Real-time observation of trigger factor function on translating ribosomes

Abstract

The contribution of co-translational chaperone functions to protein folding is poorly understood. Ribosome-associated trigger factor (TF) is the first molecular chaperone encountered by nascent polypeptides in bacteria. Here we show, using fluorescence spectroscopy to monitor TF function and structural rearrangements in real time, that TF interacts with ribosomes and translating polypeptides in a dynamic reaction cycle. Ribosome binding stabilizes TF in an open, activated conformation. Activated TF departs from the ribosome after a mean residence time of approximately 10 s, but may remain associated with the elongating nascent chain for up to 35 s, allowing entry of a new TF molecule at the ribosome docking site. The duration of nascent-chain interaction correlates with the occurrence of hydrophobic motifs in translating polypeptides, reflecting a high aggregation propensity. These findings can explain how TF prevents misfolding events during translation and may provide a paradigm for the regulation of nucleotide-independent chaperones.

Kinetic modelling indicates that fast-translating codons can coordinate cotranslational protein folding by avoiding misfolded intermediates

“…Thus, even for domains exhibiting the simplest possible folding behaviour on the ribosome, slow-translating codons can in certain circumstances affect cotranslational behaviour in different ways depending on the context. We note in addition the interesting result that molecular chaperones, such as trigger factor 46 , can serve as a cause of non-monotonic changes in domain stability 47 because of their binding to the unfolded ensemble 46 .…”

Firefly luciferase mutants as sensors of proteome stress

“…Ribosome Associated Chaperones translational folding (Agashe et al, 2004;Kaiser et al, 2006;Kramer et al, 2009). This provides sufficient time to the elongating polypeptide to receive its entire structural information requisite for the productive folding to begin.…”

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

“…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.…”