Literature summary for 4.2.99.18 extracted from

Guay, D.; Garand, C.; Reddy, S.; Schmutte, C.; Lebel, M.
The human endonuclease III enzyme is a relevant target to potentiate cisplatin cytotoxicity in Y-box-binding protein-1 overexpressing tumor cells (2008), Cancer Sci., 99, 762-769.

Search for more literature for 4.2.99.18

Activating Compound

Activating Compound	Comment	Organism
cisplatin	2fold increase in Y-box-binding protein-1/hNTH1 complex formation in transfected MCF-7 cells after a treatment of 12 microM cisplatin for 4 h	Homo sapiens
additional information	no increase in the amount of Y-box-binding protein-1/hNTH1 complex in transfected MCF-7 cells after camptothecin or mitomycin C treatments compared to untreated MCF7 cells	Homo sapiens
UV light	3fold increase in Y-box-binding protein-1/hNTH1 complex formation in transfected MCF-7 cells after a treatment with a UV irradiation dose of 40 J/m2 tested for 4 h	Homo sapiens
Y-box-binding protein-1	protein strongly stimulates in vitro the activity of hNTH1 toward DNA duplex probes containing oxidized bases, lesions prone to be present in cisplatin treated cells	Homo sapiens

Application

Application	Comment	Organism
drug development	cancer therapy, human endonuclease III enzyme is a relevant target to potentiate cisplatin cytotoxicity in Y-box binding protein-1 overexpressing tumor cells	Homo sapiens

Cloned(Commentary)

Cloned (Comment)	Organism
The human NTH1 cDNA is cloned into the PCRII plasmid and it is cloned into the EcoRI site of the pGEX-2TK vector. The hNTH1 cDNA is cut (amino acids 1-94) and cloned into the modified restriction sites in the pGEX-2TK vector. The remaining fragment (amino acids 95-312) is cloned into the pGEX-3X vector. The hNTH1 cDNA is cloned into the pET24d vector in-frame with the His-tag and into the pNTAP-B vector in-frame with the TAP-epitope. To knock down the expression of Y-box-binding protein-1, a hairpin sequence specific to Y-box-binding protein-1 is cloned into the pSuper vector. His-hNTH1 protein is produced in the BL21 bacterial strain. MCF-7 cells are transfected with a siRNA specific to hNTH1.	Homo sapiens

Cloned (Comment)

Organism

The human NTH1 cDNA is cloned into the PCRII plasmid and it is cloned into the EcoRI site of the pGEX-2TK vector. The hNTH1 cDNA is cut (amino acids 1-94) and cloned into the modified restriction sites in the pGEX-2TK vector. The remaining fragment (amino acids 95-312) is cloned into the pGEX-3X vector. The hNTH1 cDNA is cloned into the pET24d vector in-frame with the His-tag and into the pNTAP-B vector in-frame with the TAP-epitope. To knock down the expression of Y-box-binding protein-1, a hairpin sequence specific to Y-box-binding protein-1 is cloned into the pSuper vector. His-hNTH1 protein is produced in the BL21 bacterial strain. MCF-7 cells are transfected with a siRNA specific to hNTH1.

Homo sapiens

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	included in assay medium	Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.
DNA	Homo sapiens	bifunctional enzyme is involved in base excision repair, it is a bifunctional DNA glycosylase/apurinic/apyrimidinic lyase which removes hydrated, reduced, or oxidized bases from the DNA backbone as the initial step of base excision repair	fragments of DNA	-	?
additional information	Homo sapiens	hNTH1 and Y-box-binding protein-1 may be part of the same DNA repair pathway in response to cisplatin and UV treatments	?	-	?
additional information	Homo sapiens	hNTH1 binds directly to Y-box-binding protein-1 in the absence of nucleic acids, it binds to the auto-inhibitory n-terminal tail of NTH1	?	-	?
additional information	Homo sapiens	human endonuclease III is a relevant target to potentiate cisplatin cytotoxicity in Y-box-binding protein-1 overexpressing tumor cells	?	-	?

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	-	-	-

Purification (Commentary)

Purification (Comment)	Organism
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.	Homo sapiens

Source Tissue

Source Tissue	Comment	Organism	Textmining
MCF-7 cell	mammary adenocarcinoma cell line	Homo sapiens	-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
additional information	-	By adding of purified Y-box-binding protein-1 to the nuclease reaction containing 0.2 microg of TAP-hNTH1 (minimum of protein required to see cleavage of the DNA duplex), there is an 12fold increase in cleavage activity	Homo sapiens
additional information	-	no activity is detected using a probe without 8-oxoguanine residue demonstrating the specificity of hNTH1 for oxidized DNA duplex	Homo sapiens
additional information	-	TAP-hNTH1 expressing cells transfected with a small interference RNA specific to hNTH1 show decreased protein levels and nuclease activities toward the radioactive duplex 8-oxoguanine	Homo sapiens
additional information	-	the MCF7-eluted TAP-hNTH1 protein exhibits nuclease activities on a radioactive duplex containing 8-oxoguanine residues	Homo sapiens
additional information	-	Y-box-binding protein-1 binds specifically to the auto-inhibitory domain of hNTH1 to stimulating its activity	Homo sapiens
additional information	-	Y-box-binding protein-1 strongly stimulates in vitro the activity of hNTH1 toward DNA duplex probes containing oxidized bases, lesions prone to be present in cisplatin treated cells	Homo sapiens

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
DNA	bifunctional enzyme is involved in base excision repair, it is a bifunctional DNA glycosylase/apurinic/apyrimidinic lyase which removes hydrated, reduced, or oxidized bases from the DNA backbone as the initial step of base excision repair	Homo sapiens	fragments of DNA	-	?
additional information	hNTH1 and Y-box-binding protein-1 may be part of the same DNA repair pathway in response to cisplatin and UV treatments	Homo sapiens	?	-	?
additional information	hNTH1 binds directly to Y-box-binding protein-1 in the absence of nucleic acids, it binds to the auto-inhibitory n-terminal tail of NTH1	Homo sapiens	?	-	?
additional information	human endonuclease III is a relevant target to potentiate cisplatin cytotoxicity in Y-box-binding protein-1 overexpressing tumor cells	Homo sapiens	?	-	?

Synonyms

Synonyms	Comment	Organism
APE/Ref-1	-	Homo sapiens
apurinic/apyrimidinic endonuclease/redox effector factor-1	-	Homo sapiens
endonuclease III	-	Homo sapiens
hNTH1	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Homo sapiens