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Titolo:
Nuclear magnetic resonance and molecular dynamics studies on the interactions of the Ras-binding domain of Raf-1 with wild-type and mutant Ras proteins
Autore:
Terada, T; Ito, Y; Shirouzu, M; Tateno, M; Hashimoto, K; Kigawa, T; Ebisuzaki, T; Takio, K; Shibata, T; Yokoyama, S; Smith, BO; Laue, ED; Cooper, JA;
Indirizzi:
RIKEN, Inst Phys & Chem Res, Cellular Signaling Lab, Wako, Saitama 3510198, RIKEN Wako Saitama Japan 3510198 ar Signaling Lab, Wako, Saitama 3510198, RIKEN,8,nst Phys & Chem Res, Div Biomol Charecterizat, Wako, Saitama 351019 RIKEN Wako Saitama Japan 3510198 omol Charecterizat, Wako, Saitama 351019 RIKEN,,Inst Phys & Chem Res, Cellular & Mol Biol Lab, Wako, Saitama 3510198 RIKEN Wako Saitama Japan 3510198 ar & Mol Biol Lab, Wako, Saitama 3510198 RIKEN, Inst Phys & Chem Res, Sci Computat Lab, Wako, Saitama 3510198, Japan RIKEN Wako Saitama Japan 3510198 mputat Lab, Wako, Saitama 3510198, Japan Univ Tokyo, Grad Sch Sci, Dept Biophys & Biochem, Bunkyo Ku, Tokyo 1130033, Univ Tokyo Tokyo Japan 1130033 phys & Biochem, Bunkyo Ku, Tokyo 1130033, Univ,Cambridge, Dept Biochem, Cambridge Ctr Mol Recognit, Cambridge CB2 1QW Univ Cambridge Cambridge England CB2 1QW Mol Recognit, Cambridge CB2 1QW Fred Hutchinson Canc Res Ctr, Seattle, WA 98109 USA Fred Hutchinson Canc Res Ctr Seattle WA USA 98109 , Seattle, WA 98109 USA
Titolo Testata:
JOURNAL OF MOLECULAR BIOLOGY
fascicolo: 1, volume: 286, anno: 1999,
pagine: 219 - 232
SICI:
0022-2836(19990212)286:1<219:NMRAMD>2.0.ZU;2-#
Fonte:
ISI
Lingua:
ENG
Soggetto:
NUCLEOTIDE DISSOCIATION STIMULATOR; 3-DIMENSIONAL NMR-SPECTROSCOPY; CYSTEINE-RICH REGION; GTP-GAMMA-S; HA-RAS; CONFORMATIONAL CHANGE; SIGNAL-TRANSDUCTION; BACKBONE ASSIGNMENT; TERMINAL REGION; EFFECTOR-REGION;
Keywords:
Ras-binding domain; Ras; Raf-l; nuclear magnetic resonance; molecular dynamics simulation;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
74
Recensione:
Indirizzi per estratti:
Indirizzo: Yokoyama, S RIKEN,itama Phys & Chem Res, Cellular Signaling Lab, 2-1 Hirosawa, Wako, Sa RIKEN 2-1 Hirosawa Wako Saitama Japan 3510198 rosawa, Wako, Sa
Citazione:
T. Terada et al., "Nuclear magnetic resonance and molecular dynamics studies on the interactions of the Ras-binding domain of Raf-1 with wild-type and mutant Ras proteins", J MOL BIOL, 286(1), 1999, pp. 219-232

Abstract

The Ras protein and its homolog, Rap1A, have an identical "effector region" (residues 32-40) preceded by Asp30-Glu31 and Glu30-Lys31, respectively. in the complex of the "Ras-like" E30D/K31E mutant Rap1A with the Ras-bindingdomain (RBD), residues 51-131 of Raf-1, Glu31 in Rap1A forms a tight salt bridge with Lys84 in Raf-1. However, we have recently found that Raf-1 RED binding of Ras is indeed reduced by the E31K mutation, but is not affected by the E31A mutation. Here, the "Rap1A-like" D30E/E31K mutant of Ras was prepared and shown to bind the Raf-1 RED less strongly than wild-type Ras, but slightly more tightly than the E31K mutant. The backbone H-1, C-13, and N-15 magnetic resonances of the Raf-1 RED were assigned in complexes with the wild-type and D30E/E31K mutant Ras proteins in the guanosine 5'-O-(beta,gamma-imidotriphosphate)-bound form. The Lys84 residue in the Raf-1 RED exhibited a large change in chemical shift upon binding wild-type Ras, suggesting that Lys84 interacts with wild-type Ras. The D30E/E31K mutant of Ras caused nearly the same perturbations in Raf-1 chemical shifts, including that of Lys84. We hypothesized that Glu31 in Ras may not be the major salt bridge partner of Lys84 in Raf-1. A molecular dynamics simulation of a model structure of the Raf-1 RBD.Ras.GTP complex suggested that Lys84 in Raf-1 mightinstead form a tight salt bridge with Asp33 in Ras. Consistent with this, the D33A mutation in Ras greatly reduced its Raf-1 RED binding activity. Weconclude that the major salt bridge partner of Lys84 in Raf-1 may be Asp33in Ras. (C) 1999 Academic Press.

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Documento generato il 15/07/20 alle ore 21:12:27