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データを開く
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基本情報
登録情報 | データベース: PDB / ID: 7jsf | ||||||
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タイトル | Adeno-Associated Virus Helicase domain Heptamer with ssDNA | ||||||
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![]() | VIRAL PROTEIN / AAV / Protein-DNA / AAA+ / SF3 / HUH | ||||||
機能・相同性 | ![]() symbiont-mediated arrest of host cell cycle during G2/M transition / permeabilization of host organelle membrane involved in viral entry into host cell / symbiont entry into host cell via permeabilization of inner membrane / viral DNA genome replication / symbiont-mediated perturbation of host cell cycle G1/S transition checkpoint / DNA helicase / endonuclease activity / DNA replication / host cell nucleus / ATP hydrolysis activity ...symbiont-mediated arrest of host cell cycle during G2/M transition / permeabilization of host organelle membrane involved in viral entry into host cell / symbiont entry into host cell via permeabilization of inner membrane / viral DNA genome replication / symbiont-mediated perturbation of host cell cycle G1/S transition checkpoint / DNA helicase / endonuclease activity / DNA replication / host cell nucleus / ATP hydrolysis activity / DNA binding / ATP binding / metal ion binding 類似検索 - 分子機能 | ||||||
生物種 | ![]() synthetic construct (人工物) | ||||||
手法 | 電子顕微鏡法 / 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 6.7 Å | ||||||
![]() | Escalante, C.R. | ||||||
資金援助 | ![]()
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![]() | ![]() タイトル: The Cryo-EM structure of AAV2 Rep68 in complex with ssDNA reveals a malleable AAA+ machine that can switch between oligomeric states. 著者: Vishaka Santosh / Faik N Musayev / Rahul Jaiswal / Francisco Zárate-Pérez / Bram Vandewinkel / Caroline Dierckx / Molly Endicott / Kamyar Sharifi / Kelly Dryden / Els Henckaerts / Carlos R Escalante / ![]() ![]() 要旨: The adeno-associated virus (AAV) non-structural Rep proteins catalyze all the DNA transactions required for virus viability including, DNA replication, transcription regulation, genome packaging, and ...The adeno-associated virus (AAV) non-structural Rep proteins catalyze all the DNA transactions required for virus viability including, DNA replication, transcription regulation, genome packaging, and during the latent phase, site-specific integration. Rep proteins contain two multifunctional domains: an Origin Binding Domain (OBD) and a SF3 helicase domain (HD). Studies have shown that Rep proteins have a dynamic oligomeric behavior where the nature of the DNA substrate molecule modulates its oligomeric state. In the presence of ssDNA, Rep68 forms a large double-octameric ring complex. To understand the mechanisms underlying AAV Rep function, we investigated the cryo-EM and X-ray structures of Rep68-ssDNA complexes. Surprisingly, Rep68 generates hybrid ring structures where the OBD forms octameric rings while the HD forms heptamers. Moreover, the binding to ATPγS promotes a large conformational change in the entire AAA+ domain that leads the HD to form both heptamer and hexamers. The HD oligomerization is driven by an interdomain linker region that acts as a latch to 'catch' the neighboring HD subunit and is flexible enough to permit the formation of different stoichiometric ring structures. Overall, our studies show the structural basis of AAV Rep's structural flexibility required to fulfill its multifunctional role during the AAV life cycle. | ||||||
履歴 |
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構造の表示
ムービー |
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構造ビューア | 分子: ![]() ![]() |
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ダウンロードとリンク
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PDBx/mmCIF形式 | ![]() | 270.5 KB | 表示 | ![]() |
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PDB形式 | ![]() | 163 KB | 表示 | ![]() |
PDBx/mmJSON形式 | ![]() | ツリー表示 | ![]() | |
その他 | ![]() |
-検証レポート
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
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-関連構造データ
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リンク
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集合体
登録構造単位 | ![]()
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1 |
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要素
#1: タンパク質 | 分子量: 60908.703 Da / 分子数: 7 / 変異: C151S / 由来タイプ: 組換発現 由来: (組換発現) ![]() 遺伝子: Rep68 発現宿主: ![]() ![]() 参照: UniProt: P03132, DNA helicase #2: DNA鎖 | | 分子量: 1171.814 Da / 分子数: 1 / 由来タイプ: 合成 / 由来: (合成) synthetic construct (人工物) |
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-実験情報
-実験
実験 | 手法: 電子顕微鏡法 |
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EM実験 | 試料の集合状態: PARTICLE / 3次元再構成法: 単粒子再構成法 |
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試料調製
構成要素 | 名称: Helicase domain heptamer of Rep68 complx bound to ssDNA タイプ: COMPLEX / Entity ID: all / 由来: MULTIPLE SOURCES | |||||||||||||||||||||||||
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由来(天然) |
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緩衝液 | pH: 7.9 | |||||||||||||||||||||||||
緩衝液成分 |
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試料 | 濃度: 0.75 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES | |||||||||||||||||||||||||
試料支持 | グリッドの材料: COPPER / グリッドのサイズ: 400 divisions/in. / グリッドのタイプ: C-flat-1.2/1.3 | |||||||||||||||||||||||||
急速凍結 | 凍結剤: ETHANE |
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電子顕微鏡撮影
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TITAN KRIOS |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(補正後): 165000 X |
試料ホルダ | 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER |
撮影 | 電子線照射量: 92.03 e/Å2 / 検出モード: SUPER-RESOLUTION フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 実像数: 1449 |
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解析
EMソフトウェア |
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CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||
粒子像の選択 | 選択した粒子像数: 118531 | ||||||||||||
3次元再構成 | 解像度: 6.7 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 46031 / 対称性のタイプ: POINT | ||||||||||||
原子モデル構築 | プロトコル: RIGID BODY FIT | ||||||||||||
原子モデル構築 | PDB-ID: 1S9H Accession code: 1S9H / Pdb chain residue range: 225-490 / Source name: PDB / タイプ: experimental model |