NMR study of the phosphate-binding elements of Escherichia coli elongation factor Tu catalytic domain.
The phosphoryl-binding elements in the GDP-binding domain of elongation factor Tu were studied by heteronuclear proton observe methods. Five proton resonances were found below 10.5 ppm. Two of these were assigned to the amide groups of Lys 24 and Gly 83. These are conserved residues in each of the consensus sequences. Their uncharacteristic downfield proton shifts are attributed to strong hydrogen bonds to phosphate oxygens as for resonances in N-ras-p21 [Redfield, A. G., & Papastavros, M. Z. (1990) Biochemistry 29, 3509-3514]. The Lys 24 of the EF-Tu G-domain has nearly the same proton and nitrogen shifts as the corresponding Lys 16 in p21. These results suggest that this conserved lysine has a similar structural role in proteins in this class. The tentative Gly 83 resonance has no spectral analogue in p21. A mutant protein with His 84 changed to glycine was fully 15N-labeled and the proton resonance assigned to Gly 83 shifted downfield by 0.3 ppm, thereby supporting the assignment.
Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02254.
D F
DF
Lowry
R H
RH
Cool
A G
AG
Redfield
A
A
Parmeggiani
D000595:Amino Acid Sequence; D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005998:Glycine; D006153:Guanosine Diphosphate / Q000378:metabolism*; D006860:Hydrogen Bonding; D008239:Lysine; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008969:Molecular Sequence Data; D009587:Nitrogen Isotopes; D010444:Peptide Elongation Factor Tu / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D009587:Nitrogen Isotopes; D011994:Recombinant Proteins; D006153:Guanosine Diphosphate; D010444:Peptide Elongation Factor Tu; D008239:Lysine; D005998:Glycine