| Species : |
Human |
| Source : |
Insect Cells |
| Tag : |
Non |
| Description : |
This gene encodes a member of the cytochrome P450 superfamily of enzymes. The cytochrome P450 proteins are monooxygenases that catalyze many reactions involved in drug metabolism and synthesis of cholesterol, steroids and other lipids. This protein localizes to the endoplasmic reticulum and is known to metabolize many xenobiotics, including the anticonvulsive drug mephenytoin, omeprazole, diazepam and some barbiturates. Polymorphism within this gene is associated with variable ability to metabolize mephenytoin, known as the poor metabolizer and extensive metabolizer phenotypes. |
| Protein Concentration : |
9.10 mg/mL as determined using the BCA assay. Buffer: 100 mM potassium phosphate (pH 7.4), 20% glycerol, 1 mM DTT, 0.1 mM EDTA. |
| Specific Content : |
121 pmol spectral P450 per milligram of protein. |
| Reductase Activity: : |
1.61 μmol of cytochrome c reduced per minute per milligram of protein. |
| Hydroxylase Activity : |
838 pmol/min/mg of protein. Mephenytoin hydroxylation reactions were performed for 15 minutes at 37°C in 0.1 M potassium phosphate in the presence of 0.3 mM S-(+)-mephenytoin and 2 mM NADPH. The reaction was stopped by the addition of 0.5 volumes of methanol containing 0.2 mM phenobarbital (internal standard). The mixture was extracted with methylene chloride. An aliquot of the organic phase was transferred to a fresh tube, and dried down at 60°C. Analytes were dissolved by addition of 100 μL of methanol. HPLC conditions: 10 μL of sample was injected on a 4.5 mm x 70 mm 3 μm C18 HPLC column. The mobile phase was 18% acetonitrile for 12 minutes with a 97.5% methanol wash for 3 minutes, followed by an equilibration step of 18% acetonitrile for 3 minutes. Flow rate was 1.2 mL per minute with detection at 204 nm. The amount of 4-hydroxymephenytoin was calculated by comparison to a standard curve of authentic 4-hydroxymephenytoin. |
| Storage : |
−80℃; Avoid multiple freeze-thaws. |
| Safety Precautions : |
Normal precautions exercised in handling laboratory reagents should be followed. The chemical, physical, and toxicological properties of these products may not, as yet, have been thoroughly investigated. We recommend the use of gloves, lab coats, and eye protection when working with any chemical reagents. |
