Stopped-flow fluorescence studies of DNA base flipping by HhaI methyltransferase

2000 ◽  
Vol 28 (5) ◽  
pp. A468-A468
Author(s):  
S. Serva ◽  
E. Weinhold ◽  
S. Klimašauskas
Keyword(s):  
Stopped Flow ◽  
Base Flipping ◽  
Fluorescence Studies ◽  
Dna Base

scholarly journals Characterization of Fluorescent Base Analogs to Study DNA Base Flipping at the Single Molecule Level

2013 ◽  
Vol 104 (2) ◽  
pp. 346a
Author(s):  
Xochitl A. Sosa-Vazquez ◽  
Matthew Vander-Schuur ◽  
Liza Valencia ◽  
Elvin A. Aleman
Keyword(s):  
Single Molecule ◽  
Base Flipping ◽  
Single Molecule Level ◽  
Dna Base

Stopped-Flow Fluorescence Studies of HMG-Domain Protein Binding to Cisplatin-Modified DNA†

Biochemistry ◽  
2000 ◽  
Vol 39 (29) ◽  
pp. 8426-8438 ◽  
Author(s):  
Elizabeth R. Jamieson ◽  
Stephen J. Lippard
Keyword(s):  
Protein Binding ◽  
Stopped Flow ◽  
Fluorescence Studies ◽  
Modified Dna ◽  
Domain Protein

Polycyclic Aromatic DNA-Base Surrogates: High-Affinity Binding to an Adenine-Specific Base-Flipping DNA Methyltransferase

2003 ◽  
Vol 42 (33) ◽  
pp. 3958-3960 ◽  
Author(s):  
Christine Beuck ◽  
Ishwar Singh ◽  
Anupam Bhattacharya ◽  
Walburga Hecker ◽  
Virinder S. Parmar ◽  
...  
Keyword(s):  
Dna Methyltransferase ◽  
Affinity Binding ◽  
Base Flipping ◽  
High Affinity Binding ◽  
High Affinity ◽  
Dna Base ◽  
Polycyclic Aromatic

Stopped-flow fluorescence studies on binding of snake neurotoxins to acetylcholine receptor

1987 ◽  
Vol 6 (3) ◽  
Author(s):  
Toshiya Endo ◽  
Mamoru Nakanishi ◽  
Shoei Furukawa ◽  
FrancoisJ. Joubert ◽  
Nobuo Tamiya ◽  
...  
Keyword(s):  
Acetylcholine Receptor ◽  
Stopped Flow ◽  
Fluorescence Studies ◽  
Snake Neurotoxins

scholarly journals Investigation of the effect of metal ions on the reactivity of thiol groups in human 5-aminolaevulinate dehydratase

1985 ◽  
Vol 225 (3) ◽  
pp. 573-580 ◽  
Author(s):  
P N B Gibbs ◽  
M G Gore ◽  
P M Jordan
Keyword(s):  
Metal Ions ◽  
Binding Sites ◽  
The Other ◽  
Stopped Flow ◽  
Thiol Groups ◽  
Protein Fluorescence ◽  
Fluorescence Studies ◽  
Nitrobenzoic Acid ◽  
Rate Of Reaction ◽  
Molar Equivalent

The reaction of human 5-aminolaevulinate dehydratase with 5,5′-dithiobis-(2-nitrobenzoic acid) (Nbs2) results in the release of 4 molar equivalents of 5-mercapto-2-nitrobenzoic acid (Nbs) per subunit. Two of the thiol groups reacted very rapidly (groups I and II), and their rate constants were determined by stopped-flow spectrophotometry; the other two thiol groups (groups III and IV) were observed by conventional spectroscopy. Titration of the enzyme with a 1 molar equivalent concentration of Nbs2 resulted in the release of 2 molar equivalents of Nbs and the concomitant formation of an intramolecular disulphide bond between groups I and II. Removal of zinc from the holoenzyme increased the reactivity of groups I and II without significantly affecting the rate of reaction of the other groups. The reactions of the thiol groups in both the holoenzyme and apoenzyme were little affected by the presence of Pb2+ ions at concentrations that strongly inhibit the enzyme, suggesting that Zn2+ and Pb2+ ions may have independent binding sites. Protein fluorescence studies with Pb2+ and Zn2+ have shown that the binding of both metal ions results in perturbation of the protein fluorescence.

Stopped-flow fluorescence studies of the interaction of a mutant form of cytochrome b5 with lipid vesicles

1994 ◽  
Vol 4 (3) ◽  
pp. 227-233 ◽  
Author(s):  
N. Krishnamachary ◽  
Frances A. Stephenson ◽  
Alan W. Steggles ◽  
Peter W. Holloway
Keyword(s):  
Cytochrome B5 ◽  
Lipid Vesicles ◽  
Stopped Flow ◽  
Mutant Form ◽  
Fluorescence Studies

scholarly journals CRISPR-Cas9 bends and twists DNA to read its sequence

2021 ◽  
Author(s):  
Joshua C. Cofsky ◽  
Katarzyna M. Soczek ◽  
Gavin J. Knott ◽  
Eva Nogales ◽  
Jennifer A. Doudna
Keyword(s):  
Genome Editing ◽  
Target Sequence ◽  
Base Flipping ◽  
Base Pairs ◽  
Guide Rna ◽  
Target Capture ◽  
Dna Base ◽  
Protospacer Adjacent Motif ◽  
Target Dna ◽  
Dna Base Pairs

In bacterial defense and genome editing applications, the CRISPR-associated protein Cas9 searches millions of DNA base pairs to locate a 20-nucleotide, guide-RNA-complementary target sequence that abuts a protospacer-adjacent motif (PAM)1. Target capture requires Cas9 to unwind DNA at candidate sequences using an unknown ATP-independent mechanism2,3. Here we show that Cas9 sharply bends and undertwists DNA at each PAM, thereby flipping DNA nucleotides out of the duplex and toward the guide RNA for sequence interrogation. Cryo-electron-microscopy (EM) structures of Cas9:RNA:DNA complexes trapped at different states of the interrogation pathway, together with solution conformational probing, reveal that global protein rearrangement accompanies formation of an unstacked DNA hinge. Bend-induced base flipping explains how Cas9 “reads” snippets of DNA to locate target sites within a vast excess of non-target DNA, a process crucial to both bacterial antiviral immunity and genome editing. This mechanism establishes a physical solution to the problem of complementarity-guided DNA search and shows how interrogation speed and local DNA geometry may influence genome editing efficiency.

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