Transmembrane but not soluble helices fold inside the ribosome tunnel

Manuel Bañó-Polo,Anthony Hazel,Ingmarie Nilsson,Carlos Baeza-Delgado,Paul Whitley,Brayan Grau,James C Gumbart,Silvia Tamborero,Ismael Mingarro,Gunnar Von Heijne

doi:10.1038/s41467-018-07554-7

Manuel Bañó-Polo, Anthony Hazel + Show 8 more

Open Access

https://doi.org/10.1038/s41467-018-07554-7

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Abstract

Integral membrane proteins are assembled into the ER membrane via a continuous ribosome-translocon channel. The hydrophobicity and thickness of the core of the membrane bilayer leads to the expectation that transmembrane (TM) segments minimize the cost of harbouring polar polypeptide backbones by adopting a regular pattern of hydrogen bonds to form α-helices before integration. Co-translational folding of nascent chains into an α-helical conformation in the ribosomal tunnel has been demonstrated previously, but the features governing this folding are not well understood. In particular, little is known about what features influence the propensity to acquire α-helical structure in the ribosome. Using in vitro translation of truncated nascent chains trapped within the ribosome tunnel and molecular dynamics simulations, we show that folding in the ribosome is attained for TM helices but not for soluble helices, presumably facilitating SRP (signal recognition particle) recognition and/or a favourable conformation for membrane integration upon translocon entry.

Highlights

Integral membrane proteins are assembled into the ER membrane via a continuous ribosometranslocon channel
For a trapped polypeptide within the ribosome-translocon complex, the number of residues required to bridge the distance between the ribosomal P-site and the active site of the oligosaccharyl transferase (OST) can be conveniently measured by glycosylation mapping[9,14]
There are a number of previous studies demonstrating that the environment of the ribosomal tunnel is permissive for the adoption of α–helical secondary structure of some nascent polypeptide chains[9,11,15,16,33,34] along its entire length, even at the constriction[35] and very close to the ribosomal P-site[36]

Summary

Introduction

Integral membrane proteins are assembled into the ER membrane via a continuous ribosometranslocon channel. Given that a TM segment should be folded prior to its exposure to the lipidic environment for thermodynamic reasons[17,18], we considered that α-helical TM segments might achieve secondary structure in a different location/environment than helices in water-soluble proteins To address this possibility, we used truncated nascent chains trapped within the ribosome-translocon complex of a model protein (E. coli leader peptidase (Lep)) containing engineered ‘test’ sequences of amino acids with known helical propensity in their final folded forms. By translating truncated nascent polypeptide chains of different lengths we observe that test sequences containing TM sequences required a larger number of residues to reach the glycosylation acceptor site in comparison to non-TM sequences This suggests a more compact conformation for nascent polypeptides harboring TM helices, indicating that these helices are formed prior to exit from the ribosome and prior to integration into the lipid bilayer from the translocon pore. The study demonstrates that measured TM helix folding efficiencies are dependent on whether the TM sequence includes helix-breaking or polar residues, as well as on the hydrophobic length of the potential helices

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