Microscopic insight to specificity of metal ion cofactor in DNA cleavage by restriction endonuclease EcoRV

Biopolymers ◽  
2020 ◽  
Vol 111 (10) ◽  
Author(s):  
Sasthi Charan Mandal ◽  
Lakshmi Maganti ◽  
Manas Mondal ◽  
Jaydeb Chakrabarti
Keyword(s):  
Restriction Endonuclease ◽  
Dna Cleavage ◽  
Metal Ion

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.

On the Divalent Metal Ion Dependence of DNA Cleavage by Restriction Endonucleases of the EcoRI Family

2009 ◽  
Vol 393 (1) ◽  
pp. 140-160 ◽  
Author(s):  
Vera Pingoud ◽  
Wolfgang Wende ◽  
Peter Friedhoff ◽  
Monika Reuter ◽  
Jürgen Alves ◽  
...  
Keyword(s):  
Dna Cleavage ◽  
Metal Ion ◽  
Divalent Metal ◽  
Restriction Endonucleases ◽  
Divalent Metal Ion

scholarly journals Analysis of the HindIII-catalyzed reaction by time-resolved crystallography

2015 ◽  
Vol 71 (2) ◽  
pp. 256-265 ◽  
Author(s):  
Takashi Kawamura ◽  
Tomoki Kobayashi ◽  
Nobuhisa Watanabe
Keyword(s):  
Electron Density ◽  
Active Site ◽  
Dna Cleavage ◽  
Metal Ion ◽  
Manganese Ions ◽  
Metal Ion Binding ◽  
Time Resolved ◽  
Manganese Ion ◽  
Ion Binding Sites ◽  
Metal Ion Binding Sites

In order to investigate the mechanism of the reaction catalyzed by HindIII, structures of HindIII–DNA complexes with varying durations of soaking time in cryoprotectant buffer containing manganese ions were determined by the freeze-trap method. In the crystal structures of the complexes obtained after soaking for a longer duration, two manganese ions, indicated by relatively higher electron density, are clearly observed at the two metal ion-binding sites in the active site of HindIII. The increase in the electron density of the two metal-ion peaks followed distinct pathways with increasing soaking times, suggesting variation in the binding rate constant for the two metal sites. DNA cleavage is observed when the second manganese ion appears, suggesting that HindIII uses the two-metal-ion mechanism, or alternatively that its reactivity is enhanced by the binding of the second metal ion. In addition, conformational change in a loop near the active site accompanies the catalytic reaction.

scholarly journals Fluorophore ATCUN complexes: combining agent and probe for oxidative DNA cleavage

2015 ◽  
Vol 51 (62) ◽  
pp. 12395-12398 ◽  
Author(s):  
C. Wende ◽  
N. Kulak
Keyword(s):  
Dna Cleavage ◽  
Metal Ion ◽  
Fluorescent Reporter ◽  
Dna Cleaving ◽  
Oxidative Dna Cleavage

A Cu(ii)-based peptidic DNA cleaving agent equipped with a Cu(ii)-sensing fluorescent reporter allows monitoring the fate of the nucleolytic metal ion.

DNA cleavage by the EcoRV restriction endonuclease: roles of divalent metal ions in specificity and catalysis

1999 ◽  
Vol 288 (1) ◽  
pp. 87-103 ◽  
Author(s):  
Geoffrey S. Baldwin ◽  
Richard B. Sessions ◽  
Symon G. Erskine ◽  
Stephen E. Halford
Keyword(s):  
Metal Ions ◽  
Restriction Endonuclease ◽  
Dna Cleavage ◽  
Divalent Metal ◽  
Divalent Metal Ions
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