The inhibitory mechanism of protein synthesis by YoeB, an escherichia coli toxin

Yonglong Zhang, Masayori Inouye

Research output: Contribution to journalArticle

70 Citations (Scopus)

Abstract

YoeB is a toxin encoded by the yefM-yoeB antitoxin-toxin operon in the Escherichia coli genome. Here we show that YoeB, a highly potent protein synthesis inhibitor, specifically blocks translation initiation. In in vivo primer extension experiments using two different mRNAs, a major band was detected after YoeB induction at three bases downstream of the initiation codon at 2.5 min. An identical band was also detected in in vitro toeprinting experiments after the addition of YoeB to the reaction mixtures containing 70 S ribosomes and the same mRNAs, even in the absence of tRNAfMet. Notably, this band was not detected in the presence of YoeB alone, indicating that YoeB by itself does not have endoribonuclease activity under the conditions used. The 70 S ribosomes increased upon YoeB induction, and YoeB was found to be specifically associated with 50 S subunits. Using tetracycline and hygromycin B, we demonstrated that YoeB binds to the 50 S ribosomal subunit in 70 S ribosomes and interacts with the A site leading to mRNA cleavage at this site. As a result, the 3′-end portion of the mRNA was released from ribosomes, and translation initiation was effectively inhibited. These results demonstrate that YoeB primarily inhibits translation initiation.

Original languageEnglish (US)
Pages (from-to)6627-6638
Number of pages12
JournalJournal of Biological Chemistry
Volume284
Issue number11
DOIs
StatePublished - Mar 13 2009

Fingerprint

Ribosomes
Escherichia coli
Messenger RNA
Proteins
Hygromycin B
Endoribonucleases
Antitoxins
Ribosome Subunits
Protein Synthesis Inhibitors
Initiator Codon
Operon
Tetracycline
Genes
Experiments
Genome

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Biochemistry
  • Cell Biology

Cite this

@article{54aaf1be2def44a583544f19c352b711,
title = "The inhibitory mechanism of protein synthesis by YoeB, an escherichia coli toxin",
abstract = "YoeB is a toxin encoded by the yefM-yoeB antitoxin-toxin operon in the Escherichia coli genome. Here we show that YoeB, a highly potent protein synthesis inhibitor, specifically blocks translation initiation. In in vivo primer extension experiments using two different mRNAs, a major band was detected after YoeB induction at three bases downstream of the initiation codon at 2.5 min. An identical band was also detected in in vitro toeprinting experiments after the addition of YoeB to the reaction mixtures containing 70 S ribosomes and the same mRNAs, even in the absence of tRNAfMet. Notably, this band was not detected in the presence of YoeB alone, indicating that YoeB by itself does not have endoribonuclease activity under the conditions used. The 70 S ribosomes increased upon YoeB induction, and YoeB was found to be specifically associated with 50 S subunits. Using tetracycline and hygromycin B, we demonstrated that YoeB binds to the 50 S ribosomal subunit in 70 S ribosomes and interacts with the A site leading to mRNA cleavage at this site. As a result, the 3′-end portion of the mRNA was released from ribosomes, and translation initiation was effectively inhibited. These results demonstrate that YoeB primarily inhibits translation initiation.",
author = "Yonglong Zhang and Masayori Inouye",
year = "2009",
month = "3",
day = "13",
doi = "https://doi.org/10.1074/jbc.M808779200",
language = "English (US)",
volume = "284",
pages = "6627--6638",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "11",

}

The inhibitory mechanism of protein synthesis by YoeB, an escherichia coli toxin. / Zhang, Yonglong; Inouye, Masayori.

In: Journal of Biological Chemistry, Vol. 284, No. 11, 13.03.2009, p. 6627-6638.

Research output: Contribution to journalArticle

TY - JOUR

T1 - The inhibitory mechanism of protein synthesis by YoeB, an escherichia coli toxin

AU - Zhang, Yonglong

AU - Inouye, Masayori

PY - 2009/3/13

Y1 - 2009/3/13

N2 - YoeB is a toxin encoded by the yefM-yoeB antitoxin-toxin operon in the Escherichia coli genome. Here we show that YoeB, a highly potent protein synthesis inhibitor, specifically blocks translation initiation. In in vivo primer extension experiments using two different mRNAs, a major band was detected after YoeB induction at three bases downstream of the initiation codon at 2.5 min. An identical band was also detected in in vitro toeprinting experiments after the addition of YoeB to the reaction mixtures containing 70 S ribosomes and the same mRNAs, even in the absence of tRNAfMet. Notably, this band was not detected in the presence of YoeB alone, indicating that YoeB by itself does not have endoribonuclease activity under the conditions used. The 70 S ribosomes increased upon YoeB induction, and YoeB was found to be specifically associated with 50 S subunits. Using tetracycline and hygromycin B, we demonstrated that YoeB binds to the 50 S ribosomal subunit in 70 S ribosomes and interacts with the A site leading to mRNA cleavage at this site. As a result, the 3′-end portion of the mRNA was released from ribosomes, and translation initiation was effectively inhibited. These results demonstrate that YoeB primarily inhibits translation initiation.

AB - YoeB is a toxin encoded by the yefM-yoeB antitoxin-toxin operon in the Escherichia coli genome. Here we show that YoeB, a highly potent protein synthesis inhibitor, specifically blocks translation initiation. In in vivo primer extension experiments using two different mRNAs, a major band was detected after YoeB induction at three bases downstream of the initiation codon at 2.5 min. An identical band was also detected in in vitro toeprinting experiments after the addition of YoeB to the reaction mixtures containing 70 S ribosomes and the same mRNAs, even in the absence of tRNAfMet. Notably, this band was not detected in the presence of YoeB alone, indicating that YoeB by itself does not have endoribonuclease activity under the conditions used. The 70 S ribosomes increased upon YoeB induction, and YoeB was found to be specifically associated with 50 S subunits. Using tetracycline and hygromycin B, we demonstrated that YoeB binds to the 50 S ribosomal subunit in 70 S ribosomes and interacts with the A site leading to mRNA cleavage at this site. As a result, the 3′-end portion of the mRNA was released from ribosomes, and translation initiation was effectively inhibited. These results demonstrate that YoeB primarily inhibits translation initiation.

UR - http://www.scopus.com/inward/record.url?scp=65449116514&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=65449116514&partnerID=8YFLogxK

U2 - https://doi.org/10.1074/jbc.M808779200

DO - https://doi.org/10.1074/jbc.M808779200

M3 - Article

C2 - 19124462

VL - 284

SP - 6627

EP - 6638

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 11

ER -