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Human DNA Repair Enzymes
Human AP Endonuclease (APE/Ref-1)
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| Human AP
Endonuclease |
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| The Human AP Endonuclease (APE) is a multifunctional enzyme involved both in DNA repair and in facilitating the redox state of a number of DNA binding proteins. The DNA repair function of APE serves to cleave DNA at apurinic/apyrimidinic sites forming nicks in DNA which are subsequently repaired by DNA polymerases and ligases. The second, and seemingly unrelated function is the regulation of several transcription factors through the redox stimulation of DNA binding activity. These transcription factors include FOS, JUN, NF-Kb, and p53. The APE related DNA repair activity has been demonstrated to be inactivated by phosphorylation suggesting that this might be the mechanism by which the enzyme switches between the two functions. | |
| Unit Definition: | One unit is the amount of enzyme required to cleave an AP-site oligonucleotide within an oligonucleotide duplex at the rate of 1 pmol/hour at 37°C. |
| Source: | Purified from E. coli containing a recombinant plasmid harboring the human APE gene. |
| Assay Conditions: | 1X REC™ Buffer 7 (10 mM HEPES-KOH (pH 7.4), 100 mM KCl, 10 mM MgCl2), 4pmoles AP site oligonucleotide (Cat# 3851-100-01) labeled with 32P, 4 pmoles Oligo Complement B (Cat# 3849-100-02), and 1 unit human AP endonuclease in a 20 µl reaction volume and incubated for 1 hour at 37°C. |
| Applications: | • CometAssay™/FLARE™ • AP site cleavage • Redox activation of transcription factors |
| Storage: | Freeze in working aliquots at -20°C in a manual defrost freezer to avoid repeated freeze-thawing. The enzyme is supplied in buffer containing 50% glycerol. |
Products
| 4110-01K-EB | Human AP Endonuclease, 1000 Units and 10X REC™ Buffer 7, 1 ml |
| 4110-05K-EB | Human AP Endonuclease, 5000 Units and 10X REC™ Buffer 7, 5 ml |
| Human DNA Polymerase
ß |
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| We have good
news to share with you regarding our Human DNA Polymerase ß. Our efforts
to improve the expression and purification of the protein have
succeeded, and we are now able to provide the product in greater amounts
at significant cost savings. We have doubled the number of units in the
smaller size, and we now offer larger sizes at dramatically reducted
cost per unit activity. Human DNA Polymerase ß is constitutively expressed in cells and is thought to participate in the repair of DNA. The enzyme functions by filling in gaps in DNA, possibly formed following base excision repair. The activity of DNA Polymerase ß is not affected by aphidocolin. |
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| Unit Definition: | One unit is the amount of enzyme required to catalyze the incorporation of 1 nmole of dNTP into an acid-insoluble form in 60 minutes at 37°C. | ||||||||
| Source: | Purified from E. coli containing a recombinant plasmid harboring the human DNA Polymerase ß gene. | ||||||||
| Assay Conditions: | 1X REC™ Buffer 8 (50 mM Tris-Cl (pH 8.8), 10 mM MgCl2 , 100 mM KCl, 1.0 mM DTT, 10% glycerol), 50 µM dCTP, 50 µM dGTP, 50 µM dTTP, 50 µM á - 32P-dATP, and 10 µg of activated DNA in a reaction volume of 50 µl are incubated for 15 min at 37°C. | ||||||||
| Applications: |
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| Components: | Human DNA Polymerase ß Kit | ||||||||
| Catalog # | Component | Size | |||||||
| 4020-050-01
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Human DNA Polymerase ß | 50 Units | |||||||
| 4020-050-02 | ß-polymerase Control DNA | 5 µg | |||||||
| 3900-200-08 | 10X REC Buffer 8 | 1 ml | |||||||
| 4018-250 5X | REC Loading Buffer | 250 µl | |||||||
| 4019-1 | REC Water | 1 ml | |||||||
| 4020-050-04 | 1 mM Aphidicolin | 10 µl | |||||||
| Storage: | Store at -20°C in a manual defrost freezer. The enzyme is supplied in buffer containing 50% glycerol. |
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Products
| 4020-100-EB | Human DNA Polymerase ß Enzyme and Buffer, 100 Units |
| 4020-100-K | Human DNA Polymerase ß Kit, 100 Units |
| 4020-500-EB | Human DNA Polymerase ß, 500 Units |
| 4020-01K-EB | Human DNA Polymerase ß, 1000 Units |
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Human Fen-1
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| Fen-1 (Flap
Endonuclease) is an endonuclease/exonuclease that functions in base
excision repair. The enzyme shows similarity to the yeast Rad2 and Rad13
genes. Fen-1 was identified as a necessary component of Okazaki fragment
processing and functions on a number of branched DNA structures.
Recently it was shown that Fen-1 specifically associates with PCNA and
this binding stimulates Fen-1 up to 50 fold.
Notice: The supply of this Product by Trevigen conveys to you only a limited, non-transferable right to use this Product (or materials made from this Product) for purposes other than therapeutic and prophylactic applications and/or products, including, but not limited to drug screening or development. Should you desire to use this Product (or materials made from this Product) for therapeutic or prophylactic applications or products, please contact Athersys, Inc., Cleveland, Ohio, U.S.A. (www.athersys.com). |
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| Unit Definition: | One unit is the amount of enzyme required to cleave a flap DNA structure in a DNA duplex at the rate of 1 pmol/hour at 37°C. |
| Source: | Purified from E. coli containing a recombinant plasmid harboring the human FEN-1 gene. |
| Assay Conditions: | 1X REC™ Reaction Buffer 12 (50 mM Tris-HCl (pH 8.0), 10 mM MnCl2 , and 1 mM DTT), 1X BSA Additive (0.1 mg/ml BSA, 5% glycerol), 50 µl of the above 32P Flap substrate, and serial dilutions of enzyme in a reaction volume of 10 µl, incubated for 1 hour at 30°C. The reactions were stopped by the addition of 5X REC™ Loading Buffer (Cat# 4018-250) and products are resolved by 20% denaturing polyacrylamide gel electrophoresis. |
| Applications: | Excision repair research Double-strand break research |
| Storage: | Freeze in working aliquots at -20°C in a manual defrost freezer to avoid repeated freeze-thawing. The enzyme is supplied in buffer containing 50% glycerol. |
Products
| 4120-100-EB | Recombinant Human Fen-1, 100 Units and 10X REC™ Buffer 12, 1ml 10X BSA Additive, 1 ml |
| Human DNA Ligase
IV/XRCC4 Tetramer |
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| Human DNA Ligase IV/XRCC4 participates in the repair of DNA double-strand breaks in DNA via a direct nonhomologous end-joining (NHEJ) pathway. The repair involves the concerted action of several proteins, including heterodimeric Ku70/Ku80, DNA-dependent protein kinase, other accessory factors, and DNA ligase IV/CRCC4, which carries out the actual ligation reaction. DNA Ligase IV/XRCC4 exists as a tetramer containing two copies of each polypeptide with a molecular weight of approximately 300 kDa. The tetramer functions together with Ku protein, DNA-dependent protein kinase catalytic subunit, and other repair factors in a cell-free end-joining assay. | |||||||
| Source: | Recombinant human DNA Ligase IV and XRCC4. | ||||||
| Unit Definition: | One unit is the amount of enzyme required to ligate oligo (dT)24 when hybridized to poly (dA)300 at the rate of 1 pmole in 30 minutes at 25°C. | ||||||
| Assay Conditions: | Reactions of 20 µl contain 4 pmoles of 32P-oligo(dT)24 , 0.2 pmole of poly(dA)300 , 1X REC™ Buffer 13 (50 mM Tris-Cl (pH 8.0), 10 mM MgCl 2 , 25 µg/ml BSA, 10 mM DTT, 1 mM ATP), and serial dilutions of DNA Ligase IV/XRCC4 are incubated at 25°C for 30 minutes. Ligation products are resolved by 20% denaturing polyacrylamide gel electrophoresis, detected by autradiography, and quantitated. | ||||||
| Applications: |
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| Storage: | Freeze in working aliquots at -80°C in a manual defrost freezer to avoid repeat freeze-thawing. The enzyme is supplied in a buffer containing 20 mM Tris-Cl (pH 7.8), 1 mM EDTA, 50% glycerol, 0.1 M potassium acetate, and 1 mM DTT. | ||||||
Products
| 4140-100-EB | Human DNA Ligase IV/XRCC4 tetramer, 100 Units and 10X REC™ Buffer 13, 1 ml |
| 4140-500-EB | Human DNA Ligase IV/XRCC4 Tetramer, 500 Units and 10X REC™ Buffer 13, 1 ml |
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