Action of ethylenediaminetetraacetic acid, tris(hydroxymethyl)-aminomethane, and lysozyme on cell walls of Pseudomonas aeruginosa.

Abstract

Incubation of isolated cell walls of Pseudomonas aeruginosa with tris(hydroxymethyl)aminomethane (Tris), ethylenediaminetetraacetic acid (EDTA), and lysozyme, either singly or in combinations, results in partial solubilization of the cell walls.When cell walls were incubated with a combination of Tris and EDTA, approximately 30% of the total cell wall and 32% of the lipopolysaccharide fraction were solubilized. The following percentages of the individual cell wall constituents were solubilized: protein, 19%; carbohydrate, 35%; lipid, 5%; ash, 54%; and, phosphorus 23%. When lysozyme was included with Tris and EDTA, approximately 36% of the total cell wall and 86% of the lipopolysaccharide fraction were solubilized. Solubilization of the individual cell wall constituents was similarly increased.Phospholipids, which make up 7–8% of the intact cell wall of P. aeruginosa, were not released by incubation of the cell walls with these agents. Fatty acids in addition to those found in the lipopolysaccharide, however, were detected in the solubilized cell wall materials.Incubation systems composed of Tris, of EDTA, and of Tris and lysozyme solubilized cell walls to a lesser extent than incubation systems of Tris and EDTA, and of Tris, EDTA, and lysozyme. Incubation of cell walls in water, furthermore, was effective in solubilizing approximately 8% of the cell walls. Muramic acid containing materials were solubilized only in the presence of lysozyme.Materials liberated from cell walls which were incubated in systems containing EDTA exhibited single symmetrical Schlieren peaks characterized by uncorrected sedimentation constants of approximately 7 S, suggesting the release of homogenous subunits from the cell walls.Intra- and inter-molecular cross-linking by divalent cations via phosphate groups contained in lipoprotein and lipopolysaccharide components of the cell wall of P. aeruginosa is proposed.