Genetic identification of nascent peptides that induce ribosome stalling

Douglas R. Tanner, Daniel A. Cariello, Christopher Woolstenhulme, Mark A. Broadbent, Allen Buskirk

Research output: Contribution to journalArticle

Abstract

Several nascent peptides stall ribosomes during their own translation in both prokaryotes and eukaryotes. Leader peptides that induce stalling can regulate downstream gene expression. Interestingly, stalling peptides show little sequence similarity and interact with the ribosome through distinct mechanisms. To explore the scope of regulation by stalling peptides and to better understand the mechanism of stalling, we identified and characterized new examples from random libraries. We created a genetic selection that ties the life of Escherichia coli cells to stalling at a specific site. This selection relies on the natural bacterial system that rescues arrested ribosomes. We altered transfer-messenger RNA, a key component of this rescue system, to direct the completion of a necessary protein if and only if stalling occurs. We identified three classes of stalling peptides: C-terminal Pro residues, SecM-like peptides, and the novel stalling sequence FXXYXIWPP. Like the leader peptides SecM and TnaC, the FXXYXIWPP peptide induces stalling efficiently by inhibiting peptidyl transfer. The nascent peptide exit tunnel and peptidyltransferase center are implicated in this stalling event, although mutations in the ribosome affect stalling on SecM and FXXYXIWPP differently. We conclude that ribosome stalling can be caused by numerous sequences and is more common than previously believed.

Original languageEnglish (US)
Pages (from-to)34809-34818
Number of pages10
JournalJournal of Biological Chemistry
Volume284
Issue number50
DOIs
StatePublished - Dec 11 2009
Externally publishedYes

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Ribosomes
Peptides
Protein Sorting Signals
Peptidyl Transferases
Genetic Selection
Transfer RNA
Eukaryota
Gene expression
Escherichia coli
Libraries
Tunnels
Gene Expression
Messenger RNA
Mutation
Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Genetic identification of nascent peptides that induce ribosome stalling. / Tanner, Douglas R.; Cariello, Daniel A.; Woolstenhulme, Christopher; Broadbent, Mark A.; Buskirk, Allen.

In: Journal of Biological Chemistry, Vol. 284, No. 50, 11.12.2009, p. 34809-34818.

Research output: Contribution to journalArticle

Tanner, Douglas R. ; Cariello, Daniel A. ; Woolstenhulme, Christopher ; Broadbent, Mark A. ; Buskirk, Allen. / Genetic identification of nascent peptides that induce ribosome stalling. In: Journal of Biological Chemistry. 2009 ; Vol. 284, No. 50. pp. 34809-34818.
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