ジャーナル: Proc Natl Acad Sci U S A / 年: 2013 タイトル: Uncoating of common cold virus is preceded by RNA switching as determined by X-ray and cryo-EM analyses of the subviral A-particle. 著者: Angela Pickl-Herk / Daniel Luque / Laia Vives-Adrián / Jordi Querol-Audí / Damià Garriga / Benes L Trus / Nuria Verdaguer / Dieter Blaas / José R Castón / 要旨: During infection, viruses undergo conformational changes that lead to delivery of their genome into host cytosol. In human rhinovirus A2, this conversion is triggered by exposure to acid pH in the ...During infection, viruses undergo conformational changes that lead to delivery of their genome into host cytosol. In human rhinovirus A2, this conversion is triggered by exposure to acid pH in the endosome. The first subviral intermediate, the A-particle, is expanded and has lost the internal viral protein 4 (VP4), but retains its RNA genome. The nucleic acid is subsequently released, presumably through one of the large pores that open at the icosahedral twofold axes, and is transferred along a conduit in the endosomal membrane; the remaining empty capsids, termed B-particles, are shuttled to lysosomes for degradation. Previous structural analyses revealed important differences between the native protein shell and the empty capsid. Nonetheless, little is known of A-particle architecture or conformation of the RNA core. Using 3D cryo-electron microscopy and X-ray crystallography, we found notable changes in RNA-protein contacts during conversion of native virus into the A-particle uncoating intermediate. In the native virion, we confirmed interaction of nucleotide(s) with Trp(38) of VP2 and identified additional contacts with the VP1 N terminus. Study of A-particle structure showed that the VP2 contact is maintained, that VP1 interactions are lost after exit of the VP1 N-terminal extension, and that the RNA also interacts with residues of the VP3 N terminus at the fivefold axis. These associations lead to formation of a well-ordered RNA layer beneath the protein shell, suggesting that these interactions guide ordered RNA egress.
名称: Human rhinovirus 2 / タイプ: virus / ID: 1 / Name.synonym: Common cold virus / NCBI-ID: 12130 / 生物種: Human rhinovirus 2 / ウイルスタイプ: VIRION / ウイルス・単離状態: SEROTYPE / ウイルス・エンベロープ: No / ウイルス・中空状態: Yes / Syn species name: Common cold virus
宿主
生物種: Homo sapiens (ヒト) / 別称: VERTEBRATES
ウイルス殻
Shell ID: 1 / 直径: 316 Å / T番号(三角分割数): 1
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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: 50 mM sodium borate
グリッド
詳細: R 1.2/1.3 Quantifoil grids
凍結
凍結剤: ETHANE / チャンバー内湿度: 80 % / チャンバー内温度: 99 K / 装置: LEICA EM GP 手法: Using a Leica EM Grid Plunger, sample was applied per grid in a humidified chamber, left for 30 s, blotted for 0.8 s and plunge-frozen
Protocol: Rigid body. The asymmetric unit was initially treated as a single rigid body and then refined, treating each subunit as an independent rigid body
精密化
空間: RECIPROCAL / プロトコル: RIGID BODY FIT / 当てはまり具合の基準: CC