An isotope edited classical Raman difference spectroscopic study of the interactions of guanine nucleotides with elongation factor Tu and H-ras p21

D. Manor, G. Weng, Hua Deng, Robert Callender, C. X. Chen, V. Balogh-Nair, K. Delaria, F. Jurnak, R. Callender

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

We have measured the Raman spectrum of GDP bound to the elongation factor protein, EF-Tu, and the c-Harvey-ras protein, p21, two proteins of the guanine nucleotide binding family. In order to separate the Raman spectrum of the nucleotide from the much more intense protein spectrum, we investigate the feasibility of ''tagging'' the normal modes of the nucleotide by isotopic substitution, here by incorporating deuterium-labeled guanine at the C8 position into the active site. A difference spectrum between the labeled and unlabeled protein-nucleotide complex shows the changes in the Raman spectrum of the bound nucleotide that arise from the isotopic exchange. We find that surprisingly good Raman spectra of bound ligands can be obtained with this method and that the method can be easily generalized to other systems. The data show that the guanine amino group of the nucleotide interacts differently with both EF-Tu and p21 than it does with water, showing a change in hydrogen-bonding properties upon binding. On the other hand, no change in hydrogen bonding is observed at guanine's N7. The data strongly suggest that the conformation of the nucleotide when bound to EF-Tu and that p21 is the C2' endo pucker of the ribose ring and anti about the glycosidic bond. These results are compared to previous structural and chemical studies.

Original languageEnglish (US)
Pages (from-to)10914-10920
Number of pages7
JournalBiochemistry
Volume30
Issue number45
StatePublished - 1991
Externally publishedYes

Fingerprint

Peptide Elongation Factor Tu
Proto-Oncogene Proteins p21(ras)
Guanine Nucleotides
Isotopes
Nucleotides
Raman scattering
Guanine
Hydrogen Bonding
Hydrogen bonds
Proteins
Peptide Elongation Factors
ras Proteins
Ribose
Deuterium
Conformations
Catalytic Domain
Substitution reactions
Ligands
Water

ASJC Scopus subject areas

  • Biochemistry

Cite this

An isotope edited classical Raman difference spectroscopic study of the interactions of guanine nucleotides with elongation factor Tu and H-ras p21. / Manor, D.; Weng, G.; Deng, Hua; Callender, Robert; Chen, C. X.; Balogh-Nair, V.; Delaria, K.; Jurnak, F.; Callender, R.

In: Biochemistry, Vol. 30, No. 45, 1991, p. 10914-10920.

Research output: Contribution to journalArticle

Manor, D, Weng, G, Deng, H, Callender, R, Chen, CX, Balogh-Nair, V, Delaria, K, Jurnak, F & Callender, R 1991, 'An isotope edited classical Raman difference spectroscopic study of the interactions of guanine nucleotides with elongation factor Tu and H-ras p21', Biochemistry, vol. 30, no. 45, pp. 10914-10920.
Manor, D. ; Weng, G. ; Deng, Hua ; Callender, Robert ; Chen, C. X. ; Balogh-Nair, V. ; Delaria, K. ; Jurnak, F. ; Callender, R. / An isotope edited classical Raman difference spectroscopic study of the interactions of guanine nucleotides with elongation factor Tu and H-ras p21. In: Biochemistry. 1991 ; Vol. 30, No. 45. pp. 10914-10920.
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