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基本情報
登録情報 | データベース: PDB / ID: 3zq0 | ||||||
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タイトル | Visualizing GroEL-ES in the Act of Encapsulating a Non-Native Substrate Protein | ||||||
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![]() | PROTEIN FOLDING / HETEROGENEITY / ENCAPSULATION | ||||||
機能・相同性 | ![]() GroEL-GroES complex / chaperonin ATPase / virion assembly / chaperone cofactor-dependent protein refolding / protein folding chaperone / isomerase activity / ATP-dependent protein folding chaperone / response to radiation / unfolded protein binding / protein folding ...GroEL-GroES complex / chaperonin ATPase / virion assembly / chaperone cofactor-dependent protein refolding / protein folding chaperone / isomerase activity / ATP-dependent protein folding chaperone / response to radiation / unfolded protein binding / protein folding / protein-folding chaperone binding / protein refolding / response to heat / magnesium ion binding / ATP hydrolysis activity / ATP binding / identical protein binding / membrane / metal ion binding / cytosol 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: 9.2 Å | ||||||
![]() | Chen, D.-H. / Madan, D. / Weaver, J. / Lin, Z. / Schroder, G.F. / Chiu, W. / Rye, H.S. | ||||||
![]() | ![]() タイトル: Visualizing GroEL/ES in the act of encapsulating a folding protein. 著者: Dong-Hua Chen / Damian Madan / Jeremy Weaver / Zong Lin / Gunnar F Schröder / Wah Chiu / Hays S Rye / ![]() 要旨: The GroEL/ES chaperonin system is required for the assisted folding of many proteins. How these substrate proteins are encapsulated within the GroEL-GroES cavity is poorly understood. Using symmetry- ...The GroEL/ES chaperonin system is required for the assisted folding of many proteins. How these substrate proteins are encapsulated within the GroEL-GroES cavity is poorly understood. Using symmetry-free, single-particle cryo-electron microscopy, we have characterized a chemically modified mutant of GroEL (EL43Py) that is trapped at a normally transient stage of substrate protein encapsulation. We show that the symmetric pattern of the GroEL subunits is broken as the GroEL cis-ring apical domains reorient to accommodate the simultaneous binding of GroES and an incompletely folded substrate protein (RuBisCO). The collapsed RuBisCO folding intermediate binds to the lower segment of two apical domains, as well as to the normally unstructured GroEL C-terminal tails. A comparative structural analysis suggests that the allosteric transitions leading to substrate protein release and folding involve concerted shifts of GroES and the GroEL apical domains and C-terminal tails. | ||||||
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構造の表示
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構造ビューア | 分子: ![]() ![]() |
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-関連構造データ
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集合体
登録構造単位 | ![]()
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要素
#1: タンパク質 | 分子量: 55220.105 Da / 分子数: 14 / 由来タイプ: 組換発現 詳細: A GROEL VARIANT IN WHICH THE ENDOGENOUS CYS RESIDUES (138,458,519) HAVE BEEN CHANGED TO ALA (GROELCYS0) WAS MODIFIED TO CONTAIN TWO ADDITIONAL MUTATIONS, D398A AND S43C. THE D398A MUTATION ...詳細: A GROEL VARIANT IN WHICH THE ENDOGENOUS CYS RESIDUES (138,458,519) HAVE BEEN CHANGED TO ALA (GROELCYS0) WAS MODIFIED TO CONTAIN TWO ADDITIONAL MUTATIONS, D398A AND S43C. THE D398A MUTATION PREVENTS ATP HYDROLYSIS BY GROEL, WHILE THE S43C MUTATION, LOCATED AT THE TIP OF STEM LOOP AT THE BOTTOM OF THE GROEL CAVITY, PERMITS THE COVALENT ATTACHMENT OF N-1-PYRENE MALEIMIDE TO THIS POSITION. 由来: (組換発現) ![]() ![]() ![]() ![]() #2: タンパク質 | 分子量: 10400.938 Da / 分子数: 7 / 由来タイプ: 組換発現 / 由来: (組換発現) ![]() ![]() ![]() ![]() #3: 化合物 | ChemComp-ADP / #4: 化合物 | ChemComp-MG / |
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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: NON-NATIVE RUBISCO SUBSTRATE PROTEIN ENCAPSULATED INSIDE THE CAVITY OF GROEL VARIANT EL43PY CAPPED BY GROES WITH THE ASSISTANCE OF NUCLEOTIDE ATP タイプ: COMPLEX |
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緩衝液 | 名称: 50 MM HEPES, 50 MM KOAC, 10 MM MG(OAC)2, 2 MM DTT / pH: 7.6 / 詳細: 50 MM HEPES, 50 MM KOAC, 10 MM MG(OAC)2, 2 MM DTT |
試料 | 濃度: 2.4 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES |
試料支持 | 詳細: HOLEY CARBON |
急速凍結 | 装置: FEI VITROBOT MARK III / 凍結剤: ETHANE 詳細: VITRIFICATION 1 -- CRYOGEN- ETHANE, HUMIDITY- 95, TEMPERATURE- 98, INSTRUMENT- FEI VITROBOT MARK III, METHOD BLOT FOR 1 SECOND BEFORE PLUNGIN |
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電子顕微鏡撮影
顕微鏡 | モデル: JEOL KYOTO-3000SFF / 日付: 2008年8月16日 |
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電子銃 | 電子線源: ![]() |
電子レンズ | モード: OTHER / 倍率(公称値): 50000 X / 倍率(補正後): 73530 X / 最大 デフォーカス(公称値): 4700 nm / 最小 デフォーカス(公称値): 1000 nm / Cs: 1.6 mm |
試料ホルダ | 温度: 50.5 K |
撮影 | 電子線照射量: 30 e/Å2 / フィルム・検出器のモデル: GENERIC GATAN |
画像スキャン | デジタル画像の数: 2000 |
放射波長 | 相対比: 1 |
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解析
EMソフトウェア |
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CTF補正 | 詳細: EACH FRAME | ||||||||||||
対称性 | 点対称性: C1 (非対称) | ||||||||||||
3次元再構成 | 手法: FOURIER METHODS / 解像度: 9.2 Å / 粒子像の数: 17611 / ピクセルサイズ(公称値): 2.14 Å / ピクセルサイズ(実測値): 2.04 Å 倍率補正: THE RMSD BETWEEN ALL EQUATORIAL DOMAINS OF THE FITTED MODELS AND THE X-RAY STRUCTURE OF THE GROEL-GROES-ADP COMPLEX WAS CALCULATED AND THE OPTIMAL PIXEL SIZE WAS CHOSEN AS THE ONE THAT ...倍率補正: THE RMSD BETWEEN ALL EQUATORIAL DOMAINS OF THE FITTED MODELS AND THE X-RAY STRUCTURE OF THE GROEL-GROES-ADP COMPLEX WAS CALCULATED AND THE OPTIMAL PIXEL SIZE WAS CHOSEN AS THE ONE THAT LEADS TO THE SMALLEST RMSD VALUE 詳細: SUBMISSION BASED ON EXPERIMENTAL DATA FROM EMDB EMD-2326. (DEPOSITION ID: 11455). 対称性のタイプ: POINT | ||||||||||||
原子モデル構築 | プロトコル: FLEXIBLE FIT / 空間: REAL / Target criteria: Cross-correlation coefficient / 詳細: METHOD--FLEXIBLE REFINEMENT PROTOCOL--X-RAY | ||||||||||||
原子モデル構築 | PDB-ID: 1AON Accession code: 1AON / Source name: PDB / タイプ: experimental model | ||||||||||||
精密化 | 最高解像度: 9.2 Å | ||||||||||||
精密化ステップ | サイクル: LAST / 最高解像度: 9.2 Å
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