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Three-dimensional structures and properties of a transforming and a nontransforming glycine-12 mutant of p21H-ras.

Authors
  • Franken, S M
  • Scheidig, A J
  • Krengel, U
  • Rensland, H
  • Lautwein, A
  • Geyer, M
  • Scheffzek, K
  • Goody, R S
  • Kalbitzer, H R
  • Pai, E F
Type
Published Article
Journal
Biochemistry
Publication Date
Aug 24, 1993
Volume
32
Issue
33
Pages
8411–8420
Identifiers
PMID: 8357792
Source
Medline
License
Unknown

Abstract

The three-dimensional structures and biochemical properties of two mutants of the G-domain (residues 1-166) of p21H-ras, p21 (G12D) and p21 (G12P), have been determined in the triphosphate-bound form using guanosine 5'-(beta,gamma-imido)triphosphate (GppNHp). They correspond to the most frequent oncogenic and the only nononcogenic mutation of Gly-12, respectively. The G12D mutation is the only mutant analyzed so far that crystallizes in a space group different from wild type, and the atomic model of the protein shows the most drastic changes of structure around the active site as compared to wild-type p21. This is due to the interactions of the aspartic acid side chain with Tyr-32, Gln-61, and the gamma-phosphate, which result in reduced mobility of these structural elements. The interaction between the carboxylate group of Asp-12 and the gamma-phosphate is mediated by a shared proton, which we show by 31P NMR measurements to exist in solution as well. The structure of p21 (G12P) is remarkably similar to that of wild-type p21 in the active site, including the position of the nucleophilic water. The pyrrolidine ring of Pro-12 points outward and seems to be responsible for the weaker affinity toward GAP (GTPase-activating protein) and the failure of GAP to stimulate GTP hydrolysis.

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