BL4820 Lecture 4 - Glutamate Oxaloacetic Acid Transaminase (GOT) Purification -- Expt 3 Part A
4. Interference with GOT Activity Assay by Glutamate Dehydrogenase (GDH)
One problem with this method of assaying GOT is that other enzymes in the pig heart extract may also catalyze the oxidation of NADH. Therefore, you will assay for glutamate dehydrogenase activity (GDH):
NADH + 2-KG + NH4+ = Glu + NAD+
This assay will be done by omitting Asp from the standard GOT assay and monitoring A-340 nm. Here is what may go on in your standard GOT assay if there is some ammonium ion in your pig heart extract which gets into the assay:
Since you are following the oxidation of NADH by monitoring change in A-340 nm, if there is ammonium ion in your extract, the side reaction shown on the left side of the graphic above will cause extra NADH oxidation in addition to that catalyzed by GOT.
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