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

Research output: Contribution to journalArticle

6 Citations (Scopus)

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.

Original languageEnglish
Pages (from-to)25-33
Number of pages9
JournalJournal of biochemistry
Volume155
Issue number1
DOIs
Publication statusPublished - Jan 27 2014

Fingerprint

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

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

Cite this

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.

In: Journal of biochemistry, Vol. 155, No. 1, 27.01.2014, p. 25-33.

Research output: Contribution to journalArticle

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. In: Journal of biochemistry. 2014 ; Vol. 155, No. 1. pp. 25-33.
@article{27bd777c01dc42f8a767334f0faefd98,
title = "Characterization of the peripheral structures of archaeal RNase P RNA from Pyrococcus horikoshii OT3",
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.",
author = "Toshifumi Ueda and Hiroki Yamaguchi and Mitsuru Miyanoshita and Takashi Nakashima and Yoshimitsu Kakuta and Makoto Kimura",
year = "2014",
month = "1",
day = "27",
doi = "10.1093/jb/mvt092",
language = "English",
volume = "155",
pages = "25--33",
journal = "Journal of Biochemistry",
issn = "0021-924X",
publisher = "Oxford University Press",
number = "1",

}

TY - JOUR

T1 - Characterization of the peripheral structures of archaeal RNase P RNA from Pyrococcus horikoshii OT3

AU - Ueda, Toshifumi

AU - Yamaguchi, Hiroki

AU - Miyanoshita, Mitsuru

AU - Nakashima, Takashi

AU - Kakuta, Yoshimitsu

AU - Kimura, Makoto

PY - 2014/1/27

Y1 - 2014/1/27

N2 - 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.

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.

UR - http://www.scopus.com/inward/record.url?scp=84892738752&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84892738752&partnerID=8YFLogxK

U2 - 10.1093/jb/mvt092

DO - 10.1093/jb/mvt092

M3 - Article

C2 - 24143022

AN - SCOPUS:84892738752

VL - 155

SP - 25

EP - 33

JO - Journal of Biochemistry

JF - Journal of Biochemistry

SN - 0021-924X

IS - 1

ER -