ジャーナル: J Mol Biol / 年: 2010 タイトル: Mutually-induced conformational switching of RNA and coat protein underpins efficient assembly of a viral capsid. 著者: Óttar Rolfsson / Katerina Toropova / Neil A Ranson / Peter G Stockley / 要旨: Single-stranded RNA viruses package their genomes into capsids enclosing fixed volumes. We assayed the ability of bacteriophage MS2 coat protein to package large, defined fragments of its genomic, ...Single-stranded RNA viruses package their genomes into capsids enclosing fixed volumes. We assayed the ability of bacteriophage MS2 coat protein to package large, defined fragments of its genomic, single-stranded RNA. We show that the efficiency of packaging into a T=3 capsid in vitro is inversely proportional to RNA length, implying that there is a free-energy barrier to be overcome during assembly. All the RNAs examined have greater solution persistence lengths than the internal diameter of the capsid into which they become packaged, suggesting that protein-mediated RNA compaction must occur during assembly. Binding ethidium bromide to one of these RNA fragments, which would be expected to reduce its flexibility, severely inhibited packaging, consistent with this idea. Cryo-EM structures of the capsids assembled in these experiments with the sub-genomic RNAs show a layer of RNA density beneath the coat protein shell but lack density for the inner RNA shell seen in the wild-type virion. The inner layer is restored when full-length virion RNA is used in the assembly reaction, implying that it becomes ordered only when the capsid is filled, presumably because of the effects of steric and/or electrostatic repulsions. The cryo-EM results explain the length dependence of packaging. In addition, they show that for the sub-genomic fragments the strongest ordered RNA density occurs below the coat protein dimers forming the icosahedral 5-fold axes of the capsid. There is little such density beneath the proteins at the 2-fold axes, consistent with our model in which coat protein dimers binding to RNA stem-loops located at sites throughout the genome leads to switching of their preferred conformations, thus regulating the placement of the quasi-conformers needed to build the T=3 capsid. The data are consistent with mutual chaperoning of both RNA and coat protein conformations, partially explaining the ability of such viruses to assemble so rapidly and accurately.
ダウンロード / ファイル: emd_1882.map.gz / 形式: CCP4 / 大きさ: 29.8 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
注釈
This is a surface-rendered view of a Bacteriophage MS2 virus-like particle composed of purified coat protein and genomic RNA
ボクセルのサイズ
X=Y=Z: 1.87 Å
密度
表面レベル
By EMDB: 1.0 / ムービー #1: 1
最小 - 最大
-4.45316 - 5.72175
平均 (標準偏差)
0.00000000557511 (±1.0)
対称性
空間群: 1
詳細
EMDB XML:
マップ形状
Axis order
X
Y
Z
Origin
0
0
0
サイズ
200
200
200
Spacing
200
200
200
セル
A=B=C: 374 Å α=β=γ: 90 °
CCP4マップ ヘッダ情報:
mode
Image stored as Reals
Å/pix. X/Y/Z
1.87
1.87
1.87
M x/y/z
200
200
200
origin x/y/z
-0.000
-0.000
-0.000
length x/y/z
374.000
374.000
374.000
α/β/γ
90.000
90.000
90.000
start NX/NY/NZ
-56
-56
-55
NX/NY/NZ
112
112
112
MAP C/R/S
1
2
3
start NC/NR/NS
0
0
0
NC/NR/NS
200
200
200
D min/max/mean
-4.453
5.722
0.000
-
添付データ
-
試料の構成要素
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全体 : Bacteriophage MS2 virus-like particles assembled in vitro from pu...
全体
名称: Bacteriophage MS2 virus-like particles assembled in vitro from purified MS2 coat protein and genomic RNA
要素
試料: Bacteriophage MS2 virus-like particles assembled in vitro from purified MS2 coat protein and genomic RNA
ウイルス: Enterobacterio phage MS2 (ファージ)
RNA: Single stranded RNA MS2 genome
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超分子 #1000: Bacteriophage MS2 virus-like particles assembled in vitro from pu...
超分子
名称: Bacteriophage MS2 virus-like particles assembled in vitro from purified MS2 coat protein and genomic RNA タイプ: sample / ID: 1000 / 集合状態: One capsid to one RNA / Number unique components: 2
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超分子 #1: Enterobacterio phage MS2
超分子
名称: Enterobacterio phage MS2 / タイプ: virus / ID: 1 / Name.synonym: MS2 / NCBI-ID: 12022 / 生物種: Enterobacterio phage MS2 / ウイルスタイプ: VIRUS-LIKE PARTICLE / ウイルス・単離状態: STRAIN / ウイルス・エンベロープ: No / ウイルス・中空状態: No / Syn species name: MS2
名称: Single stranded RNA MS2 genome / タイプ: rna / ID: 1 / Name.synonym: Single stranded RNA MS2 genome 詳細: Single stranded RNA MS2 genome was phenol-chloroform extracted from wild type MS2 phage 分類: OTHER / Structure: SINGLE STRANDED / Synthetic?: No