Fluorescence titrations, absorption spectroscopy and stopped-flow techniques were used to study the interaction of T4 coded 32-protein (P 32) with MS2 RNA and total tRNA from E. coli under different ionic conditions. It is shown that the amount of MS2 RNA and tRNA secondary structure melted by P 32 varies markedly and reversibly within a range of ionic conditions under which the binding constant of P 32 to single-stranded nucleic acids unable to form stable hairpins remains higher than 10(8) M-1. Kinetic experiments suggest that P 32 dissociates from the MS2 RNA rewinding strand with a similar rate constant as calculated for the dissociation from single-stranded regions. Possible in vivo consequences of these findings are discussed.