ジャーナル: J Struct Biol / 年: 2013 タイトル: Dynamics in cryo EM reconstructions visualized with maximum-likelihood derived variance maps. 著者: Qiu Wang / Tsutomu Matsui / Tatiana Domitrovic / Yili Zheng / Peter C Doerschuk / John E Johnson / 要旨: CryoEM data capture the dynamic character associated with biological macromolecular assemblies by preserving the various conformations of the individual specimens at the moment of flash freezing. ...CryoEM data capture the dynamic character associated with biological macromolecular assemblies by preserving the various conformations of the individual specimens at the moment of flash freezing. Regions of high variation in the data set are apparent in the image reconstruction due to the poor density that results from the lack of superposition of these regions. These observations are qualitative and, to date, only preliminary efforts have been made to quantitate the heterogeneity in the ensemble of particles that are individually imaged. We developed and tested a quantitative method for simultaneously computing a reconstruction of the particle and a map of the space-varying heterogeneity of the particle based on an entire data set. The method uses a maximum likelihood algorithm that explicitly takes into account the continuous variability from one instance to another instance of the particle. The result describes the heterogeneity of the particle as a variance to be plotted at every voxel of the reconstructed density. The test, employing time resolved data sets of virus maturation, not only recapitulated local variations obtained with difference map analysis, but revealed a remarkable time dependent reduction in the overall particle dynamics that was unobservable with classical methods of analysis.
詳細: Data were collected using the Leginon automated electron microscopy package and data processing was performed using the Appion pipeline.
電子線
加速電圧: 120 kV / 電子線源: FIELD EMISSION GUN
電子光学系
照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD
試料ステージ
試料ホルダーモデル: GATAN LIQUID NITROGEN
実験機器
モデル: Tecnai F20 / 画像提供: FEI Company
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画像解析
詳細
The reconstruction uses a maximum likelihood algorithm that explicitly takes into account the continuous variability from one instance to another instance of the particle.
CTF補正
詳細: Each image
最終 再構成
アルゴリズム: OTHER / 解像度のタイプ: BY AUTHOR / 解像度: 21.0 Å / 解像度の算出法: FSC 0.5 CUT-OFF / 使用した粒子像数: 1200