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Endonuclease V, T.maritima (Endo V)

LO certified Recombinant enzyme Thermal inactivation at 95°C in 10 min in the presence of EDTA

Endonuclease V, T. maritima (Endo V) is a 3'-endonuclease involved in DNA repair, which initiates removal of deaminated bases from damaged DNA, including uracil, hypoxanthine, and xanthine.
  
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Thermo Scientific Endonuclease V, T. maritima (Endo V) is a 3'-endonuclease involved in DNA repair, which initiates removal of deaminated bases from damaged DNA, including uracil, hypoxanthine, and xanthine (see Figure 1).

Endonuclease V is also active toward abasic sites and urea sites, base pair mismatches, flap and pseudo Y structures, and small insertions/deletions in DNA molecules. The cleavage site generated by Endonuclease V is at the second phosphodiester bond 3' to a lesion. 

Highlights

  • Optimal activity at temperatures of 65 to 70°C

Applications

  • High-throughput methods for mutation research (see​ References 3,4)
  • Studies in mutagenesis and DNA repair
  • Mismatch cleavage
  • Genotyping

Note

Use of this enzyme in certain applications may be covered by patents and may require a license. 

When the enzyme is in excess, the primary nicked products experience a second nicking event on the complementary strand, leading to a double-stranded break. At low concentrations, however, Endonuclease V first nicks a DNA strand at the lesions located closer to the 5'-end of DNA molecule. Single-stranded DNA is cleaved with much lower efficiency. Mg2+ or Mn2+ ions are required for enzyme activity (see References 1-3).

  
Storage Condition-20 C
HazardousNo
10X Reaction Buffer500 mM Tris-acetate (pH 7.5), 500 mM KCl, 10 mM EDTA, 0.5% (v/v) Triton X-100.
Concentration5 U/µL
Definition of Activity Unit
  • One unit of the enzyme converts 1 µg of supercoiled depurinized plasmid DNA into other topological states in 30 min at 65°C.
  • Enzyme activity is assayed in the following mixture: 25 mM HEPES-NaOH (pH 7.4), 5 mM MgCl2, 5 mM DTT, 2% (v/v) glycerol, 2 µg of partially depurinated pUC19 DNA.
InactivationInactivated by heating in boiling water bath for 10 min, preferably in the presence of EDTA.
InhibitionInhibitors: no specific inhibitors have been described.
Molecular Weight25 kDa monomer.
Quality ControlThe absence of other endo-, exodeoxyribonucleases, and ribonucleases confirmed by appropriate quality tests.
SourceE.coli with a cloned nfi gene of Thermotoga maritima 
Storage BufferThe enzyme is supplied in:

20 mM HEPES-NaOH (pH 7.4), 5 mM DTT, 50 mM NaCl, 0.1% (v/v) Triton X-100 and 50% (v/v) glycerol.
Endonuclease V activity

Endonuclease V activity

Endonuclease V activity


References

  1. J. Huang et al., Multiple cleavage activities of endonuclease V from Thermotoga maritima: recognition and strand nicking mechanism. Biochemistry. 40, 8738-8748 (2001).
  2. T. M. Hitchcock et al., Cleavage of deoxyoxanosine-containing oligodeoxyribonucleotides by bacterial endonuclease V. Nucleic Acids Res. 32, 4071-4080 (2004).
  3. H. Pincas et al., High sensitivity EndoV mutation scanning through real-time ligase proofreading. Nucleic Acids Res. 32, 148 (2004).
  4. J. Huang et al., An endonuclease/ligase based mutation scanning method especially suited for analysis of neoplastic tissue. Oncogene. 21, 1909-1921 (2002).

Protocols and Product Manuals

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