ABSTRACT
Thiobacillus ferrooxidans is a Gram-negative chemolithotrophic bacterium able to oxidize ferrous iron, elemental sulfur and inorganic sulfur compounds. The oxidation of sulfur by T. ferrooxidans resulted in an expression of some outer membrane proteins (OMPs) at a level higher than that observed during ferrous iron oxidation. Among these OMPs, a protein with a molecular mass of 54 kDa was purified and 18 amino acids of the N-terminal sequence determined. Using a 54 bp PCR generated DNA product as a probe for the protein, we isolated a 4.5 kb Pst I DNA chromosomal fragment containing the corresponding gene. Sequencing 2169 bp of this fragment revealed the open reading frame codifying for the protein, consisting of 467 amino acids and a molecular mass of 49,674 Da. The mature protein was produced by the removal of a 32 amino acid signal peptide-like sequence from the N-terminus of a 499 amino acid peptide. Although no significant homology with any known protein has been found and its physiological role remains unclear, its high expression on sulfur substrates suggests a role in sulfide mineral oxidation.
Subject(s)
Bacterial Proteins/genetics , Gene Expression Regulation, Bacterial , Sulfur/metabolism , Thiobacillus/genetics , Amino Acid Sequence , Base Sequence , Cloning, Molecular , DNA, Bacterial , Molecular Sequence DataABSTRACT
The anti-invasive effect of bovine lactoferrin (BLf) and of bovine transferrin (BTf) towards L. monocytogenes, an intracellular facultative food-borne pathogen, was assayed in the enterocyte-like cell line Caco-2. When 0.5 mg/ml BLf were added during the infection time or preincubated with bacteria the number of internalized bacteria was noticeably decreased whereas BLf was ineffective when preincubated with the enterocytes or added post infection. BTf was deprived of any effect. Results from direct binding and Western blotting assays provided evidence that two L. monocytogenes surface proteins, of approximately 80 and 60 kDa, specifically reacted with BLf. These findings strongly support the hypothesis that the antiinvasive mechanism of BLf is due to its interaction with bacterial surfaces, but not to its binding with eukaryotic cells.
ABSTRACT
The influence of iron on the entry of Listeria monocytogenes into Caco-2 cells was studied. Iron availability was found to modify the surface hydrophobicity and protein profile of L. monocytogenes, with the result that cell invasion strongly increased upon bacterial growth in iron-rich medium. The enhanced invasive capability of iron-overloaded L. monocytogenes cells correlates to the higher-level expression of the inlAB virulence genes, which were positively iron regulated at the transcriptional level.
Subject(s)
Bacterial Adhesion , Bacterial Proteins/genetics , Bacterial Toxins , Gene Expression Regulation, Bacterial , Heat-Shock Proteins/genetics , Iron/metabolism , Listeria monocytogenes/pathogenicity , Listeriosis/microbiology , Bacterial Proteins/chemistry , Caco-2 Cells , Hemolysin Proteins , Hemolysis , Humans , Listeria monocytogenes/chemistry , Listeria monocytogenes/genetics , RNA, Bacterial/genetics , RNA, Messenger/genetics , Solubility , Surface Properties , Transcription, GeneticABSTRACT
The effect of growth in iron-excess or iron-limitation conditions on the invasiveness for HeLa cells of Escherichia coli HB101 carrying plasmid pRI203 which bears the invasion gene of Yersinia pseudotuberculosis was examined. Iron-limitation reduced adhesion and the number of organisms internalised by HeLa cells by about 100-fold. The reduced adhesion of iron-starved bacteria correlated with reduced hydrophobicity and the reduced invasiveness appeared to depend on the plasmid copy number, which was 3.5-fold less than in bacteria grown in iron excess.
