The Activity of a Deaminase

Most bacteria possess a battery of enzymes that specifically break down individual amino acids. In the process, the amine group on the molecule is removed and the amino acid is degraded, the reaction being known as deamination. The deaminases that effect this type of change are named for the particular amino acid substrate for which they are specific. In this experiment, we will see the effects of a phenylalanine deaminase (PDase) produced by some bacteria.

When the amino acid phenylalanine is incorporated into a culture medium in which PDase-producing bacteria are growing, the substrate is degraded to phenylpyruvic acid. The reaction is made visible by adding ferric ions, which react with the newly produced acid to form a green compound. The appearance of a green color in a medium that was colorless when inoculated is evidence of the activity of the deaminase (see colorplate 22).

Purpose	To observe the activity of PDase and to distinguish bacteria that produce it from those that do not
Materials	Slants of phenylalanine agar Dropping bottle containing 10% ferric chloride Slant cultures of Escherichia coli and Providencia stuartii

Procedures

1. Inoculate each of the two cultures on a separate slant of phenylalanine agar.
2. Incubate the new cultures at 35°C for 24 hours.
3. Examine the tubes for heavy growth. If it is adequate, run a few drops of 10% ferric chloride solution down the surface of each slant.
4. Observe the tubes for development of a green color.

Results