Substructure of the cytoplasmic membrane of Bacillus megaterium. I. Method for the fractionation of "Ghosts"

Abstract

A method of fractionation of "ghosts" was devised to identify the chemical components of the cytoplasmic membrane. The method consists of dialyzing the "ghosts" against distilled water, and then dissolving the ghosts in dilute alkali. The ghosts were fractionated into four fractions by use of differential centrifugation. The components of each fraction were analyzed in detail. The ratio of lipid to protein and content of carbohydrate were found to be different for the four fractions. The main two fractions (fractions 2 and 3) contained several types of materials. Fraction 2, which is soluble in alkali and sedimentable at 105,000 x g, contained protein and lipid in the ratio of 2:1; ribonucleic acid was not detectable. Under the electron microscope, the ghosts appeared to have released the cell's cytoplasmic contents, but many small dense particles (about 100 A in diameter) remained adherent to the membrane surface. On the other hand, fraction 2 appeared to be made up only of a membrane structure. No 100 A particles were visible in this fraction. From these results, fraction 2 seemed to be pure membrane material.