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. 2000 Jul 5;97(14):7778–7783. doi: 10.1073/pnas.97.14.7778

Figure 3.

Figure 3

The two-piece Caulobacter tmRNA is functional. (A) Ribosomal association of acceptor, coding, and precursor RNAs. The presence of each species of tmRNA in a crude ribosome preparation was assayed by primer extension on the ribosomal fraction (R) or the nonribosomal supernatant (S). (B) The ribosomal fraction was separated on a sucrose gradient, and the presence of the acceptor RNA and coding RNA in each fraction was assayed by Northern blotting. The fractions containing 70S ribosomes were determined by hybridization with a 16S RNA probe and are marked by the bar. The smearing of RNA bands is likely to be the result of nuclease activity in the ribosomal fraction. (C) Tagging activity of Caulobacter tmRNA. Western blot of a reporter protein expressed from a gene lacking an in-frame stop codon in wild-type Caulobacter (wt) or in Caulobacter with a plasmid-borne copy of SsrA-DD (wt + DD). The molecular masses (MM) indicated at the right were obtained from matrix-assisted laser desorption ionization time-of-flight mass spectrometry, and the corresponding proteins are shown schematically.