Two-dimensional 1H NMR of gene 5 protein indicates that only two aromatic rings interact significantly with oligodeoxynucleotide bases.
The interaction of gene 5 protein (G5P) with oligodeoxynucleotides is investigated by 1H NMR methods, principally two-dimensional nuclear Overhauser effect spectroscopy (NOESY). Aromatic resonances of G5P are specifically assigned from crystallographic data, while the low-field resonances of nucleotides are assigned with sequential or other procedures. Chemical shift changes that accompany binding of d(pA)4, d(A)4, d(pT)4, and d(pA)8, combined with specific protein-nucleotide nuclear Overhauser effects (NOEs) obtained from NOESY spectra, suggest that Phe-73 and Tyr-26 are the only aromatic residues that stack significantly with nucleotide bases. Chemical shift data also imply a role for Leu-28, though this has not been confirmed with intermolecular NOEs. Binding of all four oligonucleotides causes marked upfield movements (0.1-0.6 ppm) of G5P NOESY cross peaks belonging to Tyr-26, Leu-28, and Phe-73. Most other G5P spin systems, notably those of Tyr-34 and Tyr-41, do not appear to be significantly affected. In the d(pA)4-G5P complex an intermolecular NOE is observed between Tyr-26 and H1' of Ade-1, while Phe-73 has NOEs with the H2, H8, and H1' protons of Ade-2 and -3. Intramolecular NOEs seem to follow a similar pattern in the partially cooperative d(pA)8-G5P complex, though specific nucleotide resonance assignments are not possible in this case. Binding causes relatively small chemical shift changes for the base resonances in adenylyl nucleotides, suggesting that there is some, but not complete, unstacking of the bases.(ABSTRACT TRUNCATED AT 250 WORDS)
D000595:Amino Acid Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006639:Histidine; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D009838:Oligodeoxyribonucleotides / Q000378:metabolism*; D010649:Phenylalanine; D011487:Protein Conformation; D014443:Tyrosine
D004268:DNA-Binding Proteins; D009838:Oligodeoxyribonucleotides; D014443:Tyrosine; D010649:Phenylalanine; D006639:Histidine