A lipopolysaccharide-binding cell-surface protein from Salmonella minnesota. Isolation, partial characterization and occurrence in different Enterobacteriaceae.

1. Protein extracts obtained from Salmonella minnesota Re mutant cells by treatment with EDTA/NaC1 solution contain a protein which exhibits high affinity to bacterial lipopolysaccharides. The isolation and partial characterization of this lipopolysaccharide-binding protein is described. 2. The protein was purified from EDTA extracts by a two-step procedure consisting of ion-exchange chromatography on CM-Sephadex and preparative polyacrylamide gel electrophoresis at pH 9.5. The yield of the total purification procedure was around 16%. 3. The resulting protein preparation was homogeneous on the basis of disc gel electrophoresis, dodecylsulfate gel electrophoresis, isoelectric focusing in polyacrylamide gel and immunoelectrophoresis. 4. The isoelectric point of the protein was found to be 10.3 at 4 degrees C. Its molecular weight determined by dodecylsulfate gel electrophoresis is 15000. Its amino acid composition is characterized by the absence of histidine and proline, a low content in tyrosine and high amounts of alanine, lysine, aspartic and glutamic acid residues, or their respective amides. 5. The lipopolysaccharide-protein association was shown to be mainly due to ionic interactions of the basic protein with negatively charged groups (probably phosphate and pyrophosphate groups) of the lipid A moiety. 6. Purified lipopolysaccharide-binding protein is immunogenic in rabbits, thus enabling the preparation of specific antiserum. 7. The protein is located at the surface of Salmonella minnesota Re mutant cells as revealed by antiserum absorption with total bacteria. Ferritin-labelling studies further demonstrated that it is evenly spread over the entire cell surface. 8. Comparative antiserum absorption studies using smooth and rough strains of Salmonella minnesota, Salmonella typhimurium, Escherichia coli, Klebsiella and Shigella revealed the presence of lipopolysaccharide-binding protein (or a serologically cross-reacting antigen) in most of the strains tested. From these results the protein can be considered as a common antigen of Enterobacteriaceae.