[Isolation of spheroplasts from Escherichia coli 85 for aspartate-ammonia-lyase localization]

Conditions were determined for preparation of spheroplasts from E. coli under the action of lysozyme in the presence of EDTA. The preparation took from 10 to 15 min. The degree of conversion to spheroplasts was monitored spectrophotometrically at 660 nm. The spheroplasts formed were unstable in Tris-HCl buffer and immediately lysed, but they were more stable in 1 M sucrose. At lysozyme concentrations above 40 micrograms/ml of the reaction mixture, the cells lysed to a greater extent. The distribution of aspartate ammonia-lyase activity between the precipitate of the spheroplasts and the supernatant allowed the authors to suggest that aspartase should be located in the cytoplasm.