Nucleic acid binding properties of Escherichia coli ribosomal protein S1. II. Co-operativity and specificity of binding site II.

The following properties characterize the interaction of nucleic acid binding site II of Escherichia coli ribosomal protein S1 with oligo- and polyribonucleotides; all have been determined with site I complexed with oligo- or polydeoxyribonucleotides. (1) The intrinsic binding constant (K) of site II to single-stranded polyribonucleotides is fairly independent of base composition, though cytidinecontaining polymers bind with approximately threefold higher intrinsic affinities than do the comparable adenine-containing species. (2) Poly(rC) is bound to site II co-operatively; the co-operativity parameter (ω) ? 31. Poly(rA) shows no binding co-operativity. The site size (n) for both polyribonucleotides binding at site II is about ten nucleotide residues. (3) The K value for site II is ? 4 × 10⁵m^?1 for poly(rA), and ? 1 × 10⁶m^?1 for poly(rC), in 0.12 m-Na⁺. Unlike site I, the binding affinity of site II increases somewhat with increasing salt concentration, suggesting that phosphate—basic protein residue contacts are not involved. (4) Varying Mg^{2 +} concentration has no effect on K, and changes in the concentration of either Mg²⁺ or Na⁺ do not affect the magnitude of site II co-operativity. (5) Reaction of the exocyclic amino groups of poly (rC) with formaldehyde drastically reduces the affinity of site II for this polynucleotide, while the affinity of poly (rC) for site I is not altered by this treatment. (6) No major sequence specificity of K for site II is found with either homogeneous polynucleotides or the 3′ terminal dodecanucleotide of 16 S ribosomal RNA; we conclude that selectivity of S1 binding via site II depends largely on the presence or absence of base compositiondependent binding co-operativity.The binding properties of site II probably account for the ability of S1 to inhibit translation at high S1 to ribosome ratios (“factor i” activity). Possible mechanisms for the role of S1 protein as a part of the phage Qβ replicase complex and in protein synthesis are discussed in relation to the binding properties of site I and site II.