Literature summary for 4.2.99.18 extracted from

Lipton, A.S.; Heck, R.W.; Primak, S.; McNeill, D.R.; Wilson, D.M.; Ellis, P.D.
Characterization of Mg2+ binding to the DNA repair protein apurinic/apyrimidic endonuclease 1 via solid-state 25Mg NMR spectroscopy (2008), J. Am. Chem. Soc., 130, 9332-9341.

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Cloned(Commentary)

Cloned (Comment)	Organism
Recombinant human APE1 protein is overexpressed from clone pETApe1 in Escherichia coli BL21(DE3)	Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
E96Q	the mutant is expressed well in both TB and autoinducing media, the E96Q mutation prevents Mg2+ binding at this site	Homo sapiens
E96Q/D210N	mutation (APE1 ED or simply ED) is created by site-specific mutagenesis of the pETApe1 plasmid, mutant cannot bind Mg2+ in the active site	Homo sapiens

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
nucleus	-	Homo sapiens	5634	-

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	binds to APE1 and a functional APE1-substrate DNA complex with an overall stoichiometry of one Mg2+ per mole of APE1, the chemistry central to the function of APE1 is water activated by a Mg2+ ion, Mg2+ binding is an absolute requirement for the endonucleolytic activity	Homo sapiens

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
35420	-	calculated by the ExPASy Prot Param tool and confirmed by SDS-PAGE	Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
additional information	Homo sapiens	APE1 recognizes AP sites in DNA that arise either spontaneously or as enzymatic products of DNA repair glycosylases that excise substrate base lesions as part of the base excision repair (BER) response. Subsequent to damage recognition, the chemistry central to the function of APE1 is wateractivated by a Mg2+ ion followed by hydrolytic cleavage of the phosphodiester bond immediately 5' to the abasic site	?	-	?

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	P27695	-	-

Purification (Commentary)

Purification (Comment)	Organism
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	Homo sapiens

Purification (Comment)

Organism

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

Homo sapiens

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
additional information	APE1 recognizes AP sites in DNA that arise either spontaneously or as enzymatic products of DNA repair glycosylases that excise substrate base lesions as part of the base excision repair (BER) response. Subsequent to damage recognition, the chemistry central to the function of APE1 is wateractivated by a Mg2+ ion followed by hydrolytic cleavage of the phosphodiester bond immediately 5' to the abasic site	Homo sapiens	?	-	?

Synonyms

Synonyms	Comment	Organism
APE1	-	Homo sapiens
apurinic/apyrimidic endonuclease 1	member of the divalent cation-dependent phosphoesterase superfamily of proteins	Homo sapiens

pH Stability

pH Stability	pH Stability Maximum	Comment	Organism
additional information	-	lowering the pH to 4.5 simply protonated H309 and makes it unsuitable for metal binding	Homo sapiens