Crystallization and preliminary X-ray diffraction analysis of the Cmr2-Cmr3 subcomplex in the CRISPR-Cas RNA-silencing effector complex

Takuo Osawa, Hideko Inanaga, Tomoyuki Numata

研究成果: ジャーナルへの寄稿記事

1 引用 (Scopus)

抄録

Clustered, regularly interspaced, short palindromic repeat (CRISPR) loci, found in prokaryotes, are transcribed to produce CRISPR RNAs (crRNAs). The Cmr proteins (Cmr1-6) and crRNA form a ribonucleoprotein complex that degrades target RNAs derived from invading genetic elements. Cmr2dHD, a Cmr2 variant lacking the N-terminal putative HD nuclease domain, and Cmr3 were co-expressed in Escherichia coli cells and co-purified as a complex. The Cmr2dHD-Cmr3 complex was co-crystallized with 3′-AMP by the vapour-diffusion method. The crystals diffracted to 2.6 Å resolution using synchrotron radiation at the Photon Factory. The crystals belonged to the orthorhombic space group I222, with unit-cell parameters a = 103.9, b = 136.7, c = 192.0 Å. The asymmetric unit of the crystals is expected to contain one Cmr2dHD-Cmr3 complex with a Matthews coefficient of 3.0 Å 3 Da -1 and a solvent content of 59%.

元の言語英語
ページ(範囲)585-587
ページ数3
ジャーナルActa Crystallographica Section F: Structural Biology and Crystallization Communications
69
発行部数5
DOI
出版物ステータス出版済み - 5 1 2013

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Clustered Regularly Interspaced Short Palindromic Repeats
effectors
RNA Interference
Crystallization
X-Ray Diffraction
X ray diffraction analysis
RNA
crystallization
Crystals
Synchrotrons
Ribonucleoproteins
Adenosine Monophosphate
Photons
diffraction
prokaryotes
nuclease
adenosine monophosphate
crystals
x rays
Radiation

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Genetics
  • Condensed Matter Physics

これを引用

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abstract = "Clustered, regularly interspaced, short palindromic repeat (CRISPR) loci, found in prokaryotes, are transcribed to produce CRISPR RNAs (crRNAs). The Cmr proteins (Cmr1-6) and crRNA form a ribonucleoprotein complex that degrades target RNAs derived from invading genetic elements. Cmr2dHD, a Cmr2 variant lacking the N-terminal putative HD nuclease domain, and Cmr3 were co-expressed in Escherichia coli cells and co-purified as a complex. The Cmr2dHD-Cmr3 complex was co-crystallized with 3′-AMP by the vapour-diffusion method. The crystals diffracted to 2.6 {\AA} resolution using synchrotron radiation at the Photon Factory. The crystals belonged to the orthorhombic space group I222, with unit-cell parameters a = 103.9, b = 136.7, c = 192.0 {\AA}. The asymmetric unit of the crystals is expected to contain one Cmr2dHD-Cmr3 complex with a Matthews coefficient of 3.0 {\AA} 3 Da -1 and a solvent content of 59{\%}.",
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AU - Numata, Tomoyuki

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N2 - Clustered, regularly interspaced, short palindromic repeat (CRISPR) loci, found in prokaryotes, are transcribed to produce CRISPR RNAs (crRNAs). The Cmr proteins (Cmr1-6) and crRNA form a ribonucleoprotein complex that degrades target RNAs derived from invading genetic elements. Cmr2dHD, a Cmr2 variant lacking the N-terminal putative HD nuclease domain, and Cmr3 were co-expressed in Escherichia coli cells and co-purified as a complex. The Cmr2dHD-Cmr3 complex was co-crystallized with 3′-AMP by the vapour-diffusion method. The crystals diffracted to 2.6 Å resolution using synchrotron radiation at the Photon Factory. The crystals belonged to the orthorhombic space group I222, with unit-cell parameters a = 103.9, b = 136.7, c = 192.0 Å. The asymmetric unit of the crystals is expected to contain one Cmr2dHD-Cmr3 complex with a Matthews coefficient of 3.0 Å 3 Da -1 and a solvent content of 59%.

AB - Clustered, regularly interspaced, short palindromic repeat (CRISPR) loci, found in prokaryotes, are transcribed to produce CRISPR RNAs (crRNAs). The Cmr proteins (Cmr1-6) and crRNA form a ribonucleoprotein complex that degrades target RNAs derived from invading genetic elements. Cmr2dHD, a Cmr2 variant lacking the N-terminal putative HD nuclease domain, and Cmr3 were co-expressed in Escherichia coli cells and co-purified as a complex. The Cmr2dHD-Cmr3 complex was co-crystallized with 3′-AMP by the vapour-diffusion method. The crystals diffracted to 2.6 Å resolution using synchrotron radiation at the Photon Factory. The crystals belonged to the orthorhombic space group I222, with unit-cell parameters a = 103.9, b = 136.7, c = 192.0 Å. The asymmetric unit of the crystals is expected to contain one Cmr2dHD-Cmr3 complex with a Matthews coefficient of 3.0 Å 3 Da -1 and a solvent content of 59%.

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