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基本情報
登録情報 | データベース: PDB / ID: 5u1c | ||||||||||||
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タイトル | Structure of tetrameric HIV-1 Strand Transfer Complex Intasome | ||||||||||||
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![]() | VIRAL PROTEIN / integrase / integration / transposase / transesterification | ||||||||||||
機能・相同性 | ![]() RNA endonuclease activity / HIV-1 retropepsin / symbiont-mediated activation of host apoptosis / retroviral ribonuclease H / exoribonuclease H / exoribonuclease H activity / host multivesicular body / DNA integration / viral genome integration into host DNA / RNA-directed DNA polymerase ...RNA endonuclease activity / HIV-1 retropepsin / symbiont-mediated activation of host apoptosis / retroviral ribonuclease H / exoribonuclease H / exoribonuclease H activity / host multivesicular body / DNA integration / viral genome integration into host DNA / RNA-directed DNA polymerase / establishment of integrated proviral latency / viral penetration into host nucleus / RNA stem-loop binding / symbiont-mediated suppression of host gene expression / RNA-directed DNA polymerase activity / host cell / RNA-DNA hybrid ribonuclease activity / 転移酵素; リンを含む基を移すもの; 核酸を移すもの / viral nucleocapsid / endonuclease activity / DNA recombination / DNA-directed DNA polymerase / 加水分解酵素; エステル加水分解酵素 / aspartic-type endopeptidase activity / DNA-directed DNA polymerase activity / symbiont entry into host cell / lipid binding / host cell nucleus / host cell plasma membrane / structural molecule activity / virion membrane / proteolysis / DNA binding / zinc ion binding / membrane 類似検索 - 分子機能 | ||||||||||||
生物種 | ![]() ![]() ![]() ![]() ![]() | ||||||||||||
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: 3.9 Å | ||||||||||||
![]() | Lyumkis, D. / Passos, D. | ||||||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Cryo-EM structures and atomic model of the HIV-1 strand transfer complex intasome. 著者: Dario Oliveira Passos / Min Li / Renbin Yang / Stephanie V Rebensburg / Rodolfo Ghirlando / Youngmin Jeon / Nikoloz Shkriabai / Mamuka Kvaratskhelia / Robert Craigie / Dmitry Lyumkis / ![]() 要旨: Like all retroviruses, HIV-1 irreversibly inserts a viral DNA (vDNA) copy of its RNA genome into host target DNA (tDNA). The intasome, a higher-order nucleoprotein complex composed of viral integrase ...Like all retroviruses, HIV-1 irreversibly inserts a viral DNA (vDNA) copy of its RNA genome into host target DNA (tDNA). The intasome, a higher-order nucleoprotein complex composed of viral integrase (IN) and the ends of linear vDNA, mediates integration. Productive integration into host chromatin results in the formation of the strand transfer complex (STC) containing catalytically joined vDNA and tDNA. HIV-1 intasomes have been refractory to high-resolution structural studies. We used a soluble IN fusion protein to facilitate structural studies, through which we present a high-resolution cryo-electron microscopy (cryo-EM) structure of the core tetrameric HIV-1 STC and a higher-order form that adopts carboxyl-terminal domain rearrangements. The distinct STC structures highlight how HIV-1 can use the common retroviral intasome core architecture to accommodate different IN domain modules for assembly. | ||||||||||||
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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PDBx/mmCIF形式 | ![]() | 212.4 KB | 表示 | ![]() |
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PDB形式 | ![]() | 156.5 KB | 表示 | ![]() |
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その他 | ![]() |
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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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要素
-タンパク質 , 1種, 4分子 ABCD
#1: タンパク質 | 分子量: 42320.273 Da / 分子数: 4 / 変異: E152Q / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() ![]() ![]() 遺伝子: SSOP1_2677, pol / 発現宿主: ![]() ![]() 参照: UniProt: A0A157T5S7, UniProt: F2WR39, UniProt: P12497*PLUS |
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-DNA鎖 , 3種, 6分子 GHEIFJ
#2: DNA鎖 | 分子量: 3349.197 Da / 分子数: 2 / 由来タイプ: 合成 / 由来: (合成) ![]() #3: DNA鎖 | 分子量: 7015.546 Da / 分子数: 2 / 由来タイプ: 合成 由来: (合成) ![]() ![]() #4: DNA鎖 | 分子量: 11414.358 Da / 分子数: 2 / 由来タイプ: 合成 由来: (合成) ![]() ![]() ![]() |
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-非ポリマー , 2種, 4分子 ![](data/chem/img/ZN.gif)
![](data/chem/img/MG.gif)
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#5: 化合物 | #6: 化合物 | |
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-詳細
配列の詳細 | The protein is a chimera of Sso7d linked to the N-terminus of integrase via an 11-glycine linker. |
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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: complex formed by a tetrameric assembly of Sso7d-fusion HIV-1 Integrase with the product of DNA strand transfer タイプ: COMPLEX / Entity ID: #1-#4 / 由来: RECOMBINANT | |||||||||||||||||||||||||
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分子量 | 値: 0.228 MDa / 実験値: YES | |||||||||||||||||||||||||
由来(天然) | 生物種: ![]() ![]() | |||||||||||||||||||||||||
由来(組換発現) | 生物種: ![]() ![]() | |||||||||||||||||||||||||
緩衝液 | pH: 8 | |||||||||||||||||||||||||
緩衝液成分 |
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試料 | 濃度: 0.5 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: GOLD / グリッドのサイズ: 400 divisions/in. / グリッドのタイプ: Quantifoil | |||||||||||||||||||||||||
急速凍結 | 装置: HOMEMADE PLUNGER / 凍結剤: ETHANE / 湿度: 50 % / 凍結前の試料温度: 277 K 詳細: Sample containing HIV STC intasomes in SEC buffer was applied onto freshly plasma-treated (6 seconds, Gatan Solarus plasma cleaner) holey gold UltrAuFoil grids (Quantifoil), adsorbed for 30 ...詳細: Sample containing HIV STC intasomes in SEC buffer was applied onto freshly plasma-treated (6 seconds, Gatan Solarus plasma cleaner) holey gold UltrAuFoil grids (Quantifoil), adsorbed for 30 seconds, then plunged into liquid ethane using a manual cryo-plunger in an ambient environment of 4 degrees C. |
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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(公称値): 22500 X / 倍率(補正後): 38167 X / Calibrated defocus min: 1500 nm / 最大 デフォーカス(補正後): 4000 nm / Cs: 2.7 mm / C2レンズ絞り径: 100 µm / アライメント法: COMA FREE |
試料ホルダ | 凍結剤: NITROGEN 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER 最高温度: 90 K / 最低温度: 90 K |
撮影 | 平均露光時間: 20 sec. / 電子線照射量: 95 e/Å2 / 検出モード: COUNTING フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 撮影したグリッド数: 1 / 実像数: 1225 詳細: Individual frames were gain-corrected, aligned, and summed with the application of an exposure filter using MotionCor2, according to the nominal dose rate. |
画像スキャン | サンプリングサイズ: 5 µm / 横: 3838 / 縦: 3710 / 動画フレーム数/画像: 100 / 利用したフレーム数/画像: 1-100 |
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解析
ソフトウェア | 名称: PHENIX / バージョン: dev_2499: / 分類: 精密化 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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EMソフトウェア |
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CTF補正 | 詳細: performed internally in Relion and Frealign / タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
粒子像の選択 | 選択した粒子像数: 274764 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C2 (2回回転対称) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 3.9 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 83766 / アルゴリズム: FOURIER SPACE / 詳細: Resolution-limited refinement used throughout / 対称性のタイプ: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
原子モデル構築 | B value: 180 / プロトコル: FLEXIBLE FIT / 空間: REAL / Target criteria: FSC 0.5 詳細: To generate ensemble models, the complete intasome model was iteratively relaxed - using two-fold symmetry and a combination of Rosetta and Phenix - against one half map (the working map) and ...詳細: To generate ensemble models, the complete intasome model was iteratively relaxed - using two-fold symmetry and a combination of Rosetta and Phenix - against one half map (the working map) and inspected for consistency with the second half map (the free map). The model was then adjusted manually using Coot. Final ensemble modeling used half maps for all aspects of refinement and evaluation: 500 models were generated as described using Rosetta. From the 100 top-scoring models (scored by Rosetta energy), the ten models with the best map-to-model FSC were selected and refined in real space using secondary-structure restraints in Phenix. Molprobity was used throughout the refinement process. |