+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 8g0l | ||||||
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タイトル | Semi-synthetic CoA-alpha-Synuclein Constructs Trap N-terminal Acetyltransferase NatB for Binding Mechanism Studies | ||||||
要素 |
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キーワード | TRANSFERASE (転移酵素) / N-terminal acetyltransferase | ||||||
機能・相同性 | 機能・相同性情報 N-terminal peptidyl-glutamine acetylation / N-terminal methionine Nalpha-acetyltransferase NatB / N-terminal peptidyl-aspartic acid acetylation / N-terminal peptidyl-glutamic acid acetylation / NatB complex / N-terminal protein amino acid acetylation / peptide alpha-N-acetyltransferase activity / negative regulation of mitochondrial electron transport, NADH to ubiquinone / neutral lipid metabolic process / regulation of phospholipase activity ...N-terminal peptidyl-glutamine acetylation / N-terminal methionine Nalpha-acetyltransferase NatB / N-terminal peptidyl-aspartic acid acetylation / N-terminal peptidyl-glutamic acid acetylation / NatB complex / N-terminal protein amino acid acetylation / peptide alpha-N-acetyltransferase activity / negative regulation of mitochondrial electron transport, NADH to ubiquinone / neutral lipid metabolic process / regulation of phospholipase activity / negative regulation of monooxygenase activity / regulation of acyl-CoA biosynthetic process / negative regulation of dopamine uptake involved in synaptic transmission / negative regulation of norepinephrine uptake / positive regulation of glutathione peroxidase activity / positive regulation of SNARE complex assembly / positive regulation of hydrogen peroxide catabolic process / supramolecular fiber / negative regulation of transporter activity / mitochondrial membrane organization / negative regulation of chaperone-mediated autophagy / regulation of reactive oxygen species biosynthetic process / positive regulation of protein localization to cell periphery / regulation of synaptic vesicle recycling / negative regulation of platelet-derived growth factor receptor signaling pathway / negative regulation of exocytosis / regulation of glutamate secretion / response to iron(II) ion / regulation of norepinephrine uptake / dopamine biosynthetic process / SNARE complex assembly / positive regulation of neurotransmitter secretion / regulation of locomotion / synaptic vesicle priming / positive regulation of inositol phosphate biosynthetic process / regulation of macrophage activation / mitochondrial ATP synthesis coupled electron transport / dopamine uptake involved in synaptic transmission / negative regulation of microtubule polymerization / synaptic vesicle transport / dynein complex binding / positive regulation of receptor recycling / regulation of dopamine secretion / protein kinase inhibitor activity / negative regulation of thrombin-activated receptor signaling pathway / response to type II interferon / cuprous ion binding / positive regulation of exocytosis / synaptic vesicle exocytosis / positive regulation of endocytosis / cysteine-type endopeptidase inhibitor activity involved in apoptotic process / response to magnesium ion / kinesin binding / regulation of presynapse assembly / synaptic vesicle endocytosis / alpha-tubulin binding / negative regulation of serotonin uptake / localization / phospholipid metabolic process / axon terminus / supramolecular fiber organization / 封入体 / cellular response to copper ion / Hsp70 protein binding / cellular response to epinephrine stimulus / excitatory postsynaptic potential / response to interleukin-1 / adult locomotory behavior / SNARE binding / positive regulation of release of sequestered calcium ion into cytosol / fatty acid metabolic process / long-term synaptic potentiation / regulation of transmembrane transporter activity / protein tetramerization / ferrous iron binding / phosphoprotein binding / synapse organization / regulation of long-term neuronal synaptic plasticity / microglial cell activation / negative regulation of protein kinase activity / phospholipid binding / protein destabilization / PKR-mediated signaling / negative regulation of cysteine-type endopeptidase activity involved in apoptotic process / tau protein binding / positive regulation of protein serine/threonine kinase activity / receptor internalization / synaptic vesicle membrane / positive regulation of inflammatory response / activation of cysteine-type endopeptidase activity involved in apoptotic process / マイクロフィラメント / positive regulation of peptidyl-serine phosphorylation / cellular response to oxidative stress / actin binding / 細胞皮質 / 成長円錐 / histone binding / chemical synaptic transmission / postsynapse / neuron apoptotic process 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: 3.39 Å | ||||||
データ登録者 | Gardner, S.M. / Marmorstein, R. | ||||||
資金援助 | 米国, 1件
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引用 | ジャーナル: bioRxiv / 年: 2023 タイトル: Semi-synthetic CoA-α-Synuclein Constructs Trap N-terminal Acetyltransferase NatB for Binding Mechanism Studies. 著者: Buyan Pan / Sarah Gardner / Kollin Schultz / Ryann M Perez / Sunbin Deng / Marie Shimogawa / Kohei Sato / Elizabeth Rhoades / Ronen Marmorstein / E James Petersson 要旨: N-terminal acetylation is a chemical modification carried out by N-terminal acetyltransferases (NATs). A major member of this enzyme family, NatB, acts on much of the human proteome, including α- ...N-terminal acetylation is a chemical modification carried out by N-terminal acetyltransferases (NATs). A major member of this enzyme family, NatB, acts on much of the human proteome, including α-synuclein (αS), a synaptic protein that mediates vesicle trafficking. NatB acetylation of αS modulates its lipid vesicle binding properties and amyloid fibril formation, which underlies its role in the pathogenesis of Parkinson's disease. Although the molecular details of the interaction between human NatB (hNatB) and the N-terminus of αS have been resolved, whether the remainder of the protein plays a role in interacting with the enzyme is unknown. Here we execute the first synthesis, by native chemical ligation, of a bisubstrate inhibitor of NatB consisting of coenzyme A and full-length human αS, additionally incorporating two fluorescent probes for studies of conformational dynamics. We use cryo-electron microscopy (cryo-EM) to characterize the structural features of the hNatB/inhibitor complex and show that, beyond the first few residues, αS remains disordered when in complex with hNatB. We further probe changes in the αS conformation by single molecule Förster resonance energy transfer (smFRET) to reveal that the C-terminus expands when bound to hNatB. Computational models based on the cryo-EM and smFRET data help to explain the conformational changes and their implications for hNatB substrate recognition and specific inhibition of the interaction with αS. Beyond the study of αS and NatB, these experiments illustrate valuable strategies for the study of challenging structural biology targets through a combination of protein semi-synthesis, cryo-EM, smFRET, and computational modeling. | ||||||
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 | 8g0l.cif.gz | 199.7 KB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb8g0l.ent.gz | 161.7 KB | 表示 | PDB形式 |
PDBx/mmJSON形式 | 8g0l.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/g0/8g0l ftp://data.pdbj.org/pub/pdb/validation_reports/g0/8g0l | HTTPS FTP |
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-関連構造データ
関連構造データ | 29657MC 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
#1: タンパク質 | 分子量: 20390.133 Da / 分子数: 1 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: NAA20, NAT5 発現宿主: Spodoptera frugiperda (ツマジロクサヨトウ) 参照: UniProt: P61599, N-terminal methionine Nalpha-acetyltransferase NatB |
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#2: タンパク質 | 分子量: 112444.258 Da / 分子数: 1 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: NAA25, C12orf30, MDM20, NAP1 発現宿主: Spodoptera frugiperda (ツマジロクサヨトウ) 参照: UniProt: Q14CX7 |
#3: タンパク質・ペプチド | 分子量: 641.799 Da / 分子数: 1 / Fragment: UNP residues 1-5 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: SNCA, NACP, PARK1 / 発現宿主: Escherichia coli BL21 (大腸菌) / 参照: UniProt: P37840 |
#4: 化合物 | ChemComp-CMC / |
研究の焦点であるリガンドがあるか | Y |
-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: Ternary complex of NAA20, NAA25 and CoA-alpha-Synuclein タイプ: COMPLEX / Entity ID: #1-#3 / 由来: RECOMBINANT |
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分子量 | 実験値: NO |
由来(天然) | 生物種: Homo sapiens (ヒト) |
由来(組換発現) | 生物種: Spodoptera frugiperda (ツマジロクサヨトウ) 株: sf9 |
緩衝液 | pH: 7.5 |
試料 | 濃度: 4 mg/ml / 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 / 湿度: 100 % |
-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 FIELDBright-field microscopy / 最大 デフォーカス(公称値): 3000 nm / 最小 デフォーカス(公称値): 1000 nm |
撮影 | 電子線照射量: 43.9 e/Å2 / 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: 5470 |
-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: 3.39 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 192518 / 対称性のタイプ: POINT | ||||||||||||||||||||||||
拘束条件 |
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