The interaction between the bacteriophage Qβ coat protein and its specific binding site on Qβ genomic RNA was characterized by using a nitrocellulose filter binding assay. Qβ coat protein bound to a synthetic 29-nucleotide RNA hairpin with an association constant of 400 μM-1 at 4 °C, 0.2 M ionic strength, pH 6.0. Complex formation had a broad pH optimum centered around pH 6.0 and was favored by both enthalpy and entropy. The salt dependence of Ka revealed that four to five ion pairs may be formed in the complex although approximately 80% of the free energy of complex formation is contributed by nonelec-trostatic interactions. Truncation experiments revealed that coat protein binding required only the presence of a hairpin with an eight base pair stem and a three-base loop. Analysis of the binding properties of hairpin variants showed that the sequence of the stem was not important for coat protein recognition and only one of the three loop residues was essential. A bulged adenosine present in the coat protein binding site was not required for coat protein binding. Qβ coat protein binding specificity is therefore primarily achieved by the structure and not by the sequence of the operator.
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