Control of translocation through the Sec61 translocon by nascent polypeptide structure within the ribosome

Colin J. Daniel, Brian Conti, Arthur E. Johnson, William Skach

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

During polytopic protein biogenesis, multiple transmembrane segments (TMs) must pass through the ribosome exit tunnel and into the Sec61 translocon prior to insertion into the endoplasmic reticulum membrane. To investigate how movement of a newly synthesized TM along this integration pathway might be influenced by synthesis of a second TM, we used photocross-linking probes to detect the proximity of ribosomebound nascent polypeptides to Sec61α. Probes were inserted at sequential sites within TM2 of the aquaporin-1 water channel by in vitro translation of truncated mRNAs. TM2 first contacted Sec61α when the probe was positioned ∼38 residues from the ribosome peptidyltransferase center, and TM2-Sec61α photoadducts decreased markedly when the probe was >80 residues from the peptidyltransferase center. Unexpectedly, as nascent chain length was gradually extended, photocross-linking at multiple sites within TM2 abruptly and transiently decreased, indicating that TM2 initially entered, withdrew, and then re-entered Sec61. This brief reduction in TM2 photocross-linking coincided with TM3 synthesis. Replacement of TM3 with a secretory reporter domain or introduction of proline residues into TM3 changed the TM2 cross-linking profile and this biphasic behavior. These findings demonstrate that the primary and likely secondary structure of the nascent polypeptide within the ribosome exit tunnel can influence the timing with which topogenic determinants contact, enter, and pass through the translocon.

Original languageEnglish (US)
Pages (from-to)20864-20873
Number of pages10
JournalJournal of Biological Chemistry
Volume283
Issue number30
DOIs
StatePublished - Jul 25 2008

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Ribosomes
Peptidyl Transferases
Peptides
Aquaporin 1
Tunnels
Aquaporins
Protein Biosynthesis
Proline
Endoplasmic Reticulum
Chain length
Membranes
Messenger RNA
Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Control of translocation through the Sec61 translocon by nascent polypeptide structure within the ribosome. / Daniel, Colin J.; Conti, Brian; Johnson, Arthur E.; Skach, William.

In: Journal of Biological Chemistry, Vol. 283, No. 30, 25.07.2008, p. 20864-20873.

Research output: Contribution to journalArticle

Daniel, Colin J. ; Conti, Brian ; Johnson, Arthur E. ; Skach, William. / Control of translocation through the Sec61 translocon by nascent polypeptide structure within the ribosome. In: Journal of Biological Chemistry. 2008 ; Vol. 283, No. 30. pp. 20864-20873.
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