Circular dichroism study of Escherichia coli initiation factor 3 binding to nucleic acids.
The circular dichroic spectral features of (A)10-20, (C)10-20, A8UGU6, poly(A), and poly(C), at both neutral and acidic pH values and in the presence and absence of Mg2+, are significantly altered by Escherichia coli initiation factor 3 (IF3), implying the occurrence of protein-induced changes in nucleic acid secondary structure. Similarly, the circular dichroic spectral characteristics of helical poly(U), poly-(A)-poly(U), and poly(I)-poly(C) are modified by IF3. However, no structural perturbation of poly(A)-poly(U) occurs in the absence of Mg2+ by IF3. The oligonucleotides (A)10-20 and (C)10-20 at both pH 7.5 and 5.5 titrate to end point of 26 +/- 4 nucleotide residues per IF3 [except (C) 10-20 at pH 5.5 which titrates to 17 +/- 1 nucleotide residues per IF3], whereas the hairpin A8UGU6 under similar conditions at neutral pH and in the presence of Mg2+ titrates to an end point of 56 +/- 3 nucleotide residues per IF3, thereby suggesting the presence of multiple binding sites on the protein. By contrast, poly(A) and poly(C) at neutral pH and in the absence of Mg2+ titrate to an end point of 13 +/- 1 nucleotide residues per IF3. The occurrence of significant light-scattering artifacts precluded a determination of the end point stoichiometry in most other cases. The circular dichroic spectra of E. coli tRNA, MS2 RNA, phiX174 DNA, and sonicated calf thymus DNA were unaffected by IF3 at physiological concentrations. Addition of an equimolar mixture of IF3 and ribosomal protein S1 titrates the circular dichroism of poly(C) at acid pH as did S1 alone. However, addition of IF3 to mixture of poly(A) and S1 at neutral pH did not result in significant titration of the optical activity until IF3 was in excess over S1, even though filter binding assays indicate normal IF3 binding to the polynucleotide. The possible relation of these observations to the biological function of IF3 is briefly considered.
D002942:Circular Dichroism; D004926:Escherichia coli / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008274:Magnesium; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000378:metabolism*; D009843:Oligoribonucleotides / Q000378:metabolism*; D010448:Peptide Initiation Factors / Q000378:metabolism*; D011131:Polyribonucleotides / Q000378:metabolism*; D039665:Prokaryotic Initiation Factor-3; D013329:Structure-Activity Relationship
D009841:Oligonucleotides; D009843:Oligoribonucleotides; D010448:Peptide Initiation Factors; D011131:Polyribonucleotides; D039665:Prokaryotic Initiation Factor-3; D008274:Magnesium