Purification and Chemical Composition of the Protective Slime Antigen of Pseudomonas aeruginosa

Abstract

The slime obtained from Pseudomonas aeruginosa strain BI was purified by a system of ethanol precipitation, gel filtration, and ion-exchange chromatography. The slime polysaccharide was eluted as a single peak at a potassium chloride molarity of 0.30 to 0.40. The purification procedure was monitored by immunodiffusion techniques, and the number of bands was reduced from four to one, indicating the elimination of antigenic impurities that were present in the crude extracts of slime. The purified slime behaved as a homogeneous antigen, stimulating the production of a single species of antibody in rabbits. Hydrolyzed preparations of purified slime contained rhamnose, glucose, mannose, glucosamine, galactosamine, and glucuronic acid, as well as N-acetyl and O-acetyl groups. Only trace amounts of nucleic acids were detectable. A significant amount of protein was found to be associated with the carbohydrate moiety. The substrate characteristic of the slime was reaffirmed by measuring the release of hexosamines in the presence of the Pseudomonas phage 2 depolymerase PDB(2), and its activity as a protective antigen was demonstrated in passive-protection tests of mice.