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J Biol Chem. 2014 Jul 11;289(28):19294-305. doi: 10.1074/jbc.M114.563239. Epub 2014 May 7.

Signal recognition particle-ribosome binding is sensitive to nascent chain length.

Author information

1
From the Howard Hughes Medical Institute, Department of Biochemistry and Biophysics, University of California at San Francisco, San Francisco, California 94158.
2
the Department of Structural Biology, Stanford University School of Medicine, Stanford, California 94305, and the Department of Applied Physics, Stanford University, Stanford, California 94305.
3
the Department of Structural Biology, Stanford University School of Medicine, Stanford, California 94305, and.
4
the Department of Structural Biology, Stanford University School of Medicine, Stanford, California 94305, and puglisi@stanford.edu.
5
From the Howard Hughes Medical Institute, Department of Biochemistry and Biophysics, University of California at San Francisco, San Francisco, California 94158, peter@walterlab.ucsf.edu.

Abstract

The signal recognition particle (SRP) directs ribosome-nascent chain complexes (RNCs) displaying signal sequences to protein translocation channels in the plasma membrane of prokaryotes and endoplasmic reticulum of eukaryotes. It was initially proposed that SRP binds the signal sequence when it emerges from an RNC and that successful binding becomes impaired as translation extends the nascent chain, moving the signal sequence away from SRP on the ribosomal surface. Later studies drew this simple model into question, proposing that SRP binding is unaffected by nascent chain length. Here, we reinvestigate this issue using two novel and independent fluorescence resonance energy transfer assays. We show that the arrival and dissociation rates of SRP binding to RNCs vary according to nascent chain length, resulting in the highest affinity shortly after a functional signal sequence emerges from the ribosome. Moreover, we show that SRP binds RNCs in multiple and interconverting conformations, and that conversely, RNCs exist in two conformations distinguished by SRP interaction kinetics.

KEYWORDS:

Fluorescence Resonance Energy Transfer (FRET); Protein Targeting; Ribosome Function; Signal Recognition Particle (SRP); Single-Molecule Biophysics

PMID:
24808175
PMCID:
PMC4094042
DOI:
10.1074/jbc.M114.563239
[Indexed for MEDLINE]
Free PMC Article
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