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4.2.99.18: DNA-(apurinic or apyrimidinic site) lyase

This is an abbreviated version!
For detailed information about DNA-(apurinic or apyrimidinic site) lyase, go to the full flat file.

Word Map on EC 4.2.99.18

Reaction

the C-O-P bond 3' to the apurinic or apyrimidinic site in DNA is broken by a beta-elimination reaction, leaving a 3'-terminal unsaturated sugar and a product with a terminal 5'-phosphate =

Synonyms

8-oxoG DNA glycosylase, abasic (AP)-endonuclease, abasic endonuclease, AdAPE1/Ref-1, Af0371, Afogg enzyme, ALKBH1, AP 1, AP DNA endonuclease 1, AP Dnase, AP endo, AP endonuclease, AP endonuclease 1, AP endonuclease Ape1, AP endonuclease Class I, AP endonuclease I, AP endonuclease VI, AP lyase, AP site-DNA 5'-phosphomonoester-lyase, AP-endonuclease, AP-endonuclease 1, AP-endonuclease 2, AP-endonuclease/3'-5'exodeoxyribonuclease, AP-lyase, Ap1, Ap2, Ape, APE-1, APE/Ref-1, APE1, APE1/Ref-1, Ape1L, Ape2, APEN, APEX nuclease, APEX1, APLM, APN, APN-1, APN1, apurinic DNA endonuclease, apurinic endodeoxyribonuclease, apurinic endonuclease, apurinic endonuclease 1, apurinic-apyrimidinic DNA endonuclease, apurinic-apyrimidinic endodeoxyribonuclease, apurinic-apyrimidinic endonuclease, apurinic-apyrimidinic endonuclease 1, apurinic-apyrimidinic endonuclease I, apurinic/apyrimidic endonuclease 1, apurinic/Apyrimidinic (AP) endonuclease 1, apurinic/apyrimidinic AP endonuclease, apurinic/apyrimidinic DNA endonuclease 1, apurinic/apyrimidinic endonuclease, apurinic/apyrimidinic endonuclease 1, apurinic/apyrimidinic endonuclease 1/redox factor-1, apurinic/apyrimidinic endonuclease 2, apurinic/apyrimidinic endonuclease APE1, apurinic/apyrimidinic endonuclease-1, apurinic/apyrimidinic endonuclease-1/Redox factor-1, apurinic/apyrimidinic endonuclease/redox effector factor-1, Apurinic/apyrimidinic endonuclease1/redox factor 1, apurinic/apyrimidinic endonuclease1/redox factor-1, apurinic/apyrimidinic lyase, apurinic/apyrimidinic site lyase, apurinic/apyrimidinic specific endonuclease, apurinic/apyrimidinic-endonuclease, apyrimidinic endonuclease, archaeal GO glycosylase, BAP1, class II AP endonuclease LMAP, class II apurinic/apyrimidinic(AP)-endonuclease, class-II AP-endonuclease, deoxyribonuclease (apurinic or apyrimidinic), DNA lyase, DNA polymerase X, DNA-(apurinic or apyrimidinic site) lyase, E. coli endonuclease III, EC 3.1.25.2, EcoNth, Endo III, endodeoxyribonuclease, endodeoxyribonuclease III, EndoIV, endonuclease III, endonuclease IV, endonuclease VI, endonuclease VIII, Escherichia coli endonuclease III, EXO-3, ExoA, ExoA type AP endonuclease, ExoIII, HAP-1, HAP1, HAP1h, hAPE, hAPE1, hNTH, hNTH1, hOgg1 protein, HpXth, human apurinic/apyrimidinic endonuclease, human apurinic/apyrimidinic endonuclease 1, Kae1, KsgA, LplExo, Micrococcus luteus UV endonuclease, MJ0724, mjOgg, MMH, More, mtAPE, MtbXthA, multi-functional apyrimidinic endonuclease1/redox factor-1, N-glycosylase AP lyase, N-glycosylase apurinic/apyrimidinic lyase, NapE, Nei, Nfo, Ntg1, Ntg1p, Ntg2, Ntg2p, Nth, NTH1, nuclease SmnA, nuclease, apurinic endodeoxyribo-, nuclease, apurinic-apyrimidinic endodeoxyribo-, nuclease, endodeoxyribo-, III, Ogg2, oxoG DNA glycosylase, Pa-AGOG, Pa-AGOG DNA glycosylase, PAE2237, PaKae1, PALF, pE296R, PF0258, phage-T4 UV endonuclease, redox effector factor 1, redox factor-1, redox factor-1:Ref-1, Ref-1, REF-1 protein, Ref1, SisExoIII, Smx nuclease, TvoExo, UL30, UV endo V, UV endonuclease, UV endonuclease V, X-ray endonuclease III, XTH, xthA, ZAP1, zApe

ECTree

     4 Lyases
         4.2 Carbon-oxygen lyases
             4.2.99 Other carbon-oxygen lyases
                4.2.99.18 DNA-(apurinic or apyrimidinic site) lyase

Purification

Purification on EC 4.2.99.18 - DNA-(apurinic or apyrimidinic site) lyase

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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
0.3M isozyme
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1100fold
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117fold
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5 distinct apurinic endonucleases: D1, D2, D3, D4 and E
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according to protocols for purification of GFP-, GST- and His-tagged proteins
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after affinity purification and cleavage of the His-tag, the untagged proteins to near homogeneity are purified by fast protein liquid chromatography using heparin-sepharose
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AP endonuclease I and II, 400fold
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APE-1 proteins partially purified purified are visualized by Western blot using anti-human APE-1 antibody
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Bacteria are lyzed in a denaturing buffer (50 mmol NaCl and phosphate pH 7.5, 4 mol urea), the His-hNTH1 protein is purified by chromatography on a nickel column followed by gel filtration on a Superdex-200 column. The TAP-hNTH1 protein is purified from a stable MCF-7 clone expressing this protein construct with a TAP purification kit.
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by ammonium sulfate precipitation and affinity chromatography, using a HiTrap chelating column, charged with Ni2+, and a HiTrap heparin column
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by nickel-nitrilotriacetic acid affinity chromatography
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Clear supernatant from cell extract is applied to two columns in series: a strong anion exchange column (Q-Sepharose) followed by a strong cation exchange (HS50). After sample loading, both columns are washed with buffer and then the Q-Sepharose column is disconnected. APE1 bound in the HS50 column is eluted with a 50-700 mM KCl gradient. Clean APE1 fractions are pooled and then dialyzed extensively in buffer to remove glycerol, elution salts, and metal ions, followed by two runs of dialysis with EDTA-free buffer. Average yield of APE1 is 200 mg out of 15 g of cells
dialyzed mitochondrial lysates are loaded on a HiTrap Q and SP-Sepharose columns, fractions with APE activity are loaded onto a HiTrap SP column and are further purified using a HiTrap-Heparin column
First step is a Ni-NTA affinity chromatography. Then the Ni-NTA fractions containing zApe are combined and loaded on a pre-equilibrated MonoQ column after dilution to a final NaCl concentration of 50 mM. The protein is eluted using a linear NaCl gradient, and fractions containing the zApe are combined, digested with thrombin, and then subjected again to a final MonoQ column. The purified proteins are concentrated to 50-75 mg/ml. The mutant T58C is purified by using an Q-sepharose column as ion exchanger.
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full-length recombinant CSB and APE1 proteins are expressed and purified, used in direct and indirect ELISAs
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GST-EXO-3 fusion protein and GST-E68A fusion protein
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heparin-Sepharose column chromatography and Superdex 75 gel filtration
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His-tagged APE1 is purified from Escherichia coli M15 cells by using Ni-affinity chromatography with Swell-gel Nickel-chelated discs
histidine-tagged AP endonucleases are purified on Ni2+-charged HiTrap Chelating HP columns
HisTrap column chromatography and Hitrap Q column chromatography
HiTrap column chromatography, HiTrap heparin column chromatography, HiTrap Blue column chromatography, glutathione-Sepharose 4B column chromatography, hydroxyapatite column chromatography, and Resource S column chromatography
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homogeneity
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infected with bacteriophage T4
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near homogeneity
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Ni-NTA column chromatography and HisTrap chelating column chromatography
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Ni-NTA column chromatography and Superdex 75 gel filtration
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Ni-NTA column chromatography, S-Sepharose column chromatography
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Ni-NTA column chromatography, SP-Sepharose column chromatography
nickel-Sepharose resin column chromatography, gel filtration
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Ntg1p and Ntg2p
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nuclear and mitochondrial extracts are isolated
overproduced in Escherichia coli
partial
partial, endonuclease A
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Purification is done using the standard glutathione affinity purification protocol. The C65A mutant is purified as follows: cell suspension is centrifuged, supernatant is loaded on a Ni-NTA column and the protein is eluted with a linear imidazole gradient. Fractions containing C65A hApe1 are further purified on an S-Sepharose column with a linear NaCl gradient.
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purification of recombinant enzyme
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recombinant
recombinant enzyme from Escherichia coli strain Rosetta II(DE3)
recombinant enzyme from Escherichia coli strain Rosetta II(DE3) by anion exchange chromatography, dialysis, cation exchange chromatography, dialysis, and hydrophobic interaction chromatography
recombinant His-tagged enzyme from Escherichia coli strain Rosetta 2 (DE3) by nickel affinity chromatography
recombinant His-tagged wild-type and mutant enzymes by nickel affinity chromatography
recombinant human full-length and Ndelta33 APE1 is purified using a Ni-NTA Superflow and a SP-Sepharose column
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recombinant protein
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recombinant wild-type and mutant enzymes
RKO cell protein is extracted
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TAP-tagged Ntg1 is purified by a modified TAP method. To purify GST-tagged Ntg1, glutathione-Sepharose 4 Fast Flow beads are applied
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TAP-tagged Ntg2 is purified by a modified TAP method. To purify GST-tagged Ntg2, glutathione-Sepharose 4 Fast Flow beads are applied
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using a HisTrap HP column
using a metal-affinity HisTrap chelating HP column, preequilibrated with CoCl2, and size-exclusion chromatography, using a HiLoad 16/60 Superdex 200 prep grade column
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using a Ni2+-charged HiTrap Chelating HP column and a gel filtration column
using Ni-nitrilotriacetic acid agarose beads
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wild-type and mutant enzymes H270A, H270R, H270L, Q315A, F319A