SsrA-mediated trans-translation plays a role in mRNA quality control by facilitating degradation of truncated mRNAs

Western blot analysis of His₆-CRP proteins derived from the wild-type and terminatorless crp genes. (A) Schematic drawing of the wild-type and terminatorless crp genes encoding His₆-CRP in pYY100 and pYY101. The elevated arrow denotes the start and direction of transcription. The hatched and open rectangles represent the coding regions for 6 × His and CRP, respectively. The figure is not drawn to scale. The ρ-independent crp terminator is shown by horizontal arrows. (B) Western blot analysis of His₆-CRP proteins. The His₆-CRP proteins were expressed in three different strains, TA331 (ssrA⁻), W3110 (ssrA⁺), and TA371 (ssrA^DD), harboring pYY100 and pYY101. W3110 (ssrA⁺) and TA371 (ssrA^DD) express the wild-type SsrA RNA (AA) and SsrA^DD RNA (DD), respectively. The proteins were purified by Ni²⁺-NTA affinity chromatography. The purified proteins of 1 μg were subjected to Western blot analysis using anti-CRP antibodies (lanes 1–6) and anti-DD-tag antibodies (lanes 7–12). The position of the full-length His₆ CRP (24.4 kD) is shown by an asterisk. Size standards are on the right.