CAT reaction mixture 1

1 M Tris-Cl (pH 7.8), 50 μl

[¹⁴C]chloramphenicol (60 mCi/mMole, diluted in H₂O to 0.1 mCi/ml), 10 μl

acetyl-CoA (freshly prepared at a concentration of 3.5 mg/ml in H₂O), 20 μl

Prepare 80 μl of CAT reaction mixture 1 per 50 μl of cell lysate to be assayed. Assemble the reaction mixture immediately before use.

Acetyl-CoA is the S-acetylated form of coenzyme A. Two carbon units from fat and carbohydrate enter the citric acid cycle in this form. It is used as an acetylating cofactor in many biological reactions, as in this protocol, for acetylation of chloramphenicol by CAT.