ジャーナル: J Mol Biol / 年: 2020 タイトル: Phage G Structure at 6.1 Å Resolution, Condensed DNA, and Host Identity Revision to a Lysinibacillus. 著者: Brenda González / Lyman Monroe / Kunpeng Li / Rui Yan / Elena Wright / Thomas Walter / Daisuke Kihara / Susan T Weintraub / Julie A Thomas / Philip Serwer / Wen Jiang / 要旨: Phage G has the largest capsid and genome of any known propagated phage. Many aspects of its structure, assembly, and replication have not been elucidated. Herein, we present the dsDNA-packed and ...Phage G has the largest capsid and genome of any known propagated phage. Many aspects of its structure, assembly, and replication have not been elucidated. Herein, we present the dsDNA-packed and empty phage G capsid at 6.1 and 9 Å resolution, respectively, using cryo-EM for structure determination and mass spectrometry for protein identification. The major capsid protein, gp27, is identified and found to share the HK97-fold universally conserved in all previously solved dsDNA phages. Trimers of the decoration protein, gp26, sit on the 3-fold axes and are thought to enhance the interactions of the hexameric capsomeres of gp27, for other phages encoding decoration proteins. Phage G's decoration protein is longer than what has been reported in other phages, and we suspect the extra interaction surface area helps stabilize the capsid. We identified several additional capsid proteins, including a candidate for the prohead protease responsible for processing gp27. Furthermore, cryo-EM reveals a range of partially full, condensed DNA densities that appear to have no contact with capsid shell. Three analyses confirm that the phage G host is a Lysinibacillus, and not Bacillus megaterium: identity of host proteins in our mass spectrometry analyses, genome sequence of the phage G host, and host range of phage G.
名称: Bacillus virus G / タイプ: virus / ID: 1 / 親要素: 0 / NCBI-ID: 1084719 / 生物種: Bacillus virus G / ウイルスタイプ: VIRION / ウイルス・単離状態: SPECIES / ウイルス・エンベロープ: No / ウイルス・中空状態: No
ウイルス殻
Shell ID: 1 / 直径: 1600.0 Å / T番号(三角分割数): 52
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実験情報
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構造解析
手法
クライオ電子顕微鏡法
解析
単粒子再構成法
試料の集合状態
particle
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試料調製
緩衝液
pH: 7.4 詳細: 0.01 M Tris-Cl (pH 7.4), 0.01 M MgSO4, 6% polyethylene glycol MW 3350
凍結
凍結剤: ETHANE / チャンバー内湿度: 65 % / チャンバー内温度: 298 K / 装置: GATAN CRYOPLUNGE 3 詳細: Three microliters of phage G sample was deposited on a 400 mesh Ted Pella ultrathin lacey carbon grid and incubated for 30 minutes in a humid chamber on ice. The grid was then washed with 10 ...詳細: Three microliters of phage G sample was deposited on a 400 mesh Ted Pella ultrathin lacey carbon grid and incubated for 30 minutes in a humid chamber on ice. The grid was then washed with 10 microliters of 0.2 TM buffer. Using a Gatan CP3 plunger, the grid was then blotted for 9 seconds with Whatman 1 filter paper at 65% humidity, then plunge-frozen in liquid ethane. The plunge-frozen, phage G grid was then imaged using a Titan Krios equipped with a K2 detector in super-resolution mode at the Purdue Cryo-EM Facility with a nominal magnification of 8,700 resulting in 1.742 Angstroms per pixel. A total of 375 movies was collected..