Interaction of Trigger Factor with the Ribosome

Interplay between trigger factor and other protein biogenesis factors on the ribosome

“…However, those values were obtained by pelleting ribosome-TF complexes, which apparently underestimated affinities, in particular when the complexes were unstable kinetically, such as the complexes with vacant ribosomes or Lep-RNCs. The present parameters of TF-ribosome interaction also differ from published values determined using fluorescence-labelled TF [21][22][23] . In that work, half-life times around 10 s and equilibrium dissociation constants of 1-2 mM were observed for the complexes of labelled TF with non-translating ribosomes and half-life times up to 50 s for translating ribosomes exposing TF-binding nascent chains [21][22][23] .…”

“…The present parameters of TF-ribosome interaction also differ from published values determined using fluorescence-labelled TF [21][22][23] . In that work, half-life times around 10 s and equilibrium dissociation constants of 1-2 mM were observed for the complexes of labelled TF with non-translating ribosomes and half-life times up to 50 s for translating ribosomes exposing TF-binding nascent chains [21][22][23] . However, as we show here, the slow kinetics of complex formation and dissociation as well as low-affinity binding observed in those experiments were caused by the particular fluorescence label, BADAN at position 14, in TF that strongly influenced the properties of TF.…”

“…Furthermore, the TF complex with non-translating ribosomes in our analysis is unstable, with a short half-life time of about 60 ms (k off ¼ 12 s À 1 ). These results are at variance with previous studies on the kinetics of TF-ribosome interaction, obtained by monitoring the fluorescence of a BADAN label at position 14 of TF, which reported very low rates of complex formation and dissociation [21][22][23] . As described in the Supplementary Note, we have repeated the analysis with BADAN-labelled TF and observed similarly low rates ( Supplementary Fig.…”

“…The interplay of TF with other RPBs on the ribosome is not clear as well, as it has been reported that TF binds to and dissociates from ribosomes very slowly [21][22][23] , such that stably bound TF might interfere with the binding of other RPBs.…”

Sight at the end of the tunnel

“…1d). Kinetic studies 10 have suggested that trigger factor binds to the ribosome long enough for an entire chain to be synthesized, although these studies were done in the absence of polypeptide. This would imply either that the nascent chain exits through a side passage (front or back in Fig.…”