ジャーナル: Proc Natl Acad Sci U S A / 年: 2011 タイトル: RNA polymerase and transcription elongation factor Spt4/5 complex structure. 著者: Brianna J Klein / Daniel Bose / Kevin J Baker / Zahirah M Yusoff / Xiaodong Zhang / Katsuhiko S Murakami / 要旨: Spt4/5 in archaea and eukaryote and its bacterial homolog NusG is the only elongation factor conserved in all three domains of life and plays many key roles in cotranscriptional regulation and in ...Spt4/5 in archaea and eukaryote and its bacterial homolog NusG is the only elongation factor conserved in all three domains of life and plays many key roles in cotranscriptional regulation and in recruiting other factors to the elongating RNA polymerase. Here, we present the crystal structure of Spt4/5 as well as the structure of RNA polymerase-Spt4/5 complex using cryoelectron microscopy reconstruction and single particle analysis. The Spt4/5 binds in the middle of RNA polymerase claw and encloses the DNA, reminiscent of the DNA polymerase clamp and ring helicases. The transcription elongation complex model reveals that the Spt4/5 is an upstream DNA holder and contacts the nontemplate DNA in the transcription bubble. These structures reveal that the cellular RNA polymerases also use a strategy of encircling DNA to enhance its processivity as commonly observed for many nucleic acid processing enzymes including DNA polymerases and helicases.
温度: 295 K / 手法: 蒸気拡散法, シッティングドロップ法 / pH: 5.5 詳細: 0.1 M Bis-Tris (pH 5.5), 0.2 M NaCl, and 25% PEG3350, VAPOR DIFFUSION, SITTING DROP, temperature 295K
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データ収集
放射光源
由来: シンクロトロン / サイト: CHESS / ビームライン: F1 / 波長: 0.9181 Å
検出器
タイプ: ADSC QUANTUM 270 / 検出器: CCD / 日付: 2009年11月9日
放射
プロトコル: SINGLE WAVELENGTH / 単色(M)・ラウエ(L): M / 散乱光タイプ: x-ray
放射波長
波長: 0.9181 Å / 相対比: 1
反射
解像度: 1.8→30 Å / Num. obs: 44007
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解析
ソフトウェア
名称
バージョン
分類
PHASER
位相決定
REFMAC
5.5.0072
精密化
HKL-2000
データ削減
HKL-2000
データスケーリング
精密化
構造決定の手法: 分子置換 / 解像度: 1.8→30 Å / Cor.coef. Fo:Fc: 0.945 / Cor.coef. Fo:Fc free: 0.911 / SU B: 4.424 / SU ML: 0.132 / 交差検証法: THROUGHOUT / ESU R Free: 0.154 / 立体化学のターゲット値: MAXIMUM LIKELIHOOD / 詳細: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
Rfactor
反射数
%反射
Selection details
Rfree
0.29234
2208
5 %
RANDOM
Rwork
0.23644
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-
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obs
0.23928
41656
98.13 %
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溶媒の処理
イオンプローブ半径: 0.8 Å / 減衰半径: 0.8 Å / VDWプローブ半径: 1.4 Å / 溶媒モデル: MASK