MOLECULAR DISSECTION OF THE PSEUDOKNOT GOVERNING THE TRANSLATIONAL REGULATION OF ESCHERICHIA COLI RIBOSOMAL PROTEIN S15

Abstract The ribosomal proteinS15 controls its own translation by binding to a mRNA region overlapping the ribosome binding site. That region of the mRNA can fold in two mutually exclusive conformations that are in dynamic equilibrium: a structure with two hairpins and a pseudoknot. A mutational analysis provided evidence for the existence and requirement of the pseudoknotfor translational control in vivo and S15 recognition in vitro. In this study, we used chemical probingto analyze the structural consequences of mutations and their effect on the stem-loop/ pseudoknotequilibrium. Interactions between S15 and the pseudoknotstructure were further investigated by footprinting experiments. These data, combined with computer modelling and the previously published data on S15 binding and in vivo control, provide important clues on pseudoknotformation and S15 recognition. An unexpected result is that the relevant control element, here the pseudoknotform, can exist in a variety of topologically equivalent structures recognizable and shapable by S15. S15 sits on the deep groove of the co-axial stack and makes contacts with both stems, shielding the bridging adenine. The only specific sequence determinantsare found in the helix common to the pseudoknotand the hairpin structures.