Structure and dynamics of a ribosome-bound nascent chain by NMR spectroscopy

*Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, United Kingdom;
^†AG-Ribosome, Max-Planck-Institute for Molecular Genetics, Ihnestrasse 73, D-14195 Berlin, Germany; and
^‡Institut für Organische Chemie und Chemische Biologie, Johann Wolfgang Goethe-Universitaet Frankfurt am Main, Max-von-Laue-Strasse 7, D-60438 Frankfurt am Main, Germany

Edited by Adriaan Bax, National Institutes of Health, Bethesda, MD, and approved August 15, 2007 (received for review May 17, 2007)

Abstract

Protein folding in living cells is inherently coupled to protein synthesis and chain elongation. There is considerable evidence that some nascent chains fold into their native structures in a cotranslational manner before release from the ribosome, but, despite its importance, a detailed description of such a process at the atomic level remains elusive. We show here at a residue-specific level that a nascent protein chain can reach its native tertiary structure on the ribosome. By generating translation-arrested ribosomes in which the newly synthesized polypeptide chain is selectively ¹³C/¹⁵N-labeled, we observe, using ultrafast NMR techniques, a large number of resonances of a ribosome-bound nascent chain complex corresponding to a pair of C-terminally truncated immunoglobulin (Ig) domains. Analysis of these spectra reveals that the nascent chain adopts a structure in which a native-like N-terminal Ig domain is tethered to the ribosome by a largely unfolded and highly flexible C-terminal domain. Selective broadening of resonances for a group of residues that are colocalized in the structure demonstrates that there are specific but transient interactions between the ribosome and the N-terminal region of the folded Ig domain. These findings represent a step toward a detailed structural understanding of the cellular processes of cotranslational folding.

cotranslational folding

Footnotes

^§To whom correspondence may be addressed. E-mail: fucini{at}chemie-uni.frankfurt.de, cmd44{at}cam.ac.uk, or jc370{at}cam.ac.uk
Author contributions: S.-T.D.H., P.F., C.M.D., and J.C. designed research; S.-T.D.H., P.F., L.D.C., H.L., and J.C. performed research; S.-T.D.H., P.F., C.M.D., and J.C. analyzed data; and S.-T.D.H., P.F., C.M.D., and J.C. wrote the paper.
↵ ^¶Present address: Institute of Structural Molecular Biology, Department of Biochemistry and Molecular Biology, University College London, and School of Crystallography, Birkbeck College, Gower Street, London WC1E 6BT, U.K.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
This article contains supporting information online at www.pnas.org/cgi/content/full/0704664104/DC1.
Abbreviations:

CTD,

C-terminal domain;

NTD,

N-terminal domain;

RNC,

ribosome-bound nascent chain complex.