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Glutamate Decarboxylase - Assay

CAS:
9024-58-2
EC:
4.1.1.15

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