Isolation and analysis of the protoplast membrane of Bacillus megaterium

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Abstract
Optimum conditions were found for the lysis of B. megaterium KM by lysozyme. The age of culture, density of suspension and concentration of lysozyme affected the rate of lysis. Protoplast membranes were isolated by centrifugation of lysates and were exhaustively washed. Treatment with chloroform removed some lipid from the membranes, but about half of the total membrane lipid could be extracted only after partial acid hydrolysis. The defatted membranes consisted of protein together with variable amounts of ribonucleic-acid (RNA); carbohydrate was almost absent. Lipid accounted for 23% of the weight of the membrane, and included both neutral lipid and phos-pholipid. In both classes, branched-chain C15 acids made up about 80% of the total fatty-acid. The phospholipid was a kephalin, and contained small quantities of several amino-acids.