Vibrational structure of GDP and GTP bound to RAS

An isotope-edited FTIR study

Research output: Contribution to journalArticle

42 Citations (Scopus)

Abstract

A complete vibrational description of the bonding of a ligand to a protein requires the assignment of both symmetric and antisymmetric vibrational modes. The symmetric modes of isotopically enriched enzyme-bound ligands can be obtained by Raman difference spectroscopy, but until now, the antisymmetric modes, which require IR difference spectroscopy, have not been generally accessible. We have developed the methodology needed to perform IR difference spectroscopy, assign the antisymmetric modes, and accurately describe bonding. The method is used to assess the bonding changes that occur as Mg·GDP and Mg·GTP move from solution into the active site of RAS. Binding to RAS opens the nonbridging, O-P-O angle of the γ-phosphate of GTP by 2.7°, yet the angular freedom (dispersion of the O-P-O angle) of the γ-phosphate is comparable to that in solution. In contrast, the motion of the β-phosphate of GDP is highly restricted, suggesting that it positions the γ-phosphate for nucleophilic attack. The β,γbridging O-P bond of bound GTP is slightly weakened, being lengthened by 0.005 Å in the active site, corresponding to a bond order decrease of 0.012 valence unit (vu). The observed binding changes are consistent with a RAS-mediated hydrolysis mechanism that parallels that for solution hydrolysis.

Original languageEnglish (US)
Pages (from-to)4035-4043
Number of pages9
JournalBiochemistry
Volume40
Issue number13
DOIs
StatePublished - Apr 3 2001

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Fourier Transform Infrared Spectroscopy
Guanosine Triphosphate
Isotopes
Phosphates
Spectroscopy
Hydrolysis
Catalytic Domain
Spectrum Analysis
Ligands
Raman Spectrum Analysis
Enzymes
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

Vibrational structure of GDP and GTP bound to RAS : An isotope-edited FTIR study. / Cheng, H.; Sukal, S.; Deng, Hua; Leyh, Thomas S.; Callender, Robert.

In: Biochemistry, Vol. 40, No. 13, 03.04.2001, p. 4035-4043.

Research output: Contribution to journalArticle

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