L‐asparaginase synthesis by Escherichia coli B

We have studied the influence of strain of organism, temperature, and medium on the production of the antileukemic intracellular enzyme L‐asparaginase by E. coli B grown in shaken flasks. Five strains of E. coli B exhibited wide differences in their capacities to synthesize the EC‐2 form of L‐asparaginase active against leukemia. For the most productive strain, when grown in a casein hydrolysate medium, maximal production of L‐asparaginase occurred at 25°C. At this temperature, the organism required glycerol, glucose, or other mono‐saccharides to synthesize L‐asparaginase. Synthesis was stimulated when glycerol was used in place of glucose, but not in its presence. The effect of glycerol on L‐asparaginase synthesis was most evident when the cells were grown at 37°C, rather than at 25°C. With 0.25% glucose, cells had a specific activity of 409 I.U./g; with glycerol cells had a specific activity of 553 I.U./g. At 25°C, both cell and L‐asparaginase synthesis were increased by the use of 0.25% glycerol resulting in only a slight increase in specific activity of the cells. The addition of zinc, copper, manganese, iron, L‐asparagine, L‐glutamine, or L‐aspartic acid had no effect on L‐asparaginase synthesis in the casein hydrolysate medium. L‐aspartic acid (10−2 M) enhanced L‐asparaginase synthesis in a synthetic medium that lacked these metals or L‐asparagine, L‐glutamine, or L‐aspartic acid; cells grown under these conditions had a specific activity of 90 I.U./g. In the casein hydrolysate medium, cell morphology was correlated with temperature of incubation.