EMDB/PDB/SASBDBから引用されている文献のデータベース

キーワード
構造解析手法	All X線回折 NMR 電子顕微鏡小角散乱中性子線回折線維回折粉末回折赤外分光 EPR FRET 理論モデルハイブリッド
著者・登録者
ジャーナル
IF	>30 >20 >10 >5 all

タイトル	Unraveling the mechanisms of PAMless DNA interrogation by SpRY-Cas9.
ジャーナル・号・ページ	Nat Commun, Vol. 15, Issue 1, Page 3663, Year 2024
掲載日	2024年4月30日
著者	Grace N Hibshman / Jack P K Bravo / Matthew M Hooper / Tyler L Dangerfield / Hongshan Zhang / Ilya J Finkelstein / Kenneth A Johnson / David W Taylor /
PubMed 要旨	CRISPR-Cas9 is a powerful tool for genome editing, but the strict requirement for an NGG protospacer-adjacent motif (PAM) sequence immediately next to the DNA target limits the number of editable ...CRISPR-Cas9 is a powerful tool for genome editing, but the strict requirement for an NGG protospacer-adjacent motif (PAM) sequence immediately next to the DNA target limits the number of editable genes. Recently developed Cas9 variants have been engineered with relaxed PAM requirements, including SpG-Cas9 (SpG) and the nearly PAM-less SpRY-Cas9 (SpRY). However, the molecular mechanisms of how SpRY recognizes all potential PAM sequences remains unclear. Here, we combine structural and biochemical approaches to determine how SpRY interrogates DNA and recognizes target sites. Divergent PAM sequences can be accommodated through conformational flexibility within the PAM-interacting region, which facilitates tight binding to off-target DNA sequences. Nuclease activation occurs ~1000-fold slower than for Streptococcus pyogenes Cas9, enabling us to directly visualize multiple on-pathway intermediate states. Experiments with SpG position it as an intermediate enzyme between Cas9 and SpRY. Our findings shed light on the molecular mechanisms of PAMless genome editing.
リンク	Nat Commun / PubMed:38688943 / PubMed Central
手法	EM (単粒子)
解像度	2.79 - 3.69 Å
構造データ	40681 8spq EMDB-40681, PDB-8spq: SpRY-Cas9:gRNA complex targeting TGG PAM DNA 手法: EM (単粒子) / 解像度: 3.26 Å 40705 8sqh EMDB-40705, PDB-8sqh: SpRY-Cas9:gRNA complex targeting TTC PAM DNA 手法: EM (単粒子) / 解像度: 3.69 Å 40740 8srs EMDB-40740, PDB-8srs: SpRY-Cas9:gRNA complex targeting TAC PAM DNA 手法: EM (単粒子) / 解像度: 2.79 Å 41073 8t6o EMDB-41073, PDB-8t6o: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 0 bp R-loop 手法: EM (単粒子) / 解像度: 3.1 Å 41074 8t6p EMDB-41074, PDB-8t6p: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 2 bp R-loop 手法: EM (単粒子) / 解像度: 3.15 Å 41079 8t6s EMDB-41079, PDB-8t6s: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 10 bp R-loop 手法: EM (単粒子) / 解像度: 3.28 Å 41080 8t6t EMDB-41080, PDB-8t6t: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 13 bp R-loop 手法: EM (単粒子) / 解像度: 3.04 Å 41083 8t6x EMDB-41083, PDB-8t6x: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 18 bp R-loop 手法: EM (単粒子) / 解像度: 3.08 Å 41084 8t6y EMDB-41084, PDB-8t6y: SpRY-Cas9:gRNA complex bound to non-target DNA with 1 bp R-loop 手法: EM (単粒子) / 解像度: 2.88 Å 41085 8t76 EMDB-41085, PDB-8t76: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 3 bp R-loop 手法: EM (単粒子) / 解像度: 3.42 Å 41086 8t77 EMDB-41086, PDB-8t77: SpRY-Cas9:gRNA complex bound to non-target DNA with 6 bp R-loop 手法: EM (単粒子) / 解像度: 3.25 Å 41087 8t78 EMDB-41087, PDB-8t78: SpRY-Cas9:gRNA complex bound to non-target DNA with 8 bp R-loop 手法: EM (単粒子) / 解像度: 3.02 Å 41088 8t79 EMDB-41088, PDB-8t79: SpRY-Cas9:gRNA complex bound to non-target DNA with 10 bp R-loop 手法: EM (単粒子) / 解像度: 3.04 Å 41093 8t7s EMDB-41093, PDB-8t7s: SpRYmer bound to NAC PAM DNA 手法: EM (単粒子) / 解像度: 3.01 Å 41775 8tzz EMDB-41775, PDB-8tzz: SpG Cas9 with NGC PAM DNA target 手法: EM (単粒子) / 解像度: 3.56 Å 41867 8u3y EMDB-41867, PDB-8u3y: SpG Cas9 with NGG PAM DNA target 手法: EM (単粒子) / 解像度: 3.1 Å
化合物	ChemComp-MG: Unknown entry / マグネシウムジカチオン ChemComp-HOH: WATER
由来	streptococcus pyogenes (化膿レンサ球菌) streptococcus pyogenes serotype m1 (化膿レンサ球菌) escherichia phage lambda (λファージ) synthetic construct (人工物)
キーワード	IMMUNE SYSTEM / SpRY-Cas9 / CRISPR / Cas9 / IMMUNE SYSTEM/HYDROLASE / IMMUNE SYSTEM-HYDROLASE complex / PAM / R-loop / dimer / IMMUNE SYSTEM/RNA/DNA / SpG-Cas9 / IMMUNE SYSTEM-RNA-DNA complex / IMMUNE SYSTEM/DNA/RNA / IMMUNE SYSTEM-DNA-RNA complex