Low iron concentration and aconitase deficiency in a yeast frataxin homologue deficient strain.
Deletion of the yeast frataxin homologue, YFH1, elicits accumulation of iron in mitochondria and mitochondrial defects. We report here that in the presence of an iron chelator in the culture medium, the concentration of iron in mitochondria is the same in wild-type and YFH1 deletant strains. Under these conditions, the activity of the respiratory complexes is restored. However, the activity of the mitochondrial aconitase, a 4Fe-4S cluster-containing protein, remains low. The frataxin family bears homology to a bacterial protein family which confers resistance to tellurium, a metal closely related to sulfur. Yfh1p might control the synthesis of iron-sulfur clusters in mitochondria. (+info)
Tellurite-mediated thiol oxidation in Escherichia coli.
The oxyanion of tellurium, tellurite (TeO3(2-)), is toxic to most micro-organisms, particularly gram-negative bacteria. The mechanism of tellurite toxicity is presently unknown. Many heavy metals and oxyanions, including tellurite, interact with reduced thiols (RSH). To determine if tellurite interaction with RSH groups is involved in the toxicity mechanism, the RSH content of Escherichia coli cultures was assayed. After exposure to tellurite, cells were harvested and lysed in the presence of the RSH-specific reagent 5,5'-dithiobis(2-nitrobenzoic acid). Upon exposure of tellurite-susceptible cells to TeO3(2-), the RSH content decreased markedly. Resistance to potassium tellurite (Te(r)) in gram-negative bacteria is encoded by plasmids of incompatibility groups IncFI, IncP alpha, IncHI2, IncHI3 and IncHII, as well as the tehAtehB operon from the E. coli chromosome. When cells harbouring a Te(r) determinant were exposed to TeO3(2-), only a small fraction of the RSH content became oxidized. In addition to tellurite-dependent thiol oxidation, the resistance of E. coli mutants affected in proteins involved in disulfide-bond formation (dsb) was investigated. Mutant strains of dsbA and dsbB were found to be hypersensitive to tellurite (MIC 0.008-0.015 microg K2TeO3 ml(-1) compared to wild-type E. coli with MICs of 1-2 microg K2TeO3 ml(-1)). In contrast, dsbC and dsbD mutants showed no hypersensitivity. The results suggest that hypersensitivity to tellurite is reliant on the presence of an isomerase activity and not the thiol oxidase activity of the Dsb proteins. The results establish that the Te(r) determinants play an important role in maintaining homeostasis of the intracellular reducing environment within gram-negative cells through specific reactions with either TeO3(2-) or thiol:tellurium products. (+info)
Studies on the ADP-ribose pyrophosphatase subfamily of the nudix hydrolases and tentative identification of trgB, a gene associated with tellurite resistance.
Four Nudix hydrolase genes, ysa1 from Saccharomyces cerevisiae, orf209 from Escherichia coli, yqkg from Bacillus subtilis, and hi0398 from Hemophilus influenzae were amplified, cloned into an expression vector, and transformed into E. coli. The expressed proteins were purified and shown to belong to a subfamily of Nudix hydrolases active on ADP-ribose. Comparison with other members of the subfamily revealed a conserved proline 16 amino acid residues downstream of the Nudix box, common to all of the ADP-ribose pyrophosphatase subfamily. In this same region, a conserved tyrosine designates another subfamily, the diadenosine polyphosphate pyrophosphatases, while an array of eight conserved amino acids is indicative of the NADH pyrophosphatases. On the basis of these classifications, the trgB gene, a tellurite resistance factor from Rhodobacter sphaeroides, was predicted to designate an ADP-ribose pyrophosphatase. In support of this hypothesis, a highly specific ADP-ribose pyrophosphatase gene from the archaebacterium, Methanococcus jannaschii, introduced into E. coli, increased the transformant's tolerance to potassium tellurite. (+info)
Evaluation of three gamma detectors for intraoperative detection of tumors using 111In-labeled radiopharmaceuticals.
Attempts to detect tumors with intraoperative scintillation using tumor-binding radiopharmaceuticals have intensified recently. In some cases previously unknown lesions were found, but in most cases no additional lesions were detected. In this study the physical characteristics of three detector systems and their ability to detect tumors through accumulation of an 111In-labeled radiopharmaceutical were investigated. The first was a sodium iodide (NaI[TI]) detector; the second, a cesium iodide (CsI[TI]) detector; and the third, a cadmium telluride (CdTe) detector. METHODS: A body phantom and tumor phantoms (diameter 5-20 mm) made of water, agarose gel or epoxy with a density and attenuation coefficient similar to those of soft tissue were used to simulate a clinical situation. The activity concentration in the body phantom was based on reported values of 111In-octreotide in normal tissue in humans. The 111In activity concentration in the tumor phantoms varied from 3 to 80 times the 111In activity concentration in the body phantom. Data were processed to determine tumor detection levels. RESULTS: The NaI(TI) detector showed the lowest values for full width at half maximum because this detector had the best collimation, leading to a high ratio between counts from tumor and counts from background, i.e., small tumors could be detected. Because of high efficiency, the CsI(TI) detector sometimes required a somewhat shorter acquisition time to produce a statistically significant difference between tumor phantom and background. For deep-lying tumors the NaI(TI) detector was superior, whereas the CdTe detector was best suited for superficial tumors with a high activity concentration in the underlying tissue. CONCLUSION: At a maximum acquisition time of 30 s, almost all superficial tumors with a diameter of 10 mm or larger were detected if the ratio between the 111In concentration in the tumor and the 111In concentration in the background exceeded 3. However, in clinical situations, biologic variations in the uptake of 111In-octreotide in tumors and in normal tissue makes difficult the determination of a distinct detection level. For such clinical conditions, the NaI(TI) detector is the best choice because it has good resolution despite a lower efficiency. Documentation of detector characteristics is important so that clinicians can make an adequate device in relation to tumor location and receptor expression. (+info)
Selective isolation of eae-positive strains of shiga toxin-producing Escherichia coli.
Culture on cefixime, tellurite, and sorbitol-MacConkey agar after HCl treatment facilitated the growth of 410 (94%) of 436 eae-positive Shiga toxin-producing Escherichia coli (STEC) strains and 17 (16%) of 107 eae-negative STEC strains. This selectivity was closely related to acid resistance in E. coli and tellurite resistance in eae-positive STEC strains. (+info)
Production of volatile derivatives of metal(loid)s by microflora involved in anaerobic digestion of sewage sludge.
Gases released from anaerobic wastewater treatment facilities contain considerable amounts of volatile methyl and hydride derivatives of metals and metalloids, such as arsine (AsH(3)), monomethylarsine, dimethylarsine, trimethylarsine, trimethylbismuth (TMBi), elemental mercury (Hg(0)), trimethylstibine, dimethyltellurium, and tetramethyltin. Most of these compounds could be shown to be produced by pure cultures of microorganisms which are representatives of the anaerobic sewage sludge microflora, i.e., methanogenic archaea (Methanobacterium formicicum, Methanosarcina barkeri, Methanobacterium thermoautotrophicum), sulfate-reducing bacteria (Desulfovibrio vulgaris, D. gigas), and a peptolytic bacterium (Clostridium collagenovorans). Additionally, dimethylselenium and dimethyldiselenium could be detected in the headspace of most of the pure cultures. This is the first report of the production of TMBi, stibine, monomethylstibine, and dimethylstibine by a pure culture of M. formicicum. (+info)
Modification of sorbitol MacConkey medium containing cefixime and tellurite for isolation of Escherichia coli O157:H7 from radish sprouts.
A modified version of sorbitol MacConkey medium containing cefixime and tellurite (CT-SMAC medium) was produced by adding salicin and 4-methylumbelliferyl-beta-D-galactopyranoside to CT-SMAC medium; this medium was designated CT-SSMAC medium and was used to isolate Escherichia coli O157:H7 from radish sprouts. Of 101 non-E. coli bacteria isolated from radish sprouts that produced colorless colonies similar to colonies of E. coli O157:H7 grown on CT-SMAC medium, 92 (91%) formed colonies that were red to pink or were beta-galactosidase negative and colorless on CT-SSMAC medium. On the other hand, colonies of E. coli O157:H7 strains were colorless and beta-galactosidase positive on CT-SSMAC medium. Our results suggest that CT-SSMAC medium is more selective than CT-SMAC medium for isolating E. coli O157:H7. (+info)
Escherichia coli TehB requires S-adenosylmethionine as a cofactor to mediate tellurite resistance.
The Escherichia coli chromosomal determinant for tellurite resistance consists of two genes (tehA and tehB) which, when expressed on a multicopy plasmid, confer resistance to K(2)TeO(3) at 128 microg/ml, compared to the MIC of 2 microg/ml for the wild type. TehB is a cytoplasmic protein which possesses three conserved motifs (I, II, and III) found in S-adenosyl-L-methionine (SAM)-dependent non-nucleic acid methyltransferases. Replacement of the conserved aspartate residue in motif I by asparagine or alanine, or of the conserved phenylalanine in motif II by tyrosine or alanine, decreased resistance to background levels. Our results are consistent with motifs I and II in TehB being involved in SAM binding. Additionally, conformational changes in TehB are observed upon binding of both tellurite and SAM. The hydrodynamic radius of TehB measured by dynamic light scattering showed a approximately 20% decrease upon binding of both tellurite and SAM. These data suggest that TehB utilizes a methyltransferase activity in the detoxification of tellurite. (+info)