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Human P450 Liver Enzymes Ames TestThe Internal P450 1A2 Bioactivation System based on recombinant human P450 enzymes

 

 

 

 The EBPI 777/2205 has a hisD3052 mutation, a -1 frameshift mutation which affects the reading frame of a repetitive -C-G-C-G-C-G-C-G- sequence on the hisD gene.  The hisD gene encodes histidinol dehydrogenase, an enzyme involved in the terminal step of histidine biosynthesis. EBPI 777 also contains a bicistronic protein expression vector, pP4501A2-red, which contains human cytochrome P450 1A2 and human NADPH-P450 reductase cDNA.  The recombinant expression of these two proteins helps in the bioactivation of drug and foreign compounds into DNA-reactive mutagens.  The co-expression of NADPH-P450 reductase with P450 1A2 increases the bioactivation level of the system. This system acts as an in vitro mimic involving the uptake and breakdown of xenobiotic compounds by a mammal and its liver enzymes, resulting in the generation of mutagenic products inside the living system. 

EBPI Ames Express bacterial strains; expression of P450 1A2 and NADPH-P450 reductase

TA1538                  hisD3052                -1 Frameshift               Recombinant Expression of P450 1A2

TA98                      hisD3052               -1 Frameshift                Recombinant Expression of P450 1A2 

TA100                    hisG46              Base-Pair Substitution       Recombinant Expression of P450 1A2

Educational Applications

Ames Test for Classroom

Click here to learn more the role of the  Ames ExpressTM Strains for educational use.

Environmental Applications

P450 Ames Testing Environment

Click here to download EBPI's Ames Express Strain Environmental Handout

Medical Toxicology Applications

Ames Test P450 Drug Testing

Click here to learn more the role of the P450 1A2 in metabolizing of drug compounds

 Another advantage with the recombinant P450 1A2 and NADPH-P450 reductase expression system is that the reactive metabolites will be generated inside the cell rather than outside the cell, as is seen with the S9 rat liver mix.  As a result, this mode of bioactivation will allow short half-life reactive mutagens enough time to reach DNA sites allowing for an increase in sensitivity to mutagenesis.

The recombinant P450 protein also serves as a cost effective and animal friendly alternative to the S9 rat liver extract mix which is the current bioactivation standard.

Metabolically Activates Carcinogens:

P450 1A2 Internal Ames Test Bioactivation

MeIQ, Hetrocyclic amine, Alfatoxin B1

2-Amino-1-methyle-6-phenylimidazo (4,5-b) pyridine (PhIP)

3-Methyl-2-methylaminoimidazo (4,5-f)quinoline (IQ)

Polycyclic Aromatic Hydrocarbons (PAH)

The Ames ExpressTM System:

This system acts as an in vitro mimic involving the uptake and breakdown of xenobiotic compounds by a mammal and its liver enzymes, resulting in the generation of mutagenic products inside the living system. The recombinant P450 1A2 protein also serves as a cost effective and animal friendly alternative to the S9 rat liver extract mix which is the current bio-activation standard . Like EBPI`s GST expressing strains, the recombinant P450 1A2 reductase expression system is that the reactive metabolites will be generated inside the cell rather than outside the cell.  As a result, this mode of bioactivation will allow short half-life reactive mutagens enough time to reach DNA sites allowing for an increase in sensitivity to mutagenesis. 

P450 1A2 is a Phase 1 enzyme found primarily in the hepatic tissue and is responsible for metabolizing polyaromatic hydrocarbons (PAHs), aromatic amines and nitro aromatics. It performs a variety of chemical reactions on several different xenobiotic classes and is a key contributor to the mechanistic mutagenicity of well known carcinogens like benzo(a)pyrene (B[a]P), PhIP and 4-aminobiphenyl.

Click Here for information on  GST T1-1 Ames Express Bacterial Strains.

Potential Applciations:

  • Testing of pharmaceuticals for mutagenic activity
  • Testing of industrial effluents for presence of possible mutagenic compounds
  • Screening of municipal discharges for possible routine presence or spills of mutagenic compounds
  • Screening of surface and/or groundwater for mutagenic residues
  • Screening of potable water supplies for the presence of chemicals with mutagenic potential
  • Screening of water soluble air pollutants for mutagenic agents
  • Evaluation of pure or complexed raw mixtures for potential mutagenicity
  • A convenient and easy to use teaching tool for university and college laboratories

Biotoxicity Products

Please feel free to download the Biotoxicity product handout.  Products include EBPI's line of toxicity testing kits, MicroBioTests line of Toxkits along with information on Microbial Insights line of molecular tools.

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