ジャーナル: Nature / 年: 2015 タイトル: Structure of the immature HIV-1 capsid in intact virus particles at 8.8 Å resolution. 著者: Florian K M Schur / Wim J H Hagen / Michaela Rumlová / Tomáš Ruml / Barbara Müller / Hans-Georg Kräusslich / John A G Briggs / 要旨: Human immunodeficiency virus type 1 (HIV-1) assembly proceeds in two stages. First, the 55 kilodalton viral Gag polyprotein assembles into a hexameric protein lattice at the plasma membrane of the ...Human immunodeficiency virus type 1 (HIV-1) assembly proceeds in two stages. First, the 55 kilodalton viral Gag polyprotein assembles into a hexameric protein lattice at the plasma membrane of the infected cell, inducing budding and release of an immature particle. Second, Gag is cleaved by the viral protease, leading to internal rearrangement of the virus into the mature, infectious form. Immature and mature HIV-1 particles are heterogeneous in size and morphology, preventing high-resolution analysis of their protein arrangement in situ by conventional structural biology methods. Here we apply cryo-electron tomography and sub-tomogram averaging methods to resolve the structure of the capsid lattice within intact immature HIV-1 particles at subnanometre resolution, allowing unambiguous positioning of all α-helices. The resulting model reveals tertiary and quaternary structural interactions that mediate HIV-1 assembly. Strikingly, these interactions differ from those predicted by the current model based on in vitro-assembled arrays of Gag-derived proteins from Mason-Pfizer monkey virus. To validate this difference, we solve the structure of the capsid lattice within intact immature Mason-Pfizer monkey virus particles. Comparison with the immature HIV-1 structure reveals that retroviral capsid proteins, while having conserved tertiary structures, adopt different quaternary arrangements during virus assembly. The approach demonstrated here should be applicable to determine structures of other proteins at subnanometre resolution within heterogeneous environments.
ダウンロード / ファイル: emd_2708.map.gz / 形式: CCP4 / 大きさ: 6.7 GB / タイプ: IMAGE STORED AS SIGNED INTEGER (2 BYTES)
注釈
Cryo-electron tomogram of immature HIV-1 particles treated with the protease inhibitor Amprenavir
ボクセルのサイズ
X=Y=Z: 2.025 Å
密度
最小 - 最大
-32767.0 - 26529.0
平均 (標準偏差)
146.287979129999997 (±2433.4487304700001)
対称性
空間群: 1
詳細
EMDB XML:
マップ形状
Axis order
X
Y
Z
Origin
0
0
64
サイズ
2048
2048
880
Spacing
2048
2048
880
セル
A: 4147.2 Å / B: 4147.2 Å / C: 1782.0001 Å α=β=γ: 90.0 °
CCP4マップ ヘッダ情報:
mode
Image stored as Integer*27
Å/pix. X/Y/Z
2.025
2.025
2.025
M x/y/z
2048
2048
880
origin x/y/z
0.000
0.000
0.000
length x/y/z
4147.200
4147.200
1782.000
α/β/γ
90.000
90.000
90.000
start NX/NY/NZ
-180
-180
-179
NX/NY/NZ
360
360
360
MAP C/R/S
1
2
3
start NC/NR/NS
0
0
64
NC/NR/NS
2048
2048
880
D min/max/mean
-32767.000
26529.000
146.288
-
添付データ
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試料の構成要素
-
全体 : immature HIV-1 particles treated with the protease inhibitor Ampr...
全体
名称: immature HIV-1 particles treated with the protease inhibitor Amprenavir
要素
試料: immature HIV-1 particles treated with the protease inhibitor Amprenavir
ウイルス: Human immunodeficiency virus 1 (ヒト免疫不全ウイルス)
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超分子 #1000: immature HIV-1 particles treated with the protease inhibitor Ampr...
超分子
名称: immature HIV-1 particles treated with the protease inhibitor Amprenavir タイプ: sample / ID: 1000 / Number unique components: 1
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超分子 #1: Human immunodeficiency virus 1
超分子
名称: Human immunodeficiency virus 1 / タイプ: virus / ID: 1 / Name.synonym: HIV-1 / NCBI-ID: 11676 / 生物種: Human immunodeficiency virus 1 / データベース: NCBI / 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: Yes / ウイルス・中空状態: No / Syn species name: HIV-1
宿主
生物種: Homo sapiens (ヒト) / 別称: VERTEBRATES
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実験情報
-
構造解析
手法
クライオ電子顕微鏡法
解析
電子線トモグラフィー法
試料の集合状態
particle
-
試料調製
緩衝液
pH: 6.5 / 詳細: 25mM MES pH 6.5, 150mM NaCl
グリッド
詳細: C-Flat 2/2-2C glow discharged
凍結
凍結剤: ETHANE / チャンバー内湿度: 100 % / 装置: FEI VITROBOT MARK II 手法: Degassed C-Flat 2/2-2C grids were glow discharged for 30 seconds at 20 mA. Virus solution was diluted in PBS containing 10nm colloidal gold. 2 ul of this mixture was applied to a grid. Blotting time: 2 seconds
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電子顕微鏡法
顕微鏡
FEI TITAN KRIOS
アライメント法
Legacy - 非点収差: Objective lens astigmatism was corrected at nominal working magnification