Ultrastructural study of Salmonella typhimurium treated with membrane-active agents: specific reaction dansylchloride with cell envelope components.
Amino groups of cell envelope proteins, lipids, and lipopolysaccharides cannot be labeled in intact cells of Salmonella typhimurium G 30 by using 5-dimethylaminonaphthalene-1-sulfonylchloride incorporated in lecithin-cholesterol vesicles. However, application of membrane-interacting agents like tris(hydroxymethyl)aminomethane (Tris)-hydrochloride, ethylenediaminetetraacetate (Na salt) (EDTA), divalent cations, and sublethal doses of the cationic antibacterial agents polymyxin B and chlorhexidine induced specific fluorescent labeling of envelope proteins and lipids but not of cytoplasmic compounds, with the exception of a soluble protein with a molecular weight of 46,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Treatment with Tris-hydrochloride buffer produced labeling of the heat-modifiable protein B/B(+) and of proteins with molecular weights of 26,000, 22,000, and below 17,000. A combination of Tris-hydrochloride and EDTA induced additional dansylation of the major protein A and of proteins of molecular weights 80,000, 60,000, and 44,000. Polymyxin B and chlorhexidine caused similar labeling patterns. In every case, except with divalent cation treatment, protein B/B(+) was the most prominently labeled species. Phosphatidylethanolamine was dansylated up to 30%. Lipopolysaccharide was not reactive under any condition or treatment. In addition, the peptidoglycan-bound lipoprotein did not react with dansylchloride in either intact or Tris-hydrochloride-treated cells. The results are discussed with regard to a possible localization of labeled and unlabeled compounds of the cell envelope on the basis of a model placing cell envelope amino groups into ion-ion interactions with anionic components of other envelope compounds like phosphate and carboxyl groups.
D001426:Bacterial Proteins / Q000302:isolation & purification*; D002473:Cell Wall / Q000187:drug effects / Q000648:ultrastructure; D002710:Chlorhexidine / Q000494:pharmacology; D003619:Dansyl Compounds; D004492:Edetic Acid / Q000494:pharmacology; D008055:Lipids / Q000302:isolation & purification*; D008070:Lipopolysaccharides / Q000302:isolation & purification; D010714:Phosphatidylethanolamines / Q000302:isolation & purification; D011112:Polymyxin B / Q000494:pharmacology; D011135:Polysaccharides, Bacterial / Q000302:isolation & purification; D012486:Salmonella typhimurium / Q000187:drug effects / Q000648:ultrastructure*; D014325:Tromethamine / Q000494:pharmacology
D001426:Bacterial Proteins; D003619:Dansyl Compounds; D008055:Lipids; D008070:Lipopolysaccharides; D010714:Phosphatidylethanolamines; D011135:Polysaccharides, Bacterial; D014325:Tromethamine; D004492:Edetic Acid; D011112:Polymyxin B; D002710:Chlorhexidine