Cotranslational Protein Folding inside the Ribosome Exit Tunnel

Abstract

SummaryAt what point during translation do proteins fold? It is well established that proteins can fold cotranslationally outside the ribosome exit tunnel, whereas studies of folding inside the exit tunnel have so far detected only the formation of helical secondary structure and collapsed or partially structured folding intermediates. Here, using a combination of cotranslational nascent chain force measurements, inter-subunit fluorescence resonance energy transfer studies on single translating ribosomes, molecular dynamics simulations, and cryoelectron microscopy, we show that a small zinc-finger domain protein can fold deep inside the vestibule of the ribosome exit tunnel. Thus, for small protein domains, the ribosome itself can provide the kind of sheltered folding environment that chaperones provide for larger proteins.

Structure and assembly model for the Trypanosoma cruzi 60S ribosomal subunit

“…S12), where the extension of uL22, a component of the peptide exit tunnel (PET), would reach toward ES39L in yeast. The location of ES42L suggests that it may also play a role in the remote regulation of PET by uL22 (22) (Fig. 3C; SI Appendix, Fig.…”

Membrane protein serendipity

“…Even if it's not my own line of research, it's been quite an experience to help bring this new technology to Sweden. And, of course, we couldn't resist the temptation to integrate cryo-EM in some of our own projects: we now use APs to immobilize newly folded protein domains in the ribosome exit tunnel, and solve the structures using cryo-EM (35,36) (Fig. 8).…”

“…The idea was not hard to come by: test the role of positively charged residues for membrane protein topology using Lep, a protein with two transmembrane helices and a short cytoplasmic connecting loop full of Arg and Lys residues. Ross taught me the Kunkel method (40) for sitedirected mutagenesis-laborious and reeking with 32 P-and how to label proteins with 35 S. This not only led to three pairs of blue jeans made radioactive by spilled samples, but eventually to the demonstration that Lep could be "turned on its head" in the membrane by relocating positively charged residues from the cytoplasmic loop to the protein's N terminus (15) (Fig. 5).…”

Molecular simulations of cellular processes

“…The most challenging is the difficulty to simulate large systems represented at full atomistic resolution for timescales longer than a few μs. Simulations of slower processes occurring in a cell, such as protein (mis)folding, macromolecular (dis)assembly, chromosome (de)condensation, and co-translational events, are, therefore, unfeasible with today's resources (Bhattacharya et al 2013;Phillip and Schreiber 2013;Redler et al 2014;Perilla et al 2015;Nilsson et al 2015;Korolev et al 2016;Zheng and Wen 2017), unless coarser-thanatomistic representations are employed (Takada 2012;Trovato and Tozzini 2012).…”