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Literature summary for 4.2.99.18 extracted from

  • Dyrkheeva, N.S.; Lebedeva, N.A.; Lavrik, O.I.
    AP endonuclease 1 as a key enzyme in repair of apurinic/apyrimidinic sites (2016), Biochemistry (Moscow), 81, 951-967 .
    View publication on PubMed

Activating Compound

Activating Compound Comment Organism Structure
additional information the 3'-5'-exonuclease activity of APE1 towards the DNA with a 5'-pFx02group is stimulated in the presence of PARP1 Homo sapiens

Crystallization (Commentary)

Crystallization (Comment) Organism
X-ray diffraction analysis of APE1 complexes with oligomeric DNA-duplexes (11 or 15 bp) containing an AP site. There is only one bivalent metal ion in the APE1 active site present in the crystal formed at pH 4.6. In the structure produced at pH 7.5, i.e. under conditions optimal for endonuclease activity, two metal ions are located in the active site of the enzyme Homo sapiens

Protein Variants

Protein Variants Comment Organism
D219A replacement of Asp219 with alanine decreases both the DNA binding and the AP endonuclease activity of the enzyme compared to wild-type Homo sapiens
D308A replacement of Asp308 with alanine results in decrease in endonuclease activity, but the DNA binding activity is preserved compared to wild-type Homo sapiens
D90A replacement of Asp90 with alanine results in decrease in endonuclease activity, but the DNA binding activity is preserved compared to wild-type Homo sapiens
E96A replacement of Glu96 with alanine results in decrease in endonuclease activity, but the DNA binding activity is preserved compared to wild-type Homo sapiens
R177A he turnover number of the endonuclease reaction catalyzed by APE1 increases compared to wild-type when Arg177 is replaced by alanine Homo sapiens

Inhibitors

Inhibitors Comment Organism Structure
Cd2+ inhibits the enzyme to a variable degree in the cell extract Homo sapiens
Fe2+ inhibits the enzyme to a variable degree in the cell extract Homo sapiens
additional information the cleavage of single-stranded DNA with an AP site does not depend on the presence of DNA glycosylases, and it is not inhibited by the reaction product. Replacement of tetrahydrofuran with a positively charged analogue pyrrolidine decreases endonuclease activity of the APE1 21fold and the DNA binding activity by 4fold. Endonuclease activity on DNA with an tetrahydrofuran residue decreases 7300fold in 4 mM EDTA in comparison with the activity in the presence of 10 mM Mg2+, and the activity decreases only 20-30fold on DNA containing ethane and propanediol in the center of the strand instead of an AP site Homo sapiens
nucleotides dAMP across from the AP site does not cause any distortions in the helix in comparison with the undamaged DNA, while the presence dCMP and dGMP results in changes in the helical structure to a varying degree providing out-of-helix position of the nucleotide and/or AP site Homo sapiens
Pb2+ inhibits the enzyme to a variable degree in the cell extract Homo sapiens

Localization

Localization Comment Organism GeneOntology No. Textmining
chromatin APE1 is located in the area of an AP site on the strand containing the AP site relative to the DNA helix, bending the helix in the process. APE1 is fixed on the DNA through two enzyme sites. One of them containing the Met270 and Met271 residues interacts with the minor groove, and another containing Arg177,with the major DNA groove. The Arg177 residue forms a hydrogen bond with the 3'-phosphate of the AP site. APE1 stabilizes the out-of-helix position of the AP site and is efficiently anchored on the DNA. Interaction of Arg177 with DNA slows dissociation of APE1 from the product of the endonuclease reaction Homo sapiens 785
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cytoplasm
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Homo sapiens 5737
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cytoplasm
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Mus musculus 5737
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mitochondrion
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Homo sapiens 5739
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mitochondrion
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Mus musculus 5739
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additional information depending on the type of cells, APE1 is localized primarily in the nucleus, cytoplasm, or mitochondria Homo sapiens
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additional information depending on the type of cells, APE1 is localized primarily in the nucleus, cytoplasm, or mitochondria Mus musculus
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nucleus
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Homo sapiens 5634
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nucleus
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Mus musculus 5634
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Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ activates, Mg2+ or Mn2+ is required for endonuclease and exonuclease activity of APE1. Mg2+ is required for both binding of the protein with DNA and cleavage of phosphodiester bond catalyzed by APE1. Depending on the Mg2+ concentration, the limiting stage of the process can change Homo sapiens
Mn2+ activates, Mg2+ or Mn2+ is required for endonuclease and exonuclease activity of APE1 Homo sapiens
additional information there is only one bivalent metal ion in the APE1 active site present in the crystal formed at pH 4.6. In the structure produced at pH 7.5, i.e. under conditions optimal for endonuclease activity, two metal ions are located in the active site of the enzyme. MgCl2 concentration in the range 0.5-2.0 mM and low (50 mM or below) concentration of KCl are optimal for cleavage of single-stranded DNA with an AP site, while the endonuclease activity towards the double-stranded AP-DNA is the highest at 10 mM MgCl2 and 50 mM KCl or 2 mM MgCl2 and 200 mM KCl. Co2+ and Ni2+ do not affect APE1 activity Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
additional information Homo sapiens the mechanism of catalysis of the APE1 endonuclease reaction includes nucleophilic attack by a hydroxide ion on the phosphorous atom at the 5'-side from AP site. The hydroxide ion is formed from a water molecule activated by the Asp210 residue. The transition complex is stabilized via formation of hydrogen bonds with the Asn174, Asn212, and His309/Asp283 residues. Glu96 participates in binding of one Mg2+, which coordinates the leaving O3'-group, and Asp210 and His309 coordinate a second metal ion participating in formation of a hydroxide ion from a water molecule. A transition state with the phosphorous atom is formed because of the nucleophilic attack, the destabilized P-O3' bond is cleaved, and, as a result, inversion of the phosphate configuration occurs (SN2-mechanism). Slow dissociation of the APE1-DNA complex (product) prevents accumulation of single-strand breaks in DNA. The reaction rate of this step increases with increase in Mg2+ concentration and, as a result, the catalysis itself likely becomes the limiting step. Mechanism of the P-O3' bond cleavage at the 5'-side of an AP site catalyzed by human APE1 derived from the structure of the APE1-DNA complex produced by X-ray at higher resolution, overview ?
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?

Organism

Organism UniProt Comment Textmining
Homo sapiens P27695
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Mus musculus P28352
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Posttranslational Modification

Posttranslational Modification Comment Organism
acetylation APE1 is acetylated at several residues in the N-domain Homo sapiens
acetylation APE1 is acetylated at several residues in the N-domain Mus musculus
additional information APE1 is subjected to different types of posttranslational modifications: acetylation at several residues in the N-domain, nitrosylation on residues of the C-domain, ubiquitination, proteolysis, and sumoylation Homo sapiens
additional information APE1 is subjected to different types of posttranslational modifications: acetylation at several residues in the N-domain, nitrosylation on residues of the C-domain, ubiquitination, proteolysis, and sumoylation Mus musculus
nitrosylation APE1 is nitrosylated on residues of the C-domain Homo sapiens
nitrosylation APE1 is nitrosylated on residues of the C-domain Mus musculus
proteolytic modification APE1 is subjected to proteolysis Homo sapiens
proteolytic modification APE1 is subjected to proteolysis Mus musculus
sumoylation APE1 is subjected to sumoylation Homo sapiens
sumoylation APE1 is subjected to sumoylation Mus musculus
ubiquitination APE1 is subjected to ubiquination Homo sapiens
ubiquitination APE1 is subjected to ubiquination Mus musculus

Source Tissue

Source Tissue Comment Organism Textmining
cell culture
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Homo sapiens
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HeLa cell
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Homo sapiens
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additional information APE1 is synthesized in human cells in large quantities Homo sapiens
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Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
additional information the mechanism of catalysis of the APE1 endonuclease reaction includes nucleophilic attack by a hydroxide ion on the phosphorous atom at the 5'-side from AP site. The hydroxide ion is formed from a water molecule activated by the Asp210 residue. The transition complex is stabilized via formation of hydrogen bonds with the Asn174, Asn212, and His309/Asp283 residues. Glu96 participates in binding of one Mg2+, which coordinates the leaving O3'-group, and Asp210 and His309 coordinate a second metal ion participating in formation of a hydroxide ion from a water molecule. A transition state with the phosphorous atom is formed because of the nucleophilic attack, the destabilized P-O3' bond is cleaved, and, as a result, inversion of the phosphate configuration occurs (SN2-mechanism). Slow dissociation of the APE1-DNA complex (product) prevents accumulation of single-strand breaks in DNA. The reaction rate of this step increases with increase in Mg2+ concentration and, as a result, the catalysis itself likely becomes the limiting step. Mechanism of the P-O3' bond cleavage at the 5'-side of an AP site catalyzed by human APE1 derived from the structure of the APE1-DNA complex produced by X-ray at higher resolution, overview Homo sapiens ?
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?
additional information APE1 cleaved AP sites in the structure of DNA/RNAx02hybrid (DNA strand), singlex02stranded RNA, single-stranded DNA, and double-stranded regions of pseudo-triplex DNA-RNA complexes modeling replication and transcription intermediates. APE1 exhibits endonuclease activity during hydrolysis of an AP site on a single-stranded DNA. The activity on the single-stranded DNA is 20fold lower than on double-stranded DNA. As in the case of double-stranded DNA, endonuclease activity of APE1 depended on the presence of magnesium ions. the cleavage of single-stranded DNA with an AP site does not depend on the presence of DNA glycosylases, and it is not inhibited by the reaction product. Complexes of APE1 with double-stranded DNA containing an tetrahydrofuran residue in the center of a non-cleaved strand are not detected Homo sapiens ?
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?

Subunits

Subunits Comment Organism
More the N-terminal domain (about 6 kDa, 60 amino acids) is responsible for the redox function of APE1, which is independent of the repair functions of this enzyme, and it contains the nuclear export signal Homo sapiens
More the N-terminal domain (about 6 kDa, 60 amino acids) is responsible for the redox function of APE1, which is independent of the repair functions of this enzyme, and it contains the nuclear export signal Mus musculus

Synonyms

Synonyms Comment Organism
AP endonuclease 1
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Homo sapiens
AP endonuclease 1
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Mus musculus
APE1
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Homo sapiens
APE1
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Mus musculus
apurinic/apyrimidinic endonuclease 1
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Homo sapiens
apurinic/apyrimidinic endonuclease 1
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Mus musculus

Temperature Range [°C]

Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
30 42 activity range Homo sapiens

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.4
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exonuclease activity of APE1 Homo sapiens
7.5 7.8
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Homo sapiens
7.7 7.9 endonuclease activity of APE1 Homo sapiens

pH Range

pH Minimum pH Maximum Comment Organism
6.7 9 activity range Homo sapiens

Expression

Organism Comment Expression
Homo sapiens level of expression of the gene encoding APE1 increases under oxidative stress up

General Information

General Information Comment Organism
evolution basic human AP endonuclease is a multifunctional protein. AP endonucleases fall into two families depending on the similarity of their amino acid sequence with exonuclease III (ExoIII or Xth) or endonuclease IV (EndoIV or Nfo) from Escherichia coli. APE1 belongs to the large nuclease family related to ExoIII from Escherichia coli. Enzymes of the ExoIII family including APE1 exhibit several enzymatic activities manifested more or less effectively: AP endonuclease, 3'-phosphodiesterase, 3'-phosphatase, and 3'-5'-exonuclease Homo sapiens
malfunction inactivation of the APE1 gene causes early embryonic lethality in mice Mus musculus
metabolism mechanisms of AP site cleavage by enzymes from the base excision repair system (BER), overview. Enzyme Polbeta in combination with APE1 is able to perform synthesis with strand displacement and simultaneously correction of Polbeta errors with catalysis by 3'-5'-exonuclease activity of the APE1. A Schiff base is formed as an intermediate in the reactions catalyzed by AP lyases and 5'-dRP lyases. One of the repair proteins interacting with the AP sites via formation of a Schiff base is poly(ADP-ribose)polymerase 1 (PARP1). PARP1 is known as a sensor of single-stranded breaks and as a protein regulator of BER. If exogenous APE1 is added prior to irradiation, the efficiency of the PARP1 and FEN1 labeling decreases (but not of the Polbeta), these proteins compete for binding to this DNA substrate. Interaction between various enzymes and proteins participating in BER, overview Homo sapiens
additional information the N-terminal domain (about 6 kDa, 60 amino acids) is responsible for the redox function of APE1, which is independent of the repair functions of this enzyme, and it contains the nuclear export signal. The second human AP endonuclease (APE2) also belongs to the ExoIII family, but the level of its endonuclease activity is significantly lower than of APE1. Amino acids such as Asp219, Asp90, and Asp308 are functionally important, D219 in APE1 plays a key function in repair. Tyr171 does not interact directly with the AP site, but participates in the catalysis of the APE1 endonuclease reaction in the form of a phenolate ion, i.e. it attacks phosphate at the 5'-side of the AP site Homo sapiens
physiological function apurinic/apyrimidinic sites (AP sites) are among the most abundant DNA damages. They can emerge because of spontaneous hydrolysis of the N-glycoside bond and during removal of the damaged DNA bases by DNA glycosylases. In mammalian cells up to 10000 AP sites emerge daily primarily due to apurinization of DNA. The number of such damages increases dramatically during intensive oxidative stress, X-ray and UV irradiation, and other actions. The deoxyribose residues in the AP sites exist in different forms that are in equilibrium. The acyclic aldehyde form of an AP site can form a Schiff base with amino groups in proteins, most often with the epsilon-NH2 group of lysine residues. AP endonucleases are the most important enzymes involved in the DNA repair that initiates the repair of AP sites. APE1 exhibits 3'-phosphodiesterase, 3'-5'-exonuclease, and 3'-phosphatase activities. Role of APE1 in the DNA repair process and in other metabolic processes, overview. APE1 stimulates the activity of the mouse adeninex02DNA glycosylase (Myh) and enhances affinity of this enzyme to adenine/8-oxoguanine pairs in comparison with adenine/guanine pairs Mus musculus
physiological function apurinic/apyrimidinic sites (AP sites) are among the most abundant DNA damages. They can emerge because of spontaneous hydrolysis of the N-glycoside bond and during removal of the damaged DNA bases by DNA glycosylases. In mammalian cells up to 10000 AP sites emerge daily primarily due to apurinization of DNA. The number of such damages increases dramatically during intensive oxidative stress, X-ray and UV irradiation, and other actions. The deoxyribose residues in the AP sites exist in different forms that are in equilibrium. The acyclic aldehyde form of an AP site can form a Schiff base with amino groups in proteins, most often with the epsilon-NH2 group of lysine residues. AP endonucleases are the most important enzymes involved in the DNA repair that initiates the repair of AP sites. Human apurinic/apyrimidinic endonuclease 1 (APE1) is one of the key participants in the DNA base excision repair system. The major contribution to AP site cleavage in mammalian cells is provided by APE1 (over 95% of damages). APE1 hydrolyzes DNA adjacent to the 5'-end of an AP site to produce a nick with a 3'-hydroxyl group and a 5'-deoxyribose phosphate moiety. APE1 exhibits 3'-phosphodiesterase, 3'-5'-exonuclease, and 3'-phosphatase activities. APE1 is also identified as a redox factor (Ref-1). Role of APE1 in the DNA repair process and in other metabolic processes, overview. The APE1 protein is required for viability of human cell culture. APE1 is required not only for hydrolysis of AP sites generated by monofunctional DNA glycosylases, but also for processing of the products of bifunctional DNA glycosylases catalyzing beta-elimination. The enzyme is obviously involved in the cell response system to the action of some genotoxic agents. Participation of APE1 in stimulation of the DNA glycosylase, lyase, polymerase, flap endonuclease, and DNA ligase activities of BER enzymes likely reflects the role of this enzyme in coordination of different stages of DNA repair to achieve its optimal efficiency Homo sapiens