The structural mechanism of the inhibition of archaeal RelE toxin by its cognate RelB antitoxin

Masaaki Shinohara, Jin Xu Guo, Misako Mori, Takashi Nakashima, Hisanori Takagi, Etsuko Nishimoto, Shoji Yamashita, Kouhei Tsumoto, Yoshimitsu Kakuta, Makoto Kimura

研究成果: ジャーナルへの寄稿記事

6 引用 (Scopus)

抄録

The archaeal toxin, aRelE, in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 inhibits protein synthesis, whereas its cognate antitoxin, aRelB, neutralizes aRelE activity by forming a non-toxic complex, aRelB-aRelE. The structural mechanism whereby aRelB neutralizes aRelE activity was examined by biochemical and biophysical analyses. Overexpression of aRelB with an aRelE mutant (ΔC6), in which the C-terminal residues critical for aRelE activity were deleted, in Escherichia coli allowed a stable complex, aRelB-ΔC6, to be purified. Isothermal titration of aRelE or ΔC6 with aRelB indicated that the association constant (Ka) of wild-type aRelB-aRelE is similar to that of aRelB-ΔC6, demonstrating that aRelB makes little contact with the C-terminal active site of aRelE. Overexpression of deletion mutants of aRelB with aRelE indicated that either the N-terminal (pos 1-27) or C-terminal (pos. 50-67) fragment of aRelB is sufficient to counteract the toxicity of aRelE in E. coli cells and the second α-helix (α2) in aRelB plays a critical role in forming a stable complex with aRelE. The present results demonstrate that aRelB, as expected from its X-ray structure, precludes aRelE from entering the ribosome, wrapping around the molecular surface of aRelE.

元の言語英語
ページ(範囲)346-351
ページ数6
ジャーナルBiochemical and Biophysical Research Communications
400
発行部数3
DOI
出版物ステータス出版済み - 9 1 2010

Fingerprint

Antitoxins
Escherichia coli
Pyrococcus horikoshii
Archaea
Titration
Ribosomes
Toxicity
Catalytic Domain
X-Rays
Association reactions
X rays
Proteins

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

これを引用

The structural mechanism of the inhibition of archaeal RelE toxin by its cognate RelB antitoxin. / Shinohara, Masaaki; Guo, Jin Xu; Mori, Misako; Nakashima, Takashi; Takagi, Hisanori; Nishimoto, Etsuko; Yamashita, Shoji; Tsumoto, Kouhei; Kakuta, Yoshimitsu; Kimura, Makoto.

:: Biochemical and Biophysical Research Communications, 巻 400, 番号 3, 01.09.2010, p. 346-351.

研究成果: ジャーナルへの寄稿記事

Shinohara, Masaaki ; Guo, Jin Xu ; Mori, Misako ; Nakashima, Takashi ; Takagi, Hisanori ; Nishimoto, Etsuko ; Yamashita, Shoji ; Tsumoto, Kouhei ; Kakuta, Yoshimitsu ; Kimura, Makoto. / The structural mechanism of the inhibition of archaeal RelE toxin by its cognate RelB antitoxin. :: Biochemical and Biophysical Research Communications. 2010 ; 巻 400, 番号 3. pp. 346-351.
@article{0681878b5435414cbfbd85b464c497a8,
title = "The structural mechanism of the inhibition of archaeal RelE toxin by its cognate RelB antitoxin",
abstract = "The archaeal toxin, aRelE, in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 inhibits protein synthesis, whereas its cognate antitoxin, aRelB, neutralizes aRelE activity by forming a non-toxic complex, aRelB-aRelE. The structural mechanism whereby aRelB neutralizes aRelE activity was examined by biochemical and biophysical analyses. Overexpression of aRelB with an aRelE mutant (ΔC6), in which the C-terminal residues critical for aRelE activity were deleted, in Escherichia coli allowed a stable complex, aRelB-ΔC6, to be purified. Isothermal titration of aRelE or ΔC6 with aRelB indicated that the association constant (Ka) of wild-type aRelB-aRelE is similar to that of aRelB-ΔC6, demonstrating that aRelB makes little contact with the C-terminal active site of aRelE. Overexpression of deletion mutants of aRelB with aRelE indicated that either the N-terminal (pos 1-27) or C-terminal (pos. 50-67) fragment of aRelB is sufficient to counteract the toxicity of aRelE in E. coli cells and the second α-helix (α2) in aRelB plays a critical role in forming a stable complex with aRelE. The present results demonstrate that aRelB, as expected from its X-ray structure, precludes aRelE from entering the ribosome, wrapping around the molecular surface of aRelE.",
author = "Masaaki Shinohara and Guo, {Jin Xu} and Misako Mori and Takashi Nakashima and Hisanori Takagi and Etsuko Nishimoto and Shoji Yamashita and Kouhei Tsumoto and Yoshimitsu Kakuta and Makoto Kimura",
year = "2010",
month = "9",
day = "1",
doi = "10.1016/j.bbrc.2010.08.061",
language = "English",
volume = "400",
pages = "346--351",
journal = "Biochemical and Biophysical Research Communications",
issn = "0006-291X",
publisher = "Academic Press Inc.",
number = "3",

}

TY - JOUR

T1 - The structural mechanism of the inhibition of archaeal RelE toxin by its cognate RelB antitoxin

AU - Shinohara, Masaaki

AU - Guo, Jin Xu

AU - Mori, Misako

AU - Nakashima, Takashi

AU - Takagi, Hisanori

AU - Nishimoto, Etsuko

AU - Yamashita, Shoji

AU - Tsumoto, Kouhei

AU - Kakuta, Yoshimitsu

AU - Kimura, Makoto

PY - 2010/9/1

Y1 - 2010/9/1

N2 - The archaeal toxin, aRelE, in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 inhibits protein synthesis, whereas its cognate antitoxin, aRelB, neutralizes aRelE activity by forming a non-toxic complex, aRelB-aRelE. The structural mechanism whereby aRelB neutralizes aRelE activity was examined by biochemical and biophysical analyses. Overexpression of aRelB with an aRelE mutant (ΔC6), in which the C-terminal residues critical for aRelE activity were deleted, in Escherichia coli allowed a stable complex, aRelB-ΔC6, to be purified. Isothermal titration of aRelE or ΔC6 with aRelB indicated that the association constant (Ka) of wild-type aRelB-aRelE is similar to that of aRelB-ΔC6, demonstrating that aRelB makes little contact with the C-terminal active site of aRelE. Overexpression of deletion mutants of aRelB with aRelE indicated that either the N-terminal (pos 1-27) or C-terminal (pos. 50-67) fragment of aRelB is sufficient to counteract the toxicity of aRelE in E. coli cells and the second α-helix (α2) in aRelB plays a critical role in forming a stable complex with aRelE. The present results demonstrate that aRelB, as expected from its X-ray structure, precludes aRelE from entering the ribosome, wrapping around the molecular surface of aRelE.

AB - The archaeal toxin, aRelE, in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 inhibits protein synthesis, whereas its cognate antitoxin, aRelB, neutralizes aRelE activity by forming a non-toxic complex, aRelB-aRelE. The structural mechanism whereby aRelB neutralizes aRelE activity was examined by biochemical and biophysical analyses. Overexpression of aRelB with an aRelE mutant (ΔC6), in which the C-terminal residues critical for aRelE activity were deleted, in Escherichia coli allowed a stable complex, aRelB-ΔC6, to be purified. Isothermal titration of aRelE or ΔC6 with aRelB indicated that the association constant (Ka) of wild-type aRelB-aRelE is similar to that of aRelB-ΔC6, demonstrating that aRelB makes little contact with the C-terminal active site of aRelE. Overexpression of deletion mutants of aRelB with aRelE indicated that either the N-terminal (pos 1-27) or C-terminal (pos. 50-67) fragment of aRelB is sufficient to counteract the toxicity of aRelE in E. coli cells and the second α-helix (α2) in aRelB plays a critical role in forming a stable complex with aRelE. The present results demonstrate that aRelB, as expected from its X-ray structure, precludes aRelE from entering the ribosome, wrapping around the molecular surface of aRelE.

UR - http://www.scopus.com/inward/record.url?scp=77956909858&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=77956909858&partnerID=8YFLogxK

U2 - 10.1016/j.bbrc.2010.08.061

DO - 10.1016/j.bbrc.2010.08.061

M3 - Article

VL - 400

SP - 346

EP - 351

JO - Biochemical and Biophysical Research Communications

JF - Biochemical and Biophysical Research Communications

SN - 0006-291X

IS - 3

ER -