scholarly journals Structural basis of PAM-dependent target DNA recognition by the Cas9 endonuclease

Nature ◽  
2014 ◽  
Vol 513 (7519) ◽  
pp. 569-573 ◽  
Author(s):  
Carolin Anders ◽  
Ole Niewoehner ◽  
Alessia Duerst ◽  
Martin Jinek
Keyword(s):  
Dna Recognition ◽  
Structural Basis ◽  
Target Dna

scholarly journals Structural basis for DNA recognition by the transcription regulator MetR

2016 ◽  
Vol 72 (6) ◽  
pp. 417-426 ◽  
Author(s):  
Avinash S. Punekar ◽  
Jonathan Porter ◽  
Stephen B. Carr ◽  
Simon E. V. Phillips
Keyword(s):  
Molecular Level ◽  
Dna Recognition ◽  
Data Driven ◽  
Structural Basis ◽  
Methionine Biosynthesis ◽  
Dna Complexes ◽  
Winged Helix ◽  
Target Dna ◽  
Operator Sequence ◽  
Dna Complex

MetR, a LysR-type transcriptional regulator (LTTR), has been extensively studied owing to its role in the control of methionine biosynthesis in proteobacteria. A MetR homodimer binds to a 24-base-pair operator region of themetgenes and specifically recognizes the interrupted palindromic sequence 5′-TGAA-N5-TTCA-3′. Mechanistic details underlying the interaction of MetR with its target DNA at the molecular level remain unknown. In this work, the crystal structure of the DNA-binding domain (DBD) of MetR was determined at 2.16 Å resolution. MetR-DBD adopts a winged-helix–turn–helix (wHTH) motif and shares significant fold similarity with the DBD of the LTTR protein BenM. Furthermore, a data-driven macromolecular-docking strategy was used to model the structure of MetR-DBD bound to DNA, which revealed that a bent conformation of DNA is required for the recognition helix α3 and the wing loop of the wHTH motif to interact with the major and minor grooves, respectively. Comparison of the MetR-DBD–DNA complex with the crystal structures of other LTTR-DBD–DNA complexes revealed residues that may confer operator-sequence binding specificity for MetR. Taken together, the results show that MetR-DBD uses a combination of direct base-specific interactions and indirect shape recognition of the promoter to regulate the transcription ofmetgenes.

scholarly journals Structural basis for the Target DNA recognition and binding by the MYB domain of phosphate starvation response 1

FEBS Journal ◽  
2019 ◽  
Author(s):  
Meiqin Jiang ◽  
Lifang Sun ◽  
Michail N. Isupov ◽  
Jennifer A. Littlechild ◽  
Xiuling Wu ◽  
...  
Keyword(s):  
Dna Recognition ◽  
Phosphate Starvation ◽  
Structural Basis ◽  
Starvation Response ◽  
Phosphate Starvation Response ◽  
Target Dna ◽  
Myb Domain

scholarly journals Structural basis of target DNA recognition by CRISPR-Cas12k for RNA-guided DNA transposition

2021 ◽  
Author(s):  
Renjian Xiao ◽  
Shukun Wang ◽  
Ruijie Han ◽  
Zhuang Li ◽  
Clinton Gabel ◽  
...  
Keyword(s):  
Dna Recognition ◽  
Structural Basis ◽  
Structural Elements ◽  
Dna Transposition ◽  
Guide Rna ◽  
Promising Tool ◽  
Target Dna ◽  
Dna Insertion ◽  
Type V

The type V-K CRISPR-Cas system, featured by Cas12k effector with a naturally inactivated RuvC domain and associated with Tn7-like transposon for RNA-guided DNA transposition, is a promising tool for precise DNA insertion. To reveal the mechanism underlying target DNA recognition, we determined a cryo-EM structure of Cas12k from cyanobacteria Scytonema hofmanni in complex with a single guide RNA (sgRNA) and a double-stranded target DNA. Coupled with mutagenesis and in vitro DNA transposition assay, our results revealed mechanisms for the recognition of the GGTT PAM sequence and the structural elements of Cas12k critical for RNA-guided DNA transposition. These structural and mechanistic insights should aid in the development of type V-K CRISPR-transposon systems as tools for genome editing.

scholarly journals Structural basis of target DNA recognition by CRISPR-Cas12k for RNA-guided DNA transposition

2021 ◽  
Author(s):  
Renjian Xiao ◽  
Shukun Wang ◽  
Ruijie Han ◽  
Zhuang Li ◽  
Clinton Gabel ◽  
...  
Keyword(s):  
Dna Recognition ◽  
Structural Basis ◽  
Dna Transposition ◽  
Target Dna

scholarly journals Structural basis of cooperative DNA recognition by the plasmid conjugation factor, TraM

2011 ◽  
Vol 39 (15) ◽  
pp. 6775-6788 ◽  
Author(s):  
Joyce J. W. Wong ◽  
Jun Lu ◽  
Ross A. Edwards ◽  
Laura S. Frost ◽  
J. N. Mark Glover
Keyword(s):  
Dna Recognition ◽  
Structural Basis ◽  
Plasmid Conjugation

scholarly journals Mechanism of R-loop formation and conformational activation of Cas9

2021 ◽  
Author(s):  
Martin Pacesa ◽  
Martin Jinek
Keyword(s):  
Dna Cleavage ◽  
Structural Basis ◽  
Seed Region ◽  
Loop Formation ◽  
Base Pairs ◽  
Guide Rna ◽  
Target Strand ◽  
Target Dna ◽  
Dna Strand ◽  
Dna Substrate

Cas9 is a CRISPR-associated endonuclease capable of RNA-guided, site-specific DNA cleavage. The programmable activity of Cas9 has been widely utilized for genome editing applications. Despite extensive studies, the precise mechanism of target DNA binding and on-/off-target discrimination remains incompletely understood. Here we report cryo-EM structures of intermediate binding states of Streptococcus pyogenes Cas9 that reveal domain rearrangements induced by R-loop propagation and PAM-distal duplex positioning. At early stages of binding, the Cas9 REC2 and REC3 domains form a positively charged cleft that accommodates the PAM-distal duplex of the DNA substrate. Target hybridisation past the seed region positions the guide-target heteroduplex into the central binding channel and results in a conformational rearrangement of the REC lobe. Extension of the R-loop to 16 base pairs triggers the relocation of the HNH domain towards the target DNA strand in a catalytically incompetent conformation. The structures indicate that incomplete target strand pairing fails to induce the conformational displacements necessary for nuclease domain activation. Our results establish a structural basis for target DNA-dependent activation of Cas9 that advances our understanding of its off-target activity and will facilitate the development of novel Cas9 variants and guide RNA designs with enhanced specificity and activity.

scholarly journals Thermodynamic and Structural Basis for Relaxation of Specificity in Protein–DNA Recognition

2014 ◽  
Vol 426 (1) ◽  
pp. 84-104 ◽  
Author(s):  
Paul J. Sapienza ◽  
Tianyi Niu ◽  
Michael R. Kurpiewski ◽  
Arabela Grigorescu ◽  
Linda Jen-Jacobson
Keyword(s):  
Dna Recognition ◽  
Structural Basis

Structural Basis for DNA Recognition by FOXG1 and the Characterization of Disease-causing FOXG1 Mutations

2020 ◽  
Vol 432 (23) ◽  
pp. 6146-6156
Author(s):  
Shuyan Dai ◽  
Jun Li ◽  
Huajun Zhang ◽  
Xiaojuan Chen ◽  
Ming Guo ◽  
...  
Keyword(s):  
Dna Recognition ◽  
Structural Basis

Structural basis for DNA recognition and processing by UvrB

2006 ◽  
Vol 13 (4) ◽  
pp. 360-364 ◽  
Author(s):  
James J Truglio ◽  
Erkan Karakas ◽  
Benjamin Rhau ◽  
Hong Wang ◽  
Matthew J DellaVecchia ◽  
...  
Keyword(s):  
Dna Recognition ◽  
Structural Basis
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