Characterization of the peripheral structures of archaeal RNase P RNA from Pyrococcus horikoshii OT3

Toshifumi Ueda, Hiroki Yamaguchi, Mitsuru Miyanoshita, Takashi Nakashima, 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

Ribonuclease P (RNase P) is a ubiquitous trans-acting ribozyme that processes the 5′ leader sequence of precursor tRNA (pre-tRNA). The secondary structure of RNase P RNA (PhopRNA) in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 has several peripheral stem-loops. In order to investigate their functional role, we prepared six mutants, ΔP1, ΔP3, ΔP8, ΔP9, ΔP12 and ΔP15, in which the stem-loops including helices P1, P3, P8, P9, P12/12.1/12.2 and P15/16 in PhopRNA were individually deleted, respectively, and characterized them with respect to pre-tRNA cleavage activity in the presence of five proteins and also to the ability to form a complex with the proteins. The reconstituted particles containing ΔP3, ΔP8 or ΔP9 retained considerable levels of activity (35-65%), while those containing ΔP1, ΔP12 or ΔP15 had markedly reduced activity (13%). It was further found that the reconstituted particles comprising ΔP3 or ΔP15 lacked PhoPop5 and PhoRpp30, whereas those containing ΔP1, ΔP8, ΔP9 or ΔP12 bound to all five proteins. Since it is known that PhoPop5 functions in a complex with PhoRpp30, the present result suggests that the peripheral stem-loops containing P3 or P15/16 are involved in the structural formation of a catalytic site by interacting with the protein complex PhoPop5-PhoRpp30.

元の言語英語
ページ(範囲)25-33
ページ数9
ジャーナルJournal of Biochemistry
155
発行部数1
DOI
出版物ステータス出版済み - 1 27 2014

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Pyrococcus horikoshii
Ribonuclease P
RNA
RNA Precursors
Proteins
Catalytic RNA
Archaea
Catalytic Domain

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

これを引用

Characterization of the peripheral structures of archaeal RNase P RNA from Pyrococcus horikoshii OT3. / Ueda, Toshifumi; Yamaguchi, Hiroki; Miyanoshita, Mitsuru; Nakashima, Takashi; Kakuta, Yoshimitsu; Kimura, Makoto.

:: Journal of Biochemistry, 巻 155, 番号 1, 27.01.2014, p. 25-33.

研究成果: ジャーナルへの寄稿記事

Ueda, Toshifumi ; Yamaguchi, Hiroki ; Miyanoshita, Mitsuru ; Nakashima, Takashi ; Kakuta, Yoshimitsu ; Kimura, Makoto. / Characterization of the peripheral structures of archaeal RNase P RNA from Pyrococcus horikoshii OT3. :: Journal of Biochemistry. 2014 ; 巻 155, 番号 1. pp. 25-33.
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abstract = "Ribonuclease P (RNase P) is a ubiquitous trans-acting ribozyme that processes the 5′ leader sequence of precursor tRNA (pre-tRNA). The secondary structure of RNase P RNA (PhopRNA) in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 has several peripheral stem-loops. In order to investigate their functional role, we prepared six mutants, ΔP1, ΔP3, ΔP8, ΔP9, ΔP12 and ΔP15, in which the stem-loops including helices P1, P3, P8, P9, P12/12.1/12.2 and P15/16 in PhopRNA were individually deleted, respectively, and characterized them with respect to pre-tRNA cleavage activity in the presence of five proteins and also to the ability to form a complex with the proteins. The reconstituted particles containing ΔP3, ΔP8 or ΔP9 retained considerable levels of activity (35-65{\%}), while those containing ΔP1, ΔP12 or ΔP15 had markedly reduced activity (13{\%}). It was further found that the reconstituted particles comprising ΔP3 or ΔP15 lacked PhoPop5 and PhoRpp30, whereas those containing ΔP1, ΔP8, ΔP9 or ΔP12 bound to all five proteins. Since it is known that PhoPop5 functions in a complex with PhoRpp30, the present result suggests that the peripheral stem-loops containing P3 or P15/16 are involved in the structural formation of a catalytic site by interacting with the protein complex PhoPop5-PhoRpp30.",
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AB - Ribonuclease P (RNase P) is a ubiquitous trans-acting ribozyme that processes the 5′ leader sequence of precursor tRNA (pre-tRNA). The secondary structure of RNase P RNA (PhopRNA) in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 has several peripheral stem-loops. In order to investigate their functional role, we prepared six mutants, ΔP1, ΔP3, ΔP8, ΔP9, ΔP12 and ΔP15, in which the stem-loops including helices P1, P3, P8, P9, P12/12.1/12.2 and P15/16 in PhopRNA were individually deleted, respectively, and characterized them with respect to pre-tRNA cleavage activity in the presence of five proteins and also to the ability to form a complex with the proteins. The reconstituted particles containing ΔP3, ΔP8 or ΔP9 retained considerable levels of activity (35-65%), while those containing ΔP1, ΔP12 or ΔP15 had markedly reduced activity (13%). It was further found that the reconstituted particles comprising ΔP3 or ΔP15 lacked PhoPop5 and PhoRpp30, whereas those containing ΔP1, ΔP8, ΔP9 or ΔP12 bound to all five proteins. Since it is known that PhoPop5 functions in a complex with PhoRpp30, the present result suggests that the peripheral stem-loops containing P3 or P15/16 are involved in the structural formation of a catalytic site by interacting with the protein complex PhoPop5-PhoRpp30.

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