scholarly journals Promiscuous DNA cleavage by HpyAII endonuclease is modulated by the HNH catalytic residues

2020 ◽  
Vol 40 (9) ◽  
Author(s):  
Sumith Kumar ◽  
Sushant Bangru ◽  
Ritesh Kumar ◽  
Desirazu N. Rao
Keyword(s):  
Enzymatic Activity ◽  
Dna Cleavage ◽  
Metal Ion ◽  
Mutational Analysis ◽  
Restriction Enzymes ◽  
Phase Variable ◽  
Histidine Residue ◽  
Cleavage Activity ◽  
Promiscuous Dna ◽  
H Pylori

Abstract Helicobacter pylori is a carcinogenic bacterium that is responsible for 5.5% of all human gastric cancers. H. pylori codes for an unusually large number of restriction–modification (R–M) systems and several of them are strain-specific and phase-variable. HpyAII is a novel Type IIs phase-variable restriction endonuclease present in 26695 strain of H. pylori. We show that HpyAII prefers two-site substrates over one-site substrates for maximal cleavage activity. HpyAII is less stringent in metal ion requirement and shows higher cleavage activity with Ni2+ over Mg2+. Mutational analysis of the putative residues of the HNH motif of HpyAII confirms that the protein has an active HNH site for the cleavage of DNA. However, mutation of the first Histidine residue of the HNH motif to Alanine does not abolish the enzymatic activity, but instead causes loss of fidelity compared with wildtype HpyAII. Previous studies have shown that mutation of the first Histidine residue of the HNH motif of all other known HNH motif motif-containing enzymes completely abolishes enzymatic activity. We found, in the case of HpyAII, mutation of an active site residue leads to the loss of endonuclease fidelity. The present study provides further insights into the evolution of restriction enzymes.

scholarly journals Synthesis, Characterization, Antimicrobial, DNA Cleavage and Fluorescent Activity of Metal ion(II) Coordinate with 2H-Chromene Azo novel ligand

2020 ◽  
Vol 11 (2) ◽  
pp. 1953-1960
Author(s):  
Huda Muayad Nafea ◽  
Ali Muayad Nafea Al- Kawaz
Keyword(s):  
Dna Cleavage ◽  
Metal Ion ◽  
Condensation Reaction ◽  
Free Ligand ◽  
Cleavage Activity ◽  
Dna Cleavage Activity ◽  
Uv Visible ◽  
Group 2 ◽  
Conductance Measuring ◽  
Cobalt And Nickel

A new ligand 2H-chromene containing azo group 2-(4-nitrophenyl)-N-(4-(phenyldiazenyl)-2H-chromen-4-amine (AH), were synthesized from the condensation reaction (1:2)of 2'-hydroxychalcone and p-aminoazobenzene. Co(II), Cu(II) and Ni(II) complexes of the new ligand have been synthesized and characterized using C.H.N. analysis,1HNMR spectra, FT.IR, UV/Visible, magnetic susceptible, conductance measuring, and fluorescence spectral spectroscopy; 13CNMR spectroscopy of the ligand was also studied. Spectroscopic results revealed the 2H-Chromene Azo (AH) ligand behaves as monodentate chelating via the nitrogen atom of amine groupat position 4 having 1:1 [M:L] ratio; suggested that the cobalt and nickel complexes have the tetrahedralstructure and a distorted tetrahedralgeometry for the copper complex, indicating their non-electrolyte nature. The new ligand shows a fluorescence emissioncomparingwith this fluorescence quenching was noticed in its metal complexes. The antibacterial potency of the free ligand and its chelates with metal ion(II) were screened against E.coli, K.pneumoniae, Staph.aureus andB.Subtilis;The DNA cleavage activity of the free ligand and its 2H-chromene azo metal (II) complexes was performedby the gel electrophoresis process, which giventhat thesecompounds are effectiveupon DNA cleavage .

Synthesis, characterization, antimicrobial, pesticidal and DNA cleavage activity of germanium(IV) derivatives of 3-(2-methyl-2,3-dihydro-benzthiazo-2-yl)-chromen-2-one and N′-[1-2-oxo-2H-chrome-3yl-ethylidene]-hydrazinecarbodithionic acid benzyl ester ligands

2011 ◽  
Vol 34 (5-6) ◽  
pp. 139-146 ◽  
Author(s):  
Latika Dawara ◽  
Nighat Fahmi ◽  
R.V. Singh
Keyword(s):  
Antimicrobial Activity ◽  
Metal Complexes ◽  
Dna Cleavage ◽  
Metal Ion ◽  
Benzyl Ester ◽  
Germanium Atom ◽  
Conventional Heating ◽  
Molar Ratio ◽  
Cleavage Activity ◽  
Dna Cleavage Activity

Abstract The new ligands 3-(2-methyl-2,3-dihydro-benzthiazo-2-yl)-chromen-2one (AcBzH) and N′-[1-2-oxo-2H-chrome-3yl-ethylidene]-hydrazinecarbodithionic acid benzyl ester (AcBDTZH) were prepared by the reaction of 3-acetyl-2H-chromen-2-one with 2-aminothiophenol and S-benzyl dithiocarbazate, respectively. The germanium(IV) complexes have been prepared by reacting Ph3GeCl and Me3GeCl in 1:1 molar ratio with these monofunctional bidentate AcBzH and AcBDTZH ligands by using microwave as well as conventional heating methods for comparison purposes. All the synthesized compounds were characterized by elemental analyses, melting point, IR, 1H-NMR 13C-NMR, mass and X-ray powder diffraction techniques. These studies showed that the ligands coordinated to the germanium atom in a monobasic bidentate manner and trigonal bipyramidal environment around the germanium atom have been established for the complexes. To evaluate the effect of metal ion upon chelation, both ligands and their complexes have been screened for their antimicrobial activity against the various pathogenic bacterial and fungal strains. The metal complexes have shown antimicrobial activity as compared to the free ligands. The pesticidal activity and DNA cleavage activity of both ligands and their metal complexes have been tested and discussed.

scholarly journals SYNTHESIS, CHARACTERISATION, THERMAL ANALYSIS AND DNA CLEAVAGE ACTIVITY OF A NOVELZINC (II) COMPLEX OF PYRAZOLE SCHIFF BASES

2018 ◽  
Vol 5 (2) ◽  
pp. 19-23
Author(s):  
Jayanthi Eswaran
Keyword(s):  
Schiff Base ◽  
Schiff Base Ligand ◽  
Dna Cleavage ◽  
Metal Ion ◽  
Acid Hydrazide ◽  
Single Step ◽  
Cleavage Activity ◽  
Dna Cleavage Activity ◽  
Shiff Base ◽  
Ft Ir

A hydrazone Schiff base Zn(II) metal complex is synthesised from the Shiff base ligand Thiophene-2-carboxylic acid hydrazide and 1, 3-diphenyl-1H-pyrazole-4-carboxaldehyde reacted together in 1:1 mole ratio to obtain Schiff base ligand (HL) which was subsequently, allowed to react with Zn(CH3COO)2.2H2O. The Schiff base ligand and its Zn (II) complex prepared were characterized on the basis of elemental nalysis,thermogravimetry, UV-Visible spectroscopy, FT-IR spectroscopy and NMR spectroscopy. IR spectrum of the zinc complex shows that the ligand (HL) is coordinated to the metal ion in monoanionicbidentate fashion with the 1:2 metal to ligand stoichiometry. The thermal behaviour of thecomplex shows a single step decomposition pattern leaving the respective ZnO residue. The DNA cleavage activity of the complex ismonitored using agarose gel lectrophoresis method which indicates the potential of the complex to cleave supercoiled DNA.

scholarly journals Syntheses, Characterization, Antimicrobial and DNA Cleavage Activity of Cr(III) Complexes with Coumarin based Schiff Base Ligands

2021 ◽  
Vol 33 (12) ◽  
pp. 2983-2988
Author(s):  
Ramhari Meena ◽  
Anita Kumari ◽  
Naveen Sharma ◽  
Nighat Fahmi
Keyword(s):  
Schiff Base ◽  
Dna Cleavage ◽  
Metal Ion ◽  
Biological Activities ◽  
Spectroscopic Techniques ◽  
Molecular Weight Determination ◽  
Central Metal ◽  
Schiff Base Ligands ◽  
Cleavage Activity ◽  
Dna Cleavage Activity

A series of biologically important complexes of chromium(III) have been synthesized by the reaction of 3-formyl-4-chlorocoumarin hydrazinecarbothioamide (L1H) and 3-formyl-4-chlorocoumarin hydrazinecarboxamide (L2H) with CrCl3·6H2O in 1:1 and 1:2 molar ratio. All the complexes have been characterized by elemental analysis, molecular weight determination, melting point, conductivity measurements, electronic, IR, 1H NMR, 13C NMR and EPR spectroscopic techniques and X-ray diffraction. In vitro biological screening effects of the compounds were tested against the pathogenic bacterial and fungal species. Further, free ligands and their metal complexes have been screened for their DNA cleavage activity. A comparative study of the biological activities of the Schiff base ligands and their Cr(III) complexes indicates that the complexes exhibit higher antimicrobial and DNA cleavage activity than the free ligands. Physico-chemical studies and spectral data suggested a hexa-coordinated environment around the central metal ion.

scholarly journals Indirect Readout of DNA Controls Filamentation and Activation of a Sequence-Specific Endonuclease

2019 ◽  
Author(s):  
Smarajit Polley ◽  
Dmitry Lyumkis ◽  
N. C. Horton
Keyword(s):  
Conformational Changes ◽  
Dna Cleavage ◽  
Metal Ion ◽  
Structural Changes ◽  
Enzyme Mechanism ◽  
Sequence Specificity ◽  
Dna Sequence Specificity ◽  
Cleavage Activity ◽  
Dna Cleavage Activity ◽  
Indirect Readout

ABSTRACTFilament or run-on oligomer formation by enzymes is increasingly recognized as an important phenomenon with potentially unique regulatory properties and biological roles. SgrAI is an allosterically regulated type II restriction endonuclease that forms run-on oligomeric (ROO) filaments with enhanced DNA cleavage activity and altered sequence specificity. Here, we present the 3.5 Å cryo-electron microscopy structure of the ROO filament of SgrAI bound to a mimic of cleaved primary site DNA and Mg2+. Large conformational changes stabilize a second metal ion cofactor binding site within the catalytic pocket and facilitate assembling a higher-order enzyme form that is competent for rapid DNA cleavage. The structural changes illuminate the mechanistic origin of hyper-accelerated DNA cleavage activity within the filamentous SgrAI form. An analysis of the protein-DNA interface and the stacking of individual nucleotides reveals how indirect DNA readout within filamentous SgrAI enables recognition of substantially more nucleotide sequences than its low-activity form, thereby expanding DNA sequence specificity. Together, substrate DNA binding, indirect readout, and filamentation simultaneously enhance SgrAI’s catalytic activity and modulate substrate preference. This unusual enzyme mechanism may have evolved to perform the specialized functions of bacterial innate immunity in rapid defense against invading phage DNA without causing damage to the host DNA.

scholarly journals Characterization of LlaKI, a New Metal Ion-Independent Restriction Endonuclease from Lactococcus lactis KLDS4

2012 ◽  
Vol 2012 ◽  
pp. 1-6 ◽  
Author(s):  
Abdelkarim Belkebir ◽  
Houssine Azeddoug
Keyword(s):  
Restriction Endonuclease ◽  
Lactococcus Lactis ◽  
Dna Cleavage ◽  
Metal Ion ◽  
Divalent Cations ◽  
Restriction Enzymes ◽  
Restriction Endonucleases ◽  
Size Exclusion ◽  
Type Ii ◽  
Divalent Metal Ions

Requirement of divalent cations for DNA cleavage is a general feature of type II restriction enzymes with the exception of few members of this group. A new type II restriction endonuclease has been partially purified from Lactococcus lactis KLDS4. The enzyme was denoted as LlaKI and showed to recognize and cleave the same site as FokI. The enzyme displayed a denatured molecular weight of 50 kDa and behaved as a dimer in solution as evidenced by the size exclusion chromatography. To investigate the role of divalent cations in DNA cleavage by LlaKI, digestion reactions were carried out at different Mg2+, Mn2+, and Ca2+ concentrations. Unlike most of type II restriction endonucleases, LlaKI did not require divalent metal ions to cleave DNA and is one of the few metal-independent restriction endonucleases found in bacteria. The enzyme showed near-maximal levels of activity in 10 mM Tris-HCl pH 7.9, 50 mM NaCl, 10 mM MgCl2, and 1 mM dithiothreitol at 30°C. The presence of DNA modification was also determined and was correlated with the correspondent restriction enzyme.

DNA-cleavage activity of the iron(II) complex with optically active ligands, meta- and para-xylyl-linked N’,N’-dipyridylmethyl-cyclohexane-1,2-diamine

2021 ◽  
Vol 36 ◽  
pp. 127834
Author(s):  
Koichi Kato ◽  
Yoshimi Ichimaru ◽  
Yoshinori Okuno ◽  
Yoshihiro Yamaguchi ◽  
Wanchun Jin ◽  
...  
Keyword(s):  
Dna Cleavage ◽  
Optically Active ◽  
Cleavage Activity ◽  
Dna Cleavage Activity

scholarly journals A YoeB toxin cleaves both RNA and DNA

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Julia McGillick ◽  
Jessica R. Ames ◽  
Tamiko Murphy ◽  
Christina R. Bourne
Keyword(s):  
Dna Cleavage ◽  
Divalent Cations ◽  
Nuclease Activity ◽  
Dose Dependence ◽  
Cleavage Activity ◽  
Double Stranded Dna ◽  
Dna Cleavage Activity ◽  
Evolutionarily Conserved ◽  
Toxin Protein ◽  
Rna And Dna

AbstractType II toxin-antitoxin systems contain a toxin protein, which mediates diverse interactions within the bacterial cell when it is not bound by its cognate antitoxin protein. These toxins provide a rich source of evolutionarily-conserved tertiary folds that mediate diverse catalytic reactions. These properties make toxins of interest in biotechnology applications, and studies of the catalytic mechanisms continue to provide surprises. In the current work, our studies on a YoeB family toxin from Agrobacterium tumefaciens have revealed a conserved ribosome-independent non-specific nuclease activity. We have quantified the RNA and DNA cleavage activity, revealing they have essentially equivalent dose-dependence while differing in requirements for divalent cations and pH sensitivity. The DNA cleavage activity is as a nickase for any topology of double-stranded DNA, as well as cleaving single-stranded DNA. AtYoeB is able to bind to double-stranded DNA with mid-micromolar affinity. Comparison of the ribosome-dependent and -independent reactions demonstrates an approximate tenfold efficiency imparted by the ribosome. This demonstrates YoeB toxins can act as non-specific nucleases, cleaving both RNA and DNA, in the absence of being bound within the ribosome.

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