.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Bédard
authors.affiliation: Graduate Group in Biochemistry and Molecular Biophysics, Johnson Research Foundation and Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6059, USA.
authors.forename: Vignesh
authors.initials: V
authors.lastname: Kasinath
authors.affiliation: Graduate Group in Biochemistry and Molecular Biophysics, Johnson Research Foundation and Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6059, USA.
authors.forename: Jakob
authors.initials: J
authors.lastname: Dogan
authors.affiliation: Graduate Group in Biochemistry and Molecular Biophysics, Johnson Research Foundation and Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6059, USA.
authors.forename: Fiona M
authors.initials: FM
authors.lastname: Love
authors.affiliation: Graduate Group in Biochemistry and Molecular Biophysics, Johnson Research Foundation and Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6059, USA. Electronic address: wand@mail.med.upenn.edu.
authors.forename: A Joshua
authors.initials: AJ
authors.lastname: Wand
mesh_terms: D000495:Allosteric Site; D002244:Carbon / Q000737:chemistry; D002384:Catalysis; D016000:Cluster Analysis; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry; D006160:Guanosine Triphosphate / Q000737:chemistry; D006801:Humans; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D009038:Motion; D009584:Nitrogen / Q000737:chemistry; D016011:Normal Distribution; D017434:Protein Structure, Tertiary; D013696:Temperature; D013816:Thermodynamics; D020764:cdc42 GTP-Binding Protein / Q000737:chemistry*; D054462:p21-Activated Kinases / Q000737:chemistry*; D020741:rho GTP-Binding Proteins / Q000737:chemistry*
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D008024:Ligands; D002244:Carbon; D006160:Guanosine Triphosphate; C515911:PAK3 protein, human; D054462:p21-Activated Kinases; D020764:cdc42 GTP-Binding Protein; D020741:rho GTP-Binding Proteins; D009584:Nitrogen
keywords: NMR-derived order parameters; carbon methyl relaxation; distance-dependent dynamical allostery; pico-nanosecond dynamics
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