One unit will deaminate 1.0 &mgr;mole of adenosine to inosine per min at pH 7.4 at 25° C.
Reagents
0.05 M Phosphate buffer, pH 7.4.
Adenosine, 3.6 mg/ml dissolved in buffer.
Enzyme solution in buffer (1-2 U/ml).
Procedure
Set up spectrophotometer (with strip chart and temp. control) to 265 nm and 25° C.
Into a cuvette, pipette the following:
Phosphate buffer 2.97 ml
Adenosine 0.01 ml
Incubate cuvette in spectrophotometer at 25° C for 5 minutes to attain temperature equillibration and then establish blank rate if any.
Initiate reaction by adding 0.02 ml enzyme solution.
Calculate E265nm/min
Applications
Adenosine Deaminase (EC 3.5.4.4) is widely distributed in animal tissues. The highest enzyme activities are found in the mucosa of the small intestine, in the appendix and the spleen. The enzyme is present in the cytoplasmic cell fraction and also to some extent in the nucleus. The optimum pH is 7.0 to 7.4.
