+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 8cvs | ||||||
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タイトル | Human PA200-20S proteasome with MG-132 | ||||||
要素 |
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キーワード | HYDROLASE/INHIBITOR / proteolysis / protein degradation / complex / inhibitor / MG-132 / MG132 / HYDROLASE-INHIBITOR complex | ||||||
機能・相同性 | 機能・相同性情報 spermatoproteasome complex / sperm DNA condensation / peptidase activator activity / purine ribonucleoside triphosphate binding / regulation of endopeptidase activity / Regulation of ornithine decarboxylase (ODC) / proteasome core complex / Cross-presentation of soluble exogenous antigens (endosomes) / Somitogenesis / proteasomal ubiquitin-independent protein catabolic process ...spermatoproteasome complex / sperm DNA condensation / peptidase activator activity / purine ribonucleoside triphosphate binding / regulation of endopeptidase activity / Regulation of ornithine decarboxylase (ODC) / proteasome core complex / Cross-presentation of soluble exogenous antigens (endosomes) / Somitogenesis / proteasomal ubiquitin-independent protein catabolic process / immune system process / myofibril / proteasome binding / NF-kappaB binding / proteasome endopeptidase complex / proteasome core complex, beta-subunit complex / proteasome core complex, alpha-subunit complex / threonine-type endopeptidase activity / negative regulation of inflammatory response to antigenic stimulus / lysine-acetylated histone binding / : / sarcomere / proteasome complex / Regulation of activated PAK-2p34 by proteasome mediated degradation / ciliary basal body / proteolysis involved in protein catabolic process / Autodegradation of Cdh1 by Cdh1:APC/C / APC/C:Cdc20 mediated degradation of Securin / Asymmetric localization of PCP proteins / SCF-beta-TrCP mediated degradation of Emi1 / NIK-->noncanonical NF-kB signaling / Ubiquitin-dependent degradation of Cyclin D / AUF1 (hnRNP D0) binds and destabilizes mRNA / TNFR2 non-canonical NF-kB pathway / Vpu mediated degradation of CD4 / Assembly of the pre-replicative complex / Degradation of DVL / CDK-mediated phosphorylation and removal of Cdc6 / Ubiquitin Mediated Degradation of Phosphorylated Cdc25A / Dectin-1 mediated noncanonical NF-kB signaling / Cdc20:Phospho-APC/C mediated degradation of Cyclin A / lipopolysaccharide binding / SCF(Skp2)-mediated degradation of p27/p21 / Hh mutants are degraded by ERAD / Degradation of AXIN / Degradation of GLI1 by the proteasome / Activation of NF-kappaB in B cells / Hedgehog ligand biogenesis / Negative regulation of NOTCH4 signaling / Defective CFTR causes cystic fibrosis / GSK3B and BTRC:CUL1-mediated-degradation of NFE2L2 / G2/M Checkpoints / Vif-mediated degradation of APOBEC3G / Autodegradation of the E3 ubiquitin ligase COP1 / Hedgehog 'on' state / Regulation of RUNX3 expression and activity / P-body / Degradation of GLI2 by the proteasome / GLI3 is processed to GLI3R by the proteasome / Orc1 removal from chromatin / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / MAPK6/MAPK4 signaling / APC/C:Cdh1 mediated degradation of Cdc20 and other APC/C:Cdh1 targeted proteins in late mitosis/early G1 / response to virus / Degradation of beta-catenin by the destruction complex / Oxygen-dependent proline hydroxylation of Hypoxia-inducible Factor Alpha / ABC-family proteins mediated transport / response to organic cyclic compound / CLEC7A (Dectin-1) signaling / Regulation of expression of SLITs and ROBOs / FCERI mediated NF-kB activation / nuclear matrix / Regulation of PTEN stability and activity / Interleukin-1 signaling / Regulation of RAS by GAPs / Separation of Sister Chromatids / Regulation of RUNX2 expression and activity / The role of GTSE1 in G2/M progression after G2 checkpoint / UCH proteinases / KEAP1-NFE2L2 pathway / Antigen processing: Ubiquitination & Proteasome degradation / regulation of inflammatory response / Downstream TCR signaling / RUNX1 regulates transcription of genes involved in differentiation of HSCs / Neddylation / positive regulation of NF-kappaB transcription factor activity / peptidase activity / ER-Phagosome pathway / endopeptidase activity / proteasome-mediated ubiquitin-dependent protein catabolic process / secretory granule lumen / response to oxidative stress / ficolin-1-rich granule lumen / postsynapse / nuclear body / ribosome / Ub-specific processing proteases / nuclear speck / cadherin binding / DNA repair 類似検索 - 分子機能 | ||||||
生物種 | Homo sapiens (ヒト) | ||||||
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: 3.1 Å | ||||||
データ登録者 | Zhao, J. | ||||||
資金援助 | 1件
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引用 | ジャーナル: Proc Natl Acad Sci U S A / 年: 2022 タイトル: Structural insights into the human PA28-20S proteasome enabled by efficient tagging and purification of endogenous proteins. 著者: Jianhua Zhao / Suraj Makhija / Chenyu Zhou / Hanxiao Zhang / YongQiang Wang / Monita Muralidharan / Bo Huang / Yifan Cheng / 要旨: The ability to produce folded and functional proteins is a necessity for structural biology and many other biological sciences. This task is particularly challenging for numerous biomedically ...The ability to produce folded and functional proteins is a necessity for structural biology and many other biological sciences. This task is particularly challenging for numerous biomedically important targets in human cells, including membrane proteins and large macromolecular assemblies, hampering mechanistic studies and drug development efforts. Here we describe a method combining CRISPR-Cas gene editing and fluorescence-activated cell sorting to rapidly tag and purify endogenous proteins in HEK cells for structural characterization. We applied this approach to study the human proteasome from HEK cells and rapidly determined cryogenic electron microscopy structures of major proteasomal complexes, including a high-resolution structure of intact human PA28αβ-20S. Our structures reveal that PA28 with a subunit stoichiometry of 3α/4β engages tightly with the 20S proteasome. Addition of a hydrophilic peptide shows that polypeptides entering through PA28 are held in the antechamber of 20S prior to degradation in the proteolytic chamber. This study provides critical insights into an important proteasome complex and demonstrates key methodologies for the tagging of proteins from endogenous sources. | ||||||
履歴 |
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-構造の表示
構造ビューア | 分子: MolmilJmol/JSmol |
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-ダウンロードとリンク
-ダウンロード
PDBx/mmCIF形式 | 8cvs.cif.gz | 1.7 MB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb8cvs.ent.gz | 1.3 MB | 表示 | PDB形式 |
PDBx/mmJSON形式 | 8cvs.json.gz | ツリー表示 | PDBx/mmJSON形式 | |
その他 | その他のダウンロード |
-検証レポート
アーカイブディレクトリ | https://data.pdbj.org/pub/pdb/validation_reports/cv/8cvs ftp://data.pdbj.org/pub/pdb/validation_reports/cv/8cvs | HTTPS FTP |
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-関連構造データ
関連構造データ | 27015MC 7nanC 7naoC 7napC 7naqC 8cvrC 8cvtC 8cxbC M: このデータのモデリングに利用したマップデータ C: 同じ文献を引用 (文献) |
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類似構造データ | 類似検索 - 機能・相同性F&H 検索 |
-リンク
-集合体
登録構造単位 |
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1 |
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-要素
-Proteasome subunit alpha type- ... , 7種, 14分子 AOBPCQDRESFTGU
#1: タンパク質 | 分子量: 25927.535 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P25787, proteasome endopeptidase complex #2: タンパク質 | 分子量: 29525.842 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P25789, proteasome endopeptidase complex #3: タンパク質 | 分子量: 27929.891 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: O14818, proteasome endopeptidase complex #4: タンパク質 | 分子量: 26435.977 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P28066, proteasome endopeptidase complex #5: タンパク質 | 分子量: 29595.627 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P25786, proteasome endopeptidase complex #6: タンパク質 | 分子量: 28469.252 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P25788, proteasome endopeptidase complex #7: タンパク質 | 分子量: 27432.459 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P60900, proteasome endopeptidase complex |
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-Proteasome subunit beta type- ... , 7種, 14分子 HVIWJXKYLZMaNb
#8: タンパク質 | 分子量: 25321.980 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: Q99436, proteasome endopeptidase complex #9: タンパク質 | 分子量: 22972.896 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P49720, proteasome endopeptidase complex #10: タンパク質 | 分子量: 22864.277 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P49721, proteasome endopeptidase complex #11: タンパク質 | 分子量: 22484.369 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P28074, proteasome endopeptidase complex #12: タンパク質 | 分子量: 26522.396 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P20618, proteasome endopeptidase complex #13: タンパク質 | 分子量: 29231.178 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P28070, proteasome endopeptidase complex #14: タンパク質 | 分子量: 21921.836 Da / 分子数: 2 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: P28072, proteasome endopeptidase complex |
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-タンパク質 , 1種, 1分子 c
#15: タンパク質 | 分子量: 211593.344 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: Q14997 |
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-非ポリマー , 2種, 8分子
#16: 化合物 | ChemComp-LDZ / #17: 化合物 | |
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-詳細
研究の焦点であるリガンドがあるか | Y |
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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 |
-試料調製
構成要素 | 名称: PA200-20S proteasome / タイプ: COMPLEX / Entity ID: #1-#15 / 由来: NATURAL |
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由来(天然) | 生物種: Homo sapiens (ヒト) |
緩衝液 | pH: 7.5 |
試料 | 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES |
急速凍結 | 凍結剤: ETHANE |
-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: FIELD EMISSION GUN / 加速電圧: 300 kV / 照射モード: FLOOD BEAM |
電子レンズ | モード: BRIGHT FIELD / 最大 デフォーカス(公称値): 2000 nm / 最小 デフォーカス(公称値): 1000 nm |
撮影 | 電子線照射量: 30 e/Å2 / フィルム・検出器のモデル: GATAN K3 (6k x 4k) |
-解析
CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION |
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3次元再構成 | 解像度: 3.1 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 70412 / 対称性のタイプ: POINT |