FIGURE 5.

Lead(II) acetate cleavages on tmRNA in different ribosomal complexes. Lead(II) cleavage analyses were performed on four different ribosomal complexes (RCI–IV). Primer extensions of naked tmRNA were run on the same gels for comparison. Three different radiolabeled primers (GW-TM2, GW-TM3, and GW-TM5) were used for primer extension analysis, and the lead(II) concentrations for cleavage were 2.5 and 5 mM. Nucleotides U16–U17 (A) and G333–C335 (E), which are protected in the RCs due to interaction with SmpB, are indicated to the right of the autoradiograph. Nucleotides G7 and U10 (A) as well as G58 and U59 (B), which are uniquely cleaved in complexes RCII and RC IV, are marked. A few more nucleotide positions (A185, A197, A294, and A298) (C,D), which show differences in cleavage accessibility between naked tmRNA and tmRNA in different RCs, are also marked. Nucleotides A32 and A34 (A), which are protected in all RCs, are indicated as well. A detailed evaluation of cleavages is shown in Table 1. Asterisks in the figure mark the significantly increased cleavages that are in boldface in Table 1.