Dialkylfluorophosphatases of microorganisms.

Organisms were grown for periods varying from 20 to 48 hours in a peptone-yeast extract-glucose medium buffered at pH 6.8 to 7.0 with 0.05 Y phosphate buffer. Representative types which grew satisfactorily on this medium were selected for study. Cells were harvested by centrifugation and washed three times with water. The washed cells were suspended in a small volume of water, lyophilized, and stored in a desiccator over PZ06 in a cold room (10”). Lyophilized material was used in all experiments because the results with a given culture were more reproducible and corrections for endogenous metabolism were relatively low. Known amounts of cells were suspended in 0.025 M NaHCOa, and 20 to 40 mg. of the lyophilized organisms in 0.2 to 0.3 ml. were pipetted into the side arms of the Warburg flasks. The procedures previously described (2, 3) for measuring the hydrolysis of diisopropyl fluorophosphate (DFP) were used. Control experiments were carried out for non-enzymatic hydrolysis (NaHC03 + DFP) and for the acid production as measured by metabolic CO2 evolution (NaHCOs + microorganisms). Results for DFP hydrolysis were corrected for these two blanks. When activators were added to the cell suspensions, they were incubated together for 15 minutes before mixing with the substrate.