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データを開く
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基本情報
登録情報 | ![]() | |||||||||
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タイトル | G4 RNA-mediated PRC2 dimer | |||||||||
![]() | consensus map of G4 RNA-mediated PRC2 dimer | |||||||||
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![]() | PRC2 / G-quadruplex RNA / RNP complex / chromatin modifier / GENE REGULATION | |||||||||
機能・相同性 | ![]() regulation of kidney development / hepatocyte homeostasis / cellular response to trichostatin A / regulation of gliogenesis / [histone H3]-lysine27 N-trimethyltransferase / negative regulation of striated muscle cell differentiation / regulatory ncRNA-mediated heterochromatin formation / sex chromatin / CAF-1 complex / negative regulation of keratinocyte differentiation ...regulation of kidney development / hepatocyte homeostasis / cellular response to trichostatin A / regulation of gliogenesis / [histone H3]-lysine27 N-trimethyltransferase / negative regulation of striated muscle cell differentiation / regulatory ncRNA-mediated heterochromatin formation / sex chromatin / CAF-1 complex / negative regulation of keratinocyte differentiation / histone H3K27 trimethyltransferase activity / negative regulation of retinoic acid receptor signaling pathway / random inactivation of X chromosome / primary miRNA binding / skeletal muscle satellite cell maintenance involved in skeletal muscle regeneration / response to tetrachloromethane / cerebellar cortex development / facultative heterochromatin formation / histone H3K27 methyltransferase activity / positive regulation of cell cycle G1/S phase transition / NURF complex / regulation of cell fate specification / negative regulation of stem cell population maintenance / DNA replication-dependent chromatin assembly / chromatin silencing complex / NuRD complex / Transcription of E2F targets under negative control by p107 (RBL1) and p130 (RBL2) in complex with HDAC1 / regulation of stem cell differentiation / ESC/E(Z) complex / protein-lysine N-methyltransferase activity / negative regulation of stem cell differentiation / Transcription of E2F targets under negative control by DREAM complex / pronucleus / RSC-type complex / cardiac muscle hypertrophy in response to stress / Polo-like kinase mediated events / synaptic transmission, GABAergic / lncRNA binding / positive regulation of dendrite development / histone H3 methyltransferase activity / negative regulation of G1/S transition of mitotic cell cycle / negative regulation of gene expression, epigenetic / spinal cord development / G1 to G0 transition / Sin3-type complex / histone methyltransferase activity / positive regulation of stem cell population maintenance / G1/S-Specific Transcription / ATPase complex / oligodendrocyte differentiation / negative regulation of transcription elongation by RNA polymerase II / Transcriptional Regulation by E2F6 / RNA Polymerase I Transcription Initiation / negative regulation of cell differentiation / histone deacetylase complex / G0 and Early G1 / subtelomeric heterochromatin formation / negative regulation of cytokine production involved in inflammatory response / RNA polymerase II core promoter sequence-specific DNA binding / pericentric heterochromatin / ribonucleoprotein complex binding / positive regulation of epithelial to mesenchymal transition / Cyclin E associated events during G1/S transition / Cyclin A:Cdk2-associated events at S phase entry / nucleosome binding / keratinocyte differentiation / Deposition of new CENPA-containing nucleosomes at the centromere / protein localization to chromatin / enzyme activator activity / Regulation of TP53 Activity through Acetylation / methylated histone binding / SUMOylation of chromatin organization proteins / negative regulation of cell migration / B cell differentiation / transcription corepressor binding / ERCC6 (CSB) and EHMT2 (G9a) positively regulate rRNA expression / positive regulation of GTPase activity / PRC2 methylates histones and DNA / Regulation of PTEN gene transcription / Defective pyroptosis / HDACs deacetylate histones / liver regeneration / stem cell differentiation / promoter-specific chromatin binding / hippocampus development / transcription coregulator activity / heterochromatin formation / negative regulation of transforming growth factor beta receptor signaling pathway / positive regulation of MAP kinase activity / protein modification process / positive regulation of protein serine/threonine kinase activity / regulation of circadian rhythm / brain development / chromatin DNA binding / PKMTs methylate histone lysines / histone deacetylase binding / cellular response to hydrogen peroxide / Activation of anterior HOX genes in hindbrain development during early embryogenesis / HCMV Early Events / G1/S transition of mitotic cell cycle 類似検索 - 分子機能 | |||||||||
生物種 | ![]() | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 3.4 Å | |||||||||
![]() | Song J / Kasinath V | |||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Structural basis for inactivation of PRC2 by G-quadruplex RNA. 著者: Jiarui Song / Anne R Gooding / Wayne O Hemphill / Brittney D Love / Anne Robertson / Liqi Yao / Leonard I Zon / Trista E North / Vignesh Kasinath / Thomas R Cech / ![]() 要旨: Polycomb repressive complex 2 (PRC2) silences genes through trimethylation of histone H3K27. PRC2 associates with numerous precursor messenger RNAs (pre-mRNAs) and long noncoding RNAs (lncRNAs) with ...Polycomb repressive complex 2 (PRC2) silences genes through trimethylation of histone H3K27. PRC2 associates with numerous precursor messenger RNAs (pre-mRNAs) and long noncoding RNAs (lncRNAs) with a binding preference for G-quadruplex RNA. In this work, we present a 3.3-Å-resolution cryo-electron microscopy structure of PRC2 bound to a G-quadruplex RNA. Notably, RNA mediates the dimerization of PRC2 by binding both protomers and inducing a protein interface composed of two copies of the catalytic subunit EZH2, thereby blocking nucleosome DNA interaction and histone H3 tail accessibility. Furthermore, an RNA-binding loop of EZH2 facilitates the handoff between RNA and DNA, another activity implicated in PRC2 regulation by RNA. We identified a gain-of-function mutation in this loop that activates PRC2 in zebrafish. Our results reveal mechanisms for RNA-mediated regulation of a chromatin-modifying enzyme. | |||||||||
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 | ![]() | 80.2 MB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 31.6 KB 31.6 KB | 表示 表示 | ![]() |
FSC (解像度算出) | ![]() ![]() ![]() | 10.7 KB 10.7 KB 10.6 KB | 表示 表示 表示 | ![]() |
画像 | ![]() | 117.5 KB | ||
マスクデータ | ![]() | 103 MB | ![]() | |
Filedesc metadata | ![]() | 9.7 KB | ||
その他 | ![]() ![]() ![]() | 8.3 MB 80.9 MB 80.8 MB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-検証レポート
文書・要旨 | ![]() | 877.5 KB | 表示 | ![]() |
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文書・詳細版 | ![]() | 877.1 KB | 表示 | |
XML形式データ | ![]() | 17.9 KB | 表示 | |
CIF形式データ | ![]() | 23.7 KB | 表示 | |
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
関連構造データ | ![]() 8fyhMC C: 同じ文献を引用 ( M: このマップから作成された原子モデル |
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類似構造データ | 類似検索 - 機能・相同性 ![]() |
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リンク
EMDBのページ | ![]() ![]() |
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「今月の分子」の関連する項目 |
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マップ
ファイル | ![]() | ||||||||||||||||||||
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注釈 | consensus map of G4 RNA-mediated PRC2 dimer | ||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 1.06344 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 | ![]() | ||||||||||||
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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: consensus map of G4 RNA-mediated PRC2 dimer after...
ファイル | emd_29578_additional_1.map | ||||||||||||
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注釈 | consensus map of G4 RNA-mediated PRC2 dimer after postprocessing with B-factor of -40 | ||||||||||||
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 2 of consensus map
ファイル | emd_29578_half_map_1.map | ||||||||||||
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注釈 | half map 2 of consensus map | ||||||||||||
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 1 of consensus map
ファイル | emd_29578_half_map_2.map | ||||||||||||
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注釈 | half map 1 of consensus map | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
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試料の構成要素
-全体 : G4 RNA-mediated dimer of polycomb repressive complex 2
全体 | 名称: G4 RNA-mediated dimer of polycomb repressive complex 2 |
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要素 |
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-超分子 #1: G4 RNA-mediated dimer of polycomb repressive complex 2
超分子 | 名称: G4 RNA-mediated dimer of polycomb repressive complex 2 タイプ: complex / ID: 1 / 親要素: 0 / 含まれる分子: #1-#7 |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 800 KDa |
-分子 #1: Polycomb protein SUZ12
分子 | 名称: Polycomb protein SUZ12 / タイプ: protein_or_peptide / ID: 1 / コピー数: 2 / 光学異性体: LEVO |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 83.181922 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MAPQKHGGGG GGGSGPSAGS GGGGFGGSAA VAAATASGGK SGGGSCGGGG SYSASSSSSA AAAAGAAVLP VKKPKMEHVQ ADHELFLQA FEKPTQIYRF LRTRNLIAPI FLHRTLTYMS HRNSRTNIKR KTFKVDDMLS KVEKMKGEQE SHSLSAHLQL T FTGFFHKN ...文字列: MAPQKHGGGG GGGSGPSAGS GGGGFGGSAA VAAATASGGK SGGGSCGGGG SYSASSSSSA AAAAGAAVLP VKKPKMEHVQ ADHELFLQA FEKPTQIYRF LRTRNLIAPI FLHRTLTYMS HRNSRTNIKR KTFKVDDMLS KVEKMKGEQE SHSLSAHLQL T FTGFFHKN DKPSPNSENE QNSVTLEVLL VKVCHKKRKD VSCPIRQVPT GKKQVPLNPD LNQTKPGNFP SLAVSSNEFE PS NSHMVKS YSLLFRVTRP GRREFNGMIN GETNENIDVN EELPARRKRN REDGEKTFVA QMTVFDKNRR LQLLDGEYEV AMQ EMEECP ISKKRATWET ILDGKRLPPF ETFSQGPTLQ FTLRWTGETN DKSTAPIAKP LATRNSESLH QENKPGSVKP TQTI AVKES LTTDLQTRKE KDTPNENRQK LRIFYQFLYN NNTRQQTEAR DDLHCPWCTL NCRKLYSLLK HLKLCHSRFI FNYVY HPKG ARIDVSINEC YDGSYAGNPQ DIHRQPGFAF SRNGPVKRTP ITHILVCRPK RTKASMSEFL ESEDGEVEQQ RTYSSG HNR LYFHSDTCLP LRPQEMEVDS EDEKDPEWLR EKTITQIEEF SDVNEGEKEV MKLWNLHVMK HGFIADNQMN HACMLFV EN YGQKIIKKNL CRNFMLHLVS MHDFNLISIM SIDKAVTKLR EMQQKLEKGE SASPANEEIT EEQNGTANGF SEINSKEK A LETDSVSGVS KQSKKQKL UniProtKB: Polycomb protein SUZ12 |
-分子 #2: Polycomb protein EED
分子 | 名称: Polycomb protein EED / タイプ: protein_or_peptide / ID: 2 / コピー数: 2 / 光学異性体: LEVO |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 50.267691 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MSEREVSTAP AGTDMPAAKK QKLSSDENSN PDLSGDENDD AVSIESGTNT ERPDTPTNTP NAPGRKSWGK GKWKSKKCKY SFKCVNSLK EDHNQPLFGV QFNWHSKEGD PLVFATVGSN RVTLYECHSQ GEIRLLQSYV DADADENFYT CAWTYDSNTS H PLLAVAGS ...文字列: MSEREVSTAP AGTDMPAAKK QKLSSDENSN PDLSGDENDD AVSIESGTNT ERPDTPTNTP NAPGRKSWGK GKWKSKKCKY SFKCVNSLK EDHNQPLFGV QFNWHSKEGD PLVFATVGSN RVTLYECHSQ GEIRLLQSYV DADADENFYT CAWTYDSNTS H PLLAVAGS RGIIRIINPI TMQCIKHYVG HGNAINELKF HPRDPNLLLS VSKDHALRLW NIQTDTLVAI FGGVEGHRDE VL SADYDLL GEKIMSCGMD HSLKLWRINS KRMMNAIKES YDYNPNKTNR PFISQKIHFP DFSTRDIHRN YVDCVRWLGD LIL SKSCEN AIVCWKPGKM EDDIDKIKPS ESNVTILGRF DYSQCDIWYM RFSMDFWQKM LALGNQVGKL YVWDLEVEDP HKAK CTTLT HHKCGAAIRQ TSFSRDSSIL IAVCDDASIW RWDRLR UniProtKB: Polycomb protein EED |
-分子 #3: Histone-binding protein RBBP4
分子 | 名称: Histone-binding protein RBBP4 / タイプ: protein_or_peptide / ID: 3 / コピー数: 2 / 光学異性体: LEVO |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 47.709527 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MADKEAAFDD AVEERVINEE YKIWKKNTPF LYDLVMTHAL EWPSLTAQWL PDVTRPEGKD FSIHRLVLGT HTSDEQNHLV IASVQLPND DAQFDASHYD SEKGEFGGFG SVSGKIEIEI KINHEGEVNR ARYMPQNPCI IATKTPSSDV LVFDYTKHPS K PDPSGECN ...文字列: MADKEAAFDD AVEERVINEE YKIWKKNTPF LYDLVMTHAL EWPSLTAQWL PDVTRPEGKD FSIHRLVLGT HTSDEQNHLV IASVQLPND DAQFDASHYD SEKGEFGGFG SVSGKIEIEI KINHEGEVNR ARYMPQNPCI IATKTPSSDV LVFDYTKHPS K PDPSGECN PDLRLRGHQK EGYGLSWNPN LSGHLLSASD DHTICLWDIS AVPKEGKVVD AKTIFTGHTA VVEDVSWHLL HE SLFGSVA DDQKLMIWDT RSNNTSKPSH SVDAHTAEVN CLSFNPYSEF ILATGSADKT VALWDLRNLK LKLHSFESHK DEI FQVQWS PHNETILASS GTDRRLNVWD LSKIGEEQSP EDAEDGPPEL LFIHGGHTAK ISDFSWNPNE PWVICSVSED NIMQ VWQMA ENIYNDEDPE GSVDPEGQGS UniProtKB: Histone-binding protein RBBP4 |
-分子 #4: Histone-lysine N-methyltransferase EZH2
分子 | 名称: Histone-lysine N-methyltransferase EZH2 / タイプ: protein_or_peptide / ID: 4 / コピー数: 2 / 光学異性体: LEVO / EC番号: [histone H3]-lysine27 N-trimethyltransferase |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 86.149055 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MGQTGKKSEK GPVCWRKRVK SEYMRLRQLK RFRRADEVKS MFSSNRQKIL ERTEILNQEW KQRRIQPVHI LTSVSSLRGT RECSVTSDL DFPTQVIPLK TLNAVASVPI MYSWSPLQQN FMVEDETVLH NIPYMGDEVL DQDGTFIEEL IKNYDGKVHG D RECGFIND ...文字列: MGQTGKKSEK GPVCWRKRVK SEYMRLRQLK RFRRADEVKS MFSSNRQKIL ERTEILNQEW KQRRIQPVHI LTSVSSLRGT RECSVTSDL DFPTQVIPLK TLNAVASVPI MYSWSPLQQN FMVEDETVLH NIPYMGDEVL DQDGTFIEEL IKNYDGKVHG D RECGFIND EIFVELVNAL GQYNDDDDDD DGDDPEEREE KQKDLEDHRD DKESRPPRKF PSDKIFEAIS SMFPDKGTAE EL KEKYKEL TEQQLPGALP PECTPNIDGP NAKSVQREQS LHSFHTLFCR RCFKYDCFLH RKCNYSFHAT PNTYKRKNTE TAL DNKPCG PQCYQHLEGA KEFAAALTAE RIKTPPKRPG GRRRGRLPNN SSRPSTPTIN VLESKDTDSD REAGTETGGE NNDK EEEEK KDETSSSSEA NSRCQTPIKM KPNIEPPENV EWSGAEASMF RVLIGTYYDN FCAIARLIGT KTCRQVYEFR VKESS IIAP APAEDVDTPP RKKKRKHRLW AAHCRKIQLK KDGSSNHVYN YQPCDHPRQP CDSSCPCVIA QNFCEKFCQC SSECQN RFP GCRCKAQCNT KQCPCYLAVR ECDPDLCLTC GAADHWDSKN VSCKNCSIQR GSKKHLLLAP SDVAGWGIFI KDPVQKN EF ISEYCGEIIS QDEADRRGKV YDKYMCSFLF NLNNDFVVDA TRKGNKIRFA NHSVNPNCYA KVMMVNGDHR IGIFAKRA I QTGEELFFDY RYSQADALKY VGIEREMEIP UniProtKB: Histone-lysine N-methyltransferase EZH2 |
-分子 #5: protein Jumonji isoform X3
分子 | 名称: protein Jumonji isoform X3 / タイプ: protein_or_peptide / ID: 5 / コピー数: 2 / 光学異性体: LEVO |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 138.406219 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MSKERPKRNI IQKKYDDSDG IPWSEERVVR KVLYLSLKEF KNAQKRQHGE GIAGSLKSVN GLLGNDQSKA LGPASEQSEN EKDDASQVS STSNDVSSSD FEEGPSRKRP RLQAQRKFAQ SQPNSPSTTP VKTVEPLLPP PATQISDLSK RKPKTEDFLT F LCLRGSPA ...文字列: MSKERPKRNI IQKKYDDSDG IPWSEERVVR KVLYLSLKEF KNAQKRQHGE GIAGSLKSVN GLLGNDQSKA LGPASEQSEN EKDDASQVS STSNDVSSSD FEEGPSRKRP RLQAQRKFAQ SQPNSPSTTP VKTVEPLLPP PATQISDLSK RKPKTEDFLT F LCLRGSPA LPSSMVYFGS SQDEEDVEEE DDETEDVKTA NNNASSSCQS TPRKGKTHKH VHNGHVFNGS NRSTREKEPA QK HKSKETT PAKEKHIDHR ADSRREPASV AQPTATPSAG SLAKGLPANH QPPPPHRSAQ DLRKQVTLHV SKVNGVTRMS SLG AGTTSA KKIREVRPSP SKTVKYTATV TKGTVTYTKA KRELVKETKP THHKPSSAVN HTISGKTESS NAKTRKQVLS LGGA STSTG PAASGLKASS RLNPKSCTKE VGGRQLREGL RNSKRRLEEA QQVDKPQSPP KKMKGAAGIA EAPGKKASAA SAEKS LLNG HVKKEVPERS LERNRPKRAT AGKNMPGKQA HGKAEGTPCE NRSTSQPESS HKPHDPQGKP EKGIGKSGWT AMDEIP VLR PSAKEFHDPL IYIESVRAQV EKYGMCRVIP PPDWRPECKL NDEMRFVTQI QHIHKLGRRW GPNVQRLACI KKHLRSQ GI TMDELPLIGG CELDLACFFR LINEMGGMQQ VTDLKKWNKL ADMLRIPKTA QDRLAKLQEA YCQYLLSYDS LSPEEHRR L EKEVLMEKEI LEKRKGPLEG HTENDHHKFH SLPRFEPKNG LIHGVTPRNG FRSKLKEVGQ APLKTGRRRL FAQEKEVVK EEEEDKGVLN DFHKCIYKGR SVSLTTFYRT ARNIMNMCFS KEPAPAEIEQ EYWRLVEEKD CHVAVHCGKV DTNTHGSGFP VGKSEPFSR HGWNLTVLPN NTGSILRHLG AVPGVTIPWL NIGMVFSTSC WSRDQNHLPY IDYLHTGADC IWYCIPAEEE N KLEDVVHT LLQANGTPGL QMLESNVMIS PEVLCKEGIK VHRTVQQSGQ FVVCFPGSFV SKVCCGYSVS ETVHFATTQW TS MGFETAK EMKRRHIAKP FSMEKLLYQI AQAEAKKENG PTLSTISALL DELRDTELRQ RRQLFEAGLH SSARYGSHDG NST VADGKK KPRKWLQLET SERRCQICQH LCYLSMVVQE NENVVFCLEC ALRHVEKQKS CRGLKLMYRY DEEQIISLVN QICG KVSGK HGGIENCLNK PTPKRGPRKR ATVDVPPSRL PSS UniProtKB: Protein Jumonji isoform X3 |
-分子 #6: Zinc finger protein AEBP2
分子 | 名称: Zinc finger protein AEBP2 / タイプ: protein_or_peptide / ID: 6 / コピー数: 2 / 光学異性体: LEVO |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 54.535496 KDa |
組換発現 | 生物種: ![]() |
配列 | 文字列: MAAAITDMAD LEELSRLSPL PPGSPGSAAR GRAEPPEEEE EEEEEEEEAE AEAVAALLLN GGSGGGGGGG GGGVGGGEAE TMSEPSPES ASQAGEDEDE EEDDEEEEDE SSSSGGGEEE SSAESLVGSS GGSSSDETRS LSPGAASSSS GDGDGKEGLE E PKGPRGSQ ...文字列: MAAAITDMAD LEELSRLSPL PPGSPGSAAR GRAEPPEEEE EEEEEEEEAE AEAVAALLLN GGSGGGGGGG GGGVGGGEAE TMSEPSPES ASQAGEDEDE EEDDEEEEDE SSSSGGGEEE SSAESLVGSS GGSSSDETRS LSPGAASSSS GDGDGKEGLE E PKGPRGSQ GGGGGGSSSS SVVSSGGDEG YGTGGGGSSA TSGGRRGSLE MSSDGEPLSR MDSEDSISST IMDVDSTISS GR STPAMMN GQGSTTSSSK NIAYNCCWDQ CQACFNSSPD LADHIRSIHV DGQRGGVFVC LWKGCKVYNT PSTSQSWLQR HML THSGDK PFKCVVGGCN ASFASQGGLA RHVPTHFSQQ NSSKVSSQPK AKEESPSKAG MNKRRKLKNK RRRSLPRPHD FFDA QTLDA IRHRAICFNL SAHIESLGKG HSVVFHSTVI AKRKEDSGKI KLLLHWMPED ILPDVWVNES ERHQLKTKVV HLSKL PKDT ALLLDPNIYR TMPQKRLKRT LIRKVFNLYL SKQ UniProtKB: Zinc finger protein AEBP2 |
-分子 #7: G4 RNA
分子 | 名称: G4 RNA / タイプ: rna / ID: 7 / コピー数: 1 |
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由来(天然) | 生物種: ![]() |
分子量 | 理論値: 7.955823 KDa |
配列 | 文字列: GGGUAAGGGU AAGGGUAAGG GUAA |
-分子 #8: ZINC ION
分子 | 名称: ZINC ION / タイプ: ligand / ID: 8 / コピー数: 14 / 式: ZN |
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分子量 | 理論値: 65.409 Da |
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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![]() | 単粒子再構成法 |
試料の集合状態 | particle |
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試料調製
緩衝液 | pH: 7.9 構成要素:
詳細: RNP complex buffer (25 mM HEPES pH 7.9, 50 mM KCl, 2 mM MgCl2, 10% glycerol, and 1mM TCEP) EM preparation buffer I (25 mM HEPES pH 7.9, 50 mM KCl, 2.5% glycerol, and 1mM TCEP) EM preparation ...詳細: RNP complex buffer (25 mM HEPES pH 7.9, 50 mM KCl, 2 mM MgCl2, 10% glycerol, and 1mM TCEP) EM preparation buffer I (25 mM HEPES pH 7.9, 50 mM KCl, 2.5% glycerol, and 1mM TCEP) EM preparation buffer II (25 mM HEPES pH 7.9, 50 mM KCl, 2.5% glycerol, 0.01%NP-40, and 1mM TCEP). | ||||||||||||||||||
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グリッド | モデル: Quantifoil R2/1 / 材質: GOLD / メッシュ: 300 / 支持フィルム - 材質: CARBON / 支持フィルム - トポロジー: CONTINUOUS / 支持フィルム - Film thickness: 1.5 | ||||||||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 90 % / チャンバー内温度: 281 K / 装置: LEICA PLUNGER / 詳細: 3s of single side blotting. | ||||||||||||||||||
詳細 | We used streptavidin-affinity grid preparation method with biotin-labeled RNA at 100 nM concentration. PRC2 was applied in excess at 600 nM. |
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電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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特殊光学系 | エネルギーフィルター - スリット幅: 20 eV |
撮影 | フィルム・検出器のモデル: GATAN K3 (6k x 4k) / 撮影したグリッド数: 1 / 実像数: 18632 / 平均電子線量: 60.0 e/Å2 / 詳細: 60 frames per movie |
電子線 | 加速電圧: 300 kV / 電子線源: ![]() |
電子光学系 | 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 0.01 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): 0.6 µm / 倍率(公称値): 81000 |
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 |
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |