A membrane system was prepared from Escherichia coli K12 by the treatment with lysozyme [EC 3.2.1.17] and EDTA, followed by osmotic disruption in the presence of 0.005 M MgCl2. Electron microscopic observation revealed that the system was composed of cytoplasmic membranes to which an appreciable amount of ribosomes was attached. The ribosomes were released from the system upon washing with a Mg++-free medium. The system prepared with 0.005 M MgCl2 contained about 50% protein, 40% RNA, 30% DNA and 80% phosphatide of the cells. These values, except that of phosphatide, were, however, continuous functions of the concentration of magnesium ions with which the membrane systems were prepared. The membrane system actively incorporated 3H-leucine into the protein and 32P1 into the RNA and the DNA fractions. Chloramphenicol, actinomycin D and mitomycin G strongly inhibited the incorporation into protein, RNA and DNA, respectively. Qualitative difference in the membrane-bound and soluble components was investigated. Succinate oxidase system was localized specifically in the membrane, while glucose-6-phosphate dehydrogenase [EG 1.1.1.49] was detected only in the supernatant fraction. Hatch's polarity ratios of the bulk proteins were found to be 1.32 for the membrane and 1.57 for the supernatant. As much as 78% of the total phosphatides was found in the membrane system. The major lipids were phosphatidyl ethanolamine and phosphatidyl glycerol in both fractions. Possibility that the activity of the membrane system would have been attributed to the contaminated intact cells and/or undisrupted spheroplasts was eliminated from several evidences including viable cell counts of the system.
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Nagata et al. (1967) studied this question.