Nascentome analysis uncovers futile protein synthesis in escherichia coli

Koreaki Ito, Yuhei Chadani, Kenta Nakamori, Shinobu Chiba, Yoshinori Akiyama, Tatsuhiko Abo

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

Although co-translational biological processes attract much attention, no general and easy method has been available to detect cellular nascent polypeptide chains, which we propose to call collectively a "nascentome." We developed a method to selectively detect polypeptide portions of cellular polypeptidyl-tRNAs and used it to study the generality of the quality control reactions that rescue dead-end translation complexes. To detect nascent polypeptides, having their growing ends covalently attached to a tRNA, cellular extracts are separated by SDS-PAGE in two dimensions, first with the peptidyl-tRNA ester bonds preserved and subsequently after their in-gel cleavage. Pulse-labeled nascent polypeptides of Escherichia coli form a characteristic line below the main diagonal line, because each of them had contained a tRNA of nearly uniform size in the first-dimension electrophoresis but not in the second-dimension. The detection of nascent polypeptides, separately from any translation-completed polypeptides or degradation products thereof, allows us to follow their fates to gain deeper insights into protein biogenesis and quality control pathways. It was revealed that polypeptidyl-tRNAs were significantly stabilized in E. coli upon dysfunction of the tmRNA-ArfA ribosome-rescuing system, whose function had only been studied previously using model constructs. Our results suggest that E. coli cells are intrinsically producing aberrant translation products, which are normally eliminated by the ribosome-rescuing mechanisms.

Original languageEnglish
Article numbere28413
JournalPLoS One
Volume6
Issue number12
DOIs
Publication statusPublished - Dec 5 2011

Fingerprint

Escherichia coli
polypeptides
protein synthesis
Transfer RNA
Peptides
translation (genetics)
Proteins
ribosomes
Ribosomes
Quality Control
quality control
Quality control
Biological Phenomena
Electrophoresis
electrophoresis
polyacrylamide gel electrophoresis
Polyacrylamide Gel Electrophoresis
Esters
Gels
esters

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)

Cite this

Nascentome analysis uncovers futile protein synthesis in escherichia coli. / Ito, Koreaki; Chadani, Yuhei; Nakamori, Kenta; Chiba, Shinobu; Akiyama, Yoshinori; Abo, Tatsuhiko.

In: PLoS One, Vol. 6, No. 12, e28413, 05.12.2011.

Research output: Contribution to journalArticle

Ito, K, Chadani, Y, Nakamori, K, Chiba, S, Akiyama, Y & Abo, T 2011, 'Nascentome analysis uncovers futile protein synthesis in escherichia coli', PLoS One, vol. 6, no. 12, e28413. https://doi.org/10.1371/journal.pone.0028413
Ito, Koreaki ; Chadani, Yuhei ; Nakamori, Kenta ; Chiba, Shinobu ; Akiyama, Yoshinori ; Abo, Tatsuhiko. / Nascentome analysis uncovers futile protein synthesis in escherichia coli. In: PLoS One. 2011 ; Vol. 6, No. 12.
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