+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-41778 | |||||||||
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タイトル | Map from local refinement (focused on CRBN) of DDB1dB:CRBN:PT-179:SD40, conformation 1 | |||||||||
マップデータ | Unsharpened main map | |||||||||
試料 |
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キーワード | ubiquitin (ユビキチン) / CRBN / directed evolution / Zinc finger (ジンクフィンガー) / IMiD / Molecular glue / Ubiquitin ligase (ユビキチンリガーゼ) / TRANSFERASE (転移酵素) | |||||||||
機能・相同性 | 機能・相同性情報 negative regulation of monoatomic ion transmembrane transport / positive regulation by virus of viral protein levels in host cell / epigenetic programming in the zygotic pronuclei / spindle assembly involved in female meiosis / Cul4-RING E3 ubiquitin ligase complex / UV-damage excision repair / biological process involved in interaction with symbiont / regulation of mitotic cell cycle phase transition / WD40-repeat domain binding / Cul4A-RING E3 ubiquitin ligase complex ...negative regulation of monoatomic ion transmembrane transport / positive regulation by virus of viral protein levels in host cell / epigenetic programming in the zygotic pronuclei / spindle assembly involved in female meiosis / Cul4-RING E3 ubiquitin ligase complex / UV-damage excision repair / biological process involved in interaction with symbiont / regulation of mitotic cell cycle phase transition / WD40-repeat domain binding / Cul4A-RING E3 ubiquitin ligase complex / Cul4B-RING E3 ubiquitin ligase complex / ubiquitin ligase complex scaffold activity / negative regulation of reproductive process / negative regulation of developmental process / locomotory exploration behavior / cullin family protein binding / positive regulation of Wnt signaling pathway / viral release from host cell / ectopic germ cell programmed cell death / negative regulation of protein-containing complex assembly / positive regulation of viral genome replication / positive regulation of gluconeogenesis / proteasomal protein catabolic process / nucleotide-excision repair / Recognition of DNA damage by PCNA-containing replication complex / positive regulation of protein-containing complex assembly / DNA Damage Recognition in GG-NER / regulation of circadian rhythm / Dual Incision in GG-NER / Transcription-Coupled Nucleotide Excision Repair (TC-NER) / Formation of TC-NER Pre-Incision Complex / Wntシグナル経路 / Formation of Incision Complex in GG-NER / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / positive regulation of protein catabolic process / cellular response to UV / rhythmic process / protein-macromolecule adaptor activity / site of double-strand break / Neddylation / ubiquitin-dependent protein catabolic process / proteasome-mediated ubiquitin-dependent protein catabolic process / Potential therapeutics for SARS / transmembrane transporter binding / chromosome, telomeric region / damaged DNA binding / protein ubiquitination / DNA修復 / apoptotic process / DNA damage response / protein-containing complex binding / 核小体 / negative regulation of apoptotic process / perinuclear region of cytoplasm / protein-containing complex / DNA binding / extracellular space / extracellular exosome / 核質 / 生体膜 / metal ion binding / 細胞核 / 細胞質基質 / 細胞質 類似検索 - 分子機能 | |||||||||
生物種 | Homo sapiens (ヒト) | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 2.55 Å | |||||||||
データ登録者 | Roy Burman SS / Hunkeler M / Fischer ES | |||||||||
資金援助 | 米国, 1件
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引用 | ジャーナル: Science / 年: 2024 タイトル: Continuous evolution of compact protein degradation tags regulated by selective molecular glues. 著者: Jaron A M Mercer / Stephan J DeCarlo / Shourya S Roy Burman / Vedagopuram Sreekanth / Andrew T Nelson / Moritz Hunkeler / Peter J Chen / Katherine A Donovan / Praveen Kokkonda / Praveen K ...著者: Jaron A M Mercer / Stephan J DeCarlo / Shourya S Roy Burman / Vedagopuram Sreekanth / Andrew T Nelson / Moritz Hunkeler / Peter J Chen / Katherine A Donovan / Praveen Kokkonda / Praveen K Tiwari / Veronika M Shoba / Arghya Deb / Amit Choudhary / Eric S Fischer / David R Liu / 要旨: Conditional protein degradation tags (degrons) are usually >100 amino acids long or are triggered by small molecules with substantial off-target effects, thwarting their use as specific modulators of ...Conditional protein degradation tags (degrons) are usually >100 amino acids long or are triggered by small molecules with substantial off-target effects, thwarting their use as specific modulators of endogenous protein levels. We developed a phage-assisted continuous evolution platform for molecular glue complexes (MG-PACE) and evolved a 36-amino acid zinc finger (ZF) degron (SD40) that binds the ubiquitin ligase substrate receptor cereblon in complex with PT-179, an orthogonal thalidomide derivative. Endogenous proteins tagged in-frame with SD40 using prime editing are degraded by otherwise inert PT-179. Cryo-electron microscopy structures of SD40 in complex with ligand-bound cereblon revealed mechanistic insights into the molecular basis of SD40's activity and specificity. Our efforts establish a system for continuous evolution of molecular glue complexes and provide ZF tags that overcome shortcomings associated with existing degrons. | |||||||||
履歴 |
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-構造の表示
添付画像 |
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-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_41778.map.gz | 63.1 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-41778-v30.xml emd-41778.xml | 28 KB 28 KB | 表示 表示 | EMDBヘッダ |
FSC (解像度算出) | emd_41778_fsc.xml | 10.5 KB | 表示 | FSCデータファイル |
画像 | emd_41778.png | 79.4 KB | ||
マスクデータ | emd_41778_msk_1.map | 125 MB | マスクマップ | |
Filedesc metadata | emd-41778.cif.gz | 7.6 KB | ||
その他 | emd_41778_additional_1.map.gz emd_41778_additional_2.map.gz emd_41778_half_map_1.map.gz emd_41778_half_map_2.map.gz | 109.4 MB 117.9 MB 115.8 MB 115.8 MB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-41778 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-41778 | HTTPS FTP |
-関連構造データ
-リンク
EMDBのページ | EMDB (EBI/PDBe) / EMDataResource |
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「今月の分子」の関連する項目 |
-マップ
ファイル | ダウンロード / ファイル: emd_41778.map.gz / 形式: CCP4 / 大きさ: 125 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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注釈 | Unsharpened main map | ||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 0.8715 Å | ||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||
詳細 | EMDB XML:
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-添付データ
-マスク #1
ファイル | emd_41778_msk_1.map | ||||||||||||
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投影像・断面図 |
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密度ヒストグラム |
-追加マップ: Main map, post-processed with DeepEMhancer
ファイル | emd_41778_additional_1.map | ||||||||||||
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注釈 | Main map, post-processed with DeepEMhancer | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-追加マップ: Main map, sharpened
ファイル | emd_41778_additional_2.map | ||||||||||||
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注釈 | Main map, sharpened | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: half map A
ファイル | emd_41778_half_map_1.map | ||||||||||||
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注釈 | half map A | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: half map B
ファイル | emd_41778_half_map_2.map | ||||||||||||
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注釈 | half map B | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-試料の構成要素
-全体 : Ternary Complex of DDB1dB:CRBN:PT-179 with SD40
全体 | 名称: Ternary Complex of DDB1dB:CRBN:PT-179 with SD40 |
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要素 |
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-超分子 #1: Ternary Complex of DDB1dB:CRBN:PT-179 with SD40
超分子 | 名称: Ternary Complex of DDB1dB:CRBN:PT-179 with SD40 / タイプ: complex / ID: 1 / 親要素: 0 / 含まれる分子: all |
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分子量 | 理論値: 201 KDa |
-分子 #1: DNA damage-binding protein 1
分子 | 名称: DNA damage-binding protein 1 / タイプ: protein_or_peptide / ID: 1 / 光学異性体: LEVO |
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由来(天然) | 生物種: Homo sapiens (ヒト) |
組換発現 | 生物種: Trichoplusia ni (イラクサキンウワバ) |
配列 | 文字列: MGSSHHHHHH SAVDENLYFQ GGGRMSYNYV VTAQKPTAVN GCVTGHFTSA EDLNLLIAKN TRLEIYVVTA EGLRPVKEVG MYGKIAVME LFRPKGESKD LLFILTAKYN ACILEYKQSG ESIDIITRAH GNVQDRIGRP SETGIIGIID PECRMIGLRL Y DGLFKVIP ...文字列: MGSSHHHHHH SAVDENLYFQ GGGRMSYNYV VTAQKPTAVN GCVTGHFTSA EDLNLLIAKN TRLEIYVVTA EGLRPVKEVG MYGKIAVME LFRPKGESKD LLFILTAKYN ACILEYKQSG ESIDIITRAH GNVQDRIGRP SETGIIGIID PECRMIGLRL Y DGLFKVIP LDRDNKELKA FNIRLEELHV IDVKFLYGCQ APTICFVYQD PQGRHVKTYE VSLREKEFNK GPWKQENVEA EA SMVIAVP EPFGGAIIIG QESITYHNGD KYLAIAPPII KQSTIVCHNR VDPNGSRYLL GDMEGRLFML LLEKEEQMDG TVT LKDLRV ELLGETSIAE CLTYLDNGVV FVGSRLGDSQ LVKLNVDSNE QGSYVVAMET FTNLGPIVDM CVVDLERQGQ GQLV TCSGA FKEGSLRIIR NGIGGNGNSG EIQKLHIRTV PLYESPRKIC YQEVSQCFGV LSSRIEVQDT SGGTTALRPS ASTQA LSSS VSSSKLFSSS TAPHETSFGE EVEVHNLLII DQHTFEVLHA HQFLQNEYAL SLVSCKLGKD PNTYFIVGTA MVYPEE AEP KQGRIVVFQY SDGKLQTVAE KEVKGAVYSM VEFNGKLLAS INSTVRLYEW TTEKELRTEC NHYNNIMALY LKTKGDF IL VGDLMRSVLL LAYKPMEGNF EEIARDFNPN WMSAVEILDD DNFLGAENAF NLFVCQKDSA ATTDEERQHL QEVGLFHL G EFVNVFCHGS LVMQNLGETS TPTQGSVLFG TVNGMIGLVT SLSESWYNLL LDMQNRLNKV IKSVGKIEHS FWRSFHTER KTEPATGFID GDLIESFLDI SRPKMQEVVA NLQYDDGSGM KREATADDLI KVVEELTRIH UniProtKB: DNA damage-binding protein 1 |
-分子 #2: Protein cereblon
分子 | 名称: Protein cereblon / タイプ: protein_or_peptide / ID: 2 / 光学異性体: LEVO |
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由来(天然) | 生物種: Homo sapiens (ヒト) |
組換発現 | 生物種: Trichoplusia ni (イラクサキンウワバ) |
配列 | 文字列: MDYKDDDDKS AVDENLYFQG GGRGGSAHIV MVDAYKPTKG GSGMAGEGDQ QDAAHNMGNH LPLLPAESEE EDEMEVEDQD SKEAKKPNI INFDTSLPTS HTYLGADMEE FHGRTLHDDD SCQVIPVLPQ VMMILIPGQT LPLQLFHPQE VSMVRNLIQK D RTFAVLAY ...文字列: MDYKDDDDKS AVDENLYFQG GGRGGSAHIV MVDAYKPTKG GSGMAGEGDQ QDAAHNMGNH LPLLPAESEE EDEMEVEDQD SKEAKKPNI INFDTSLPTS HTYLGADMEE FHGRTLHDDD SCQVIPVLPQ VMMILIPGQT LPLQLFHPQE VSMVRNLIQK D RTFAVLAY SNVQEREAQF GTTAEIYAYR EEQDFGIEIV KVKAIGRQRF KVLELRTQSD GIQQAKVQIL PECVLPSTMS AV QLESLNK CQIFPSKPVS REDQCSYKWW QKYQKRKFHC ANLTSWPRWL YSLYDAETLM DRIKKQLREW DENLKDDSLP SNP IDFSYR VAACLPIDDV LRIQLLKIGS AIQRLRCELD IMNKCTSLCC KQCQETEITT KNEIFSLSLC GPMAAYVNPH GYVH ETLTV YKACNLNLIG RPSTEHSWFP GYAWTVAQCK ICASHIGWKF TATKKDMSPQ KFWGLTRSAL LPTIPDTEDE ISPDK VILC L UniProtKB: Protein cereblon |
-分子 #3: Maltose/maltodextrin-binding periplasmic protein,SD40
分子 | 名称: Maltose/maltodextrin-binding periplasmic protein,SD40 タイプ: protein_or_peptide / ID: 3 詳細: IKZF1/ZFP91 fusion construct that was further engineered to enhance binding of cereblon/DDB1 in the presence of IMiD derivatives 光学異性体: LEVO |
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由来(天然) | 生物種: Homo sapiens (ヒト) |
組換発現 | 生物種: Escherichia coli (大腸菌) |
配列 | 文字列: MGLNDIFEAQ KIEWHEGSSH HHHHHGSSKI EEGKLVIWIN GDKGYNGLAE VGKKFEKDTG IKVTVEHPDK LEEKFPQVAA TGDGPDIIF WAHDRFGGYA QSGLLAEITP DKAFQDKLYP FTWDAVRYNG KLIAYPIAVE ALSLIYNKDL LPNPPKTWEE I PALDKELK ...文字列: MGLNDIFEAQ KIEWHEGSSH HHHHHGSSKI EEGKLVIWIN GDKGYNGLAE VGKKFEKDTG IKVTVEHPDK LEEKFPQVAA TGDGPDIIF WAHDRFGGYA QSGLLAEITP DKAFQDKLYP FTWDAVRYNG KLIAYPIAVE ALSLIYNKDL LPNPPKTWEE I PALDKELK AKGKSALMFN LQEPYFTWPL IAADGGYAFK YENGKYDIKD VGVDNAGAKA GLTFLVDLIK NKHMNADTDY SI AEAAFNK GETAMTINGP WAWSNIDTSK VNYGVTVLPT FKGQPSKPFV GVLSAGINAA SPNKELAKEF LENYLLTDEG LEA VNKDKP LGAVALKSYE EELAKDPRIA ATMENAQKGE IMPNIPQMSA FWYAVRTAVI NAASGRQTVD EALKDAQTRI TKLE VLFQG PDYKDDDDKS GGGGLLLFCP ICGFTCRQKG NLLRHINLHT GEKLFKYHLY |
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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解析 | 単粒子再構成法 |
試料の集合状態 | particle |
-試料調製
濃度 | 0.2625 mg/mL | ||||||||||||
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緩衝液 | pH: 7 構成要素:
詳細: 20 mM HEPES/NaOH pH 7.0, 150 mM NaCl, and 3 mM TCEP. DMSO concentrations were kept below 2% (v/v) | ||||||||||||
グリッド | モデル: Quantifoil R1.2/1.3 / 材質: GOLD / メッシュ: 300 / 支持フィルム - 材質: GOLD / 支持フィルム - トポロジー: HOLEY ARRAY / 支持フィルム - Film thickness: 50 / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 時間: 120 sec. / 前処理 - 雰囲気: AIR / 前処理 - 気圧: 0.039 kPa 詳細: Grids (Quantifoil UltrAuFoil R 1.2/1.3) were glow discharged in PELCO easiGlow (20 mA, 120s, 39 Pa) | ||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 90 % / チャンバー内温度: 283 K / 装置: LEICA EM GP 詳細: Leica EM-GP plunge freezer with chamber conditions of 10 C and 90% relative humidity. Grids were first pre-incubated with 4 uL of 10 uM CRBN-agnostic IKZF1_140-196_Q146A,G151N for 1 minute ...詳細: Leica EM-GP plunge freezer with chamber conditions of 10 C and 90% relative humidity. Grids were first pre-incubated with 4 uL of 10 uM CRBN-agnostic IKZF1_140-196_Q146A,G151N for 1 minute and then blotted from behind for 4 s. Immediately, 4 uL of mixture 1 diluted 10-fold--with the dilution buffer during the 1-minute incubation time--was applied to the grids before blotting for 4 s and plunging into liquid ethane at -181 C.. | ||||||||||||
詳細 | DDB1dB_CRBN, PT-179, and SD40 were mixed and incubated on ice for 1 hour at final concentration of 10.5, 105, and 21 uM, respectively. Then diluted 10-fold before blotting. |
-電子顕微鏡法
顕微鏡 | TFS KRIOS |
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電子線 | 加速電圧: 300 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | C2レンズ絞り径: 50.0 µm / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / 最大 デフォーカス(公称値): 2.0 µm / 最小 デフォーカス(公称値): 0.8 µm / 倍率(公称値): 105000 |
特殊光学系 | エネルギーフィルター - 名称: GIF Bioquantum / エネルギーフィルター - スリット幅: 20 eV |
試料ステージ | 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER ホルダー冷却材: NITROGEN |
撮影 | フィルム・検出器のモデル: GATAN K3 BIOQUANTUM (6k x 4k) 撮影したグリッド数: 1 / 実像数: 16340 / 平均露光時間: 2.497 sec. / 平均電子線量: 52.0 e/Å2 詳細: Movies (50 frames, two movies per hole) collected using beam-shift with 9 holes per stage position (3x3 pattern) |
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |
-画像解析
-原子モデル構築 1
精密化 | プロトコル: OTHER |
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