Location of sulfate-binding protein in Salmonella typhimurium.
A method is described for location of proteins in bacteria. It depends upon two techniques. One technique is the inactivation of the protein by a reagent which is incapable of penetrating the bacterial membrane (permeability barrier). Proteins inside this membrane cannot be inactivated unless the cells are disrupted; proteins on or outside the membrane can be inactivated. The second technique depends upon inactivation of the protein by specific antibody. Antibody should not penetrate the external bacterial wall, and therefore should only inactivate proteins that are on the wall surface. Thus, proteins can be localized inside the membrane, in the wall-membrane area, or outside the wall. One reagent developed for use with the first technique is diazo-7-amino-1,3-naphthalene-disulfonate. It inactivated beta-galactoside transport, but not beta-galactosidase of intact Escherichia coli. Similarly, it inactivated sulfate binding and transport but not uridine phosphorylase activity of Salmonella typhimurium. This indicates that the sulfate-binding protein is on or outside the cell membrane, and that uridine phosphorylase is inside the cell. The organic mercurial compounds used also showed that the sensitive parts of the sulfate and alpha-methylglucoside transport systems are less reactive than the sensitive part of the beta-galactoside system. Antibody to the sulfate-binding protein inactivated the purified protein but did not inactivate this protein when intact bacteria were employed. Thus, it appears that the sulfate-binding protein does not protrude outside the cell wall. The conclusion that the binding protein is located in the wall-membrane region is supported by its release upon spheroplast formation or osmotic shock, and also by its ability to combine with sulfate in bacteria which cannot transport sulfate into the cell.
D001391:Azo Compounds / Q000494:pharmacology; D001426:Bacterial Proteins; D001692:Biological Transport / Q000187:drug effects; D002247:Carbon Isotopes; D002462:Cell Membrane / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism; D004492:Edetic Acid / Q000494:pharmacology; D004926:Escherichia coli; D005696:Galactosidases / Q000037:antagonists & inhibitors; D006027:Glycosides / Q000378:metabolism; D007106:Immune Sera / Q000494:pharmacology; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins / Q000037:antagonists & inhibitors; D008628:Mercury / Q000494:pharmacology; D011523:Protoplasts / Q000378:metabolism; D012486:Salmonella typhimurium / Q000187:drug effects / Q000378:metabolism*; D013431:Sulfates / Q000378:metabolism*; D013451:Sulfonic Acids / Q000494:pharmacology; D013460:Sulfur Isotopes; D014166:Transferases / Q000037:antagonists & inhibitors; D014529:Uridine / Q000378:metabolism
D001391:Azo Compounds; D001426:Bacterial Proteins; D002247:Carbon Isotopes; D006027:Glycosides; D007106:Immune Sera; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins; D013431:Sulfates; D013451:Sulfonic Acids; D013460:Sulfur Isotopes; D004492:Edetic Acid; D014166:Transferases; D005696:Galactosidases; D008628:Mercury; D014529:Uridine