Figure 4.

Efficiency of tagging of tmRNA targets. ΔssrA expressing tmRNA-His was used to estimate the fraction of DasR protein, which becomes tagged by the tmRNA-mediated trans-translation mechanism. (A) Western analysis using His antibodies of tmRNA-tagged proteins and of purified EF-Tu1(His)₆ (excess of 10 pmol; control for column capacity). (B) Analysis of tmRNA-tagged fractions using DasR and EF-Tu1 antibodies and compared with Streptomyces coelicolor M600 and its ssrA mutant. The amount of protein loaded for this experiment was equivalent to 100 μl of culture. DasR, development of aerial mycelium and spores regulator; EF, elongation factor; pellet, bound fraction; sup., unbound fraction; tmRNA, transfer–messenger RNA; total, extract before separation.