Glutamate Decarboxylase Assay

Method: The reaction velocity is measured by a conventional Warburg manometric technique where glutamate is converted to aminobutyrate. One unit releases one micromole of CO₂ per minute at 37°C and pH 5.0 under the specified conditions.

Reagents

• 0.5 M Sodium acetate buffer, pH 5.0
• 0.05 M L-glutamate in 0.5 M sodium acetate, pH 5.0
• 3.0 M Sodium chloride

Enzyme

Immediately prior to use, dilute the enzyme with reagent grade water to a concentration of 0.5 - 3.0 units/ml.

Procedure

Into the main well of the Warburg flask pipette the following:

0.5 M sodium acetate, pH 5.0	1.0 ml
3.0 M sodium chloride	0.1 ml
Reagent grade water	1.3 ml
Enzyme dilution	0.1 ml

Pipette 0.5 ml of 0.05 M L-glutamate into side arm. Include one flask with no enzyme as a blank and one flask with 3.0 ml water as a thermal barometer. Assemble flasks and incubate for 10 - 15 minutes to achieve temperature equilibration. Tip in substrate and measure CO₂ evolution at 2 - 3 minute intervals for 30 minutes.

Calculation