Human Apurinic/Apyrimidinic Endonuclease 1 (APE 1)
Source:
An E. coli strain which carries the cloned human APE 1 gene.
Human apurinic/apyrimidic (AP) endonuclease (APE 1) cleaves the phosphodiester backbone a DNA strand immediately 5' to an AP site via a hydrolytic mechanism. Cleavage generates a single-strand DNA break with a 3'-hydroxyl and 5'-deoxyribose phosphate terminus. In addition to endonuclease activity, APE 1 has been reported to have weak DNA 3'-diesterase, 3' to 5' exonuclease, and RNase H activities. APE 1 shares homology with Escherichia coli exonuclease III protein, and is also referred to as HAP1 or Ref-1.
Storage Buffer:
10mM Tris-HCl (pH 7.5), 50mM KCl, 1mM DTT, 0.1mM EDTA, 0.05% Tween®-20, 50% Glycerol
Concentration:
10 units/μl
Unit Definition:
One unit is defined as the amount of enzyme required to cleave 20 pmol of an oligonucleotide duplex containing a single AP site in a total reaction volume of 10 μl in 1 hour at 37°C. An AP site is created by treating 20 pmol of an oligonucleotide duplex containing a single uracil residue with 1 unit of E. coli Uracil-DNA Glycosylase (UDG, PN 71960) for 2 min at 37°C.
10X APE 1 Reaction Buffer (1 ml included):
200mM Tris-acetate (pH 8.0), 500mM potassium acetate, 100mM magnesium acetate, 10mM DTT.
References:
1. Demple, B. et al. (1991) Proc. Natl. Acad. Sci. USA 88, 11450-11454.
2. Barzilay, G. et al. (1995) Nucl. Acids Res., 23,1544-1550.
3. Barzilay, G. et al. (1995) Nature Struc. Biol. 2,451-468.
4. Robson, C. N. and Hickson, D. I. (1991) Nucl. Acids Res. 19, 5519-5523.
5. Flaherty, D. M. (2001) Am. J. Respir. Cell. Mol.Biol. 25, 664-667.
Applications:
• Single cell gel electrophoresis (comet assay)
• Alkaline elution
• Alkaline unwinding
• Modified nick translation
Storage:
Shipped on dry ice. Store at -20°C.