- Open Access
Proteomic profiling of L-cysteine induced selenite resistance in Enterobacter sp. YSU
© Jasenec et al; licensee BioMed Central Ltd. 2009
- Received: 6 May 2009
- Accepted: 28 August 2009
- Published: 28 August 2009
Enterobacter sp. YSU is resistant to several different heavy metal salts, including selenite. A previous study using M-9 minimal medium showed that when the selenite concentration was 100,000 times higher than the sulfate concentration, selenite entered Escherichia coli cells using two pathways: a specific and a non-specific pathway. In the specific pathway, selenite entered the cells through a yet to be characterized channel dedicated for selenite. In the non-specific pathway, selenite entered the cells through a sulfate permease channel. Addition of L-cystine, an L-cysteine dimer, appeared to indirectly decrease selenite import into the cell through the non-specific pathway. However, it did not affect the level of selenite transport into the cell through the specific pathway.
Growth curves using M-9 minimal medium containing 40 mM selenite and 1 mM sulfate showed that Enterobacter sp. YSU grew when L-cysteine was present but died when it was absent. Differential protein expression analysis by two dimensional gel electrophoresis showed that CysK was present in cultures containing selenite and lacking L-cysteine but absent in cultures containing both selenite and L-cysteine. Additional RT-PCR studies demonstrated that transcripts for the sulfate permease genes, cysA, cysT and cysW, were down-regulated in the presence of L-cysteine.
L-cysteine appeared to confer selenite resistance upon Enterobacter sp. YSU by decreasing the level of selenite transport into the cell through the non-specific pathway.
- Sequence Coverage
- Outer Membrane Protein
- Elemental Selenium
Selenium is an important cofactor in some mammalian and bacterial enzymes [1–5]. It is found in mammalian glutathione peroxidase  and bacterial formate dehydrogenase  in the form of selenocysteine. A series of Escherichia coli (E. coli) proteins, SelA, SelB, SelC and SelD, incorporate selenium into selenocysteine which is then inserted into proteins. The mechanisms that transport selenite into the cell are not well understood. Once it enters the cell, it may be reduced to selenide by glutathione  or thioredoxin . Then, SelD, a selenophosphate synthetase, catalyzes a reaction with selenide and ATP to make selenophosphate [10, 11]. SelA, a selenocysteine synthetase, uses selenophosphate to convert serine to selenocysteine , which is carried by SelC, a special tRNA . Finally, SelB, a translation factor, inserts selenocysteine at a UGA stop codon. The mRNAs of proteins that encode selenocysteine also contain a special stem loop structure or selenocysteine insertion sequence (SECIS) which stalls the ribosome and allows SelB to insert selenocysteine at the UGA stop codon [14, 15].
Although selenium is an important cofactor, too much can be toxic. Selenium inhibits the growth of E. coli when it is present at overabundant concentrations as selenite, but non-toxic when it is present as elemental selenium . Selenite reacts with glutathione and other thiol containing proteins to produce highly reactive superoxides [16–19], which may kill the cells by damaging their DNA and lipids [18, 20].
Selenite and selenate resistant bacteria appear to remove excess selenium by reducing it to elemental selenium or methylating it . A strain of Stenotrophomonas maltophilia (S. maltophilia) isolated from a selenium-contaminated drainage pond reduces selenate and selenite to elemental selenium and deposits it near the cell surface and in the surrounding growth medium . In addition, Cupriavidus metallidurans (C. metallidurans) CH34, a multi-metal resistant strain  from a zinc contaminated site, is resistant to selenate and selenite and reduces both oxyanions to elemental selenium and alkyl selenide under aerobic conditions [23, 24]. A C. metallidurans CH34 protein, DedA, which was identified by transposon mutagenesis, may be involved in selenite uptake . A set of E. coli proteins, YgfK, YgfM and YgfN, which were also identified by transposon mutagenesis, act as an enzyme that reduces selenate to elemental selenium . Finally, some bacterial strains also methylate selenite and selenate by encoding a thiopurine methyltransferase, which produces volatile dimethyl selenide and dimethyl diselenide [27, 28].
Understanding how selenite enters the cell is important to understanding the mechanisms of selenite resistance. An early study in E. coli shows that selenite and sulfate follow a Michaelis-Menten model when they are transported into the cell and compete competitively with each other . Using a low sulfate, M-9 minimal medium containing 10 μM sulfate and 75Se-labeled selenite mixed with 1 M unlabeled selenite, Muller et al define two pathways for selenium incorporation in E. coli: (1) specific and (2) non-specific [30, 31]. In the specific pathway, selenite enters the cell through a pathway dedicated for selenite and becomes incorporated into selenocysteine intentionally. In the non-specific pathway, selenite enters the cell through the sulfate transport system and becomes incorporated unintentionally to produce selenocysteine instead of L-cysteine. When 10 μg/ml of L-cystine, an L-cysteine dimer, is added to the medium, the wild type E. coli strain incorporates selenium randomly into cellular proteins, presumably via the non-specific pathway. However, when 60 μg/ml L-cystine is added to the medium, the amount of randomly incorporated selenium in wild type E. coli decreases dramatically. These results suggest that feedback inhibition blocks both the biosynthesis pathway for L-cysteine and the accidental incorporation of selenium into proteins. In support of this hypothesis, cysK mutants, which lack the ability to add sulfide to O-acetylserine to generate L-cysteine, fail to randomly incorporate radioactive selenite into protein through the non-specific pathway [30, 31]. Thus, under most conditions when selenite concentrations are lower than sulfate concentrations, selenite is excluded from the non-specific pathway. However, when selenite concentrations are higher than sulfate concentrations, selenite is transported into the cell through the non-specific pathway instead of sulfate.
Enterobacter sp. YSU is a multi-metal resistant strain that grows in the presence of mercury, cadmium, zinc, gold, silver, arsenic and selenium . It does not grow in M-9 minimal medium containing 1 mM sulfate and 40 mM selenite. However, growth inhibition is relieved by the addition of L-cysteine. Here, we define L-cysteine-dependent selenite resistance in Enterobacter sp. YSU using growth curves, 2-D gel electrophoresis and reverse-transcriptase-polymerase chain reactions (RT-PCR).
Requirement of L-cysteine for selenite resistance
Growth curves used for proteomic analysis
Two stationary phase cultures of Enterobacter sp. YSU grown in M-9 minimal medium containing and lacking 40 μg/ml L-cysteine were diluted 1:20 into fresh medium containing and lacking L-cysteine. Growth was followed by measuring turbidity every 30 minutes. After 2.5 hours of growth, the NCNS and CNS samples were taken for protein analysis. The cultures were split into equal volumes, and selenite or water was added to give the four conditions. One hour after selenite was added, the NCS and CS samples were taken for protein analysis.
Two-dimensional gel electrophoresis (2DGE)
Proteins in spots that appeared at equivalent intensities in gels of samples prepared from cells grown under all four conditions
Identification of select protein spots that appeared under all four conditions (NCNS, NCS, CNS and CS)a.
NCBI Accession (Version)
Theo. Mr/pI (kDa)
Estimated Expt. Mr/pI (kDa)
MS/MS Peptide Sequence
Equally intense under all conditions
Equally intense under all conditions
protein chain elongation factor EF-Ts
NCNS & NCS
conserved hypothetical lipobinding protein
Only in NCS
branched-chain amino-acid aminotransferase
Only in NCS
outer membrane protein II
NCNS & NS
translation elongation factor EF-Tu
NCNS & NS
translation elongation factor EF-Tu
In 6 but not in 5
7, 10, 11
NCS & CS
outer membrane protein II
NCS & CS
NCS & CS
putative membrane component hydrogenase
Only in spot 7, NCS
Only in spot 10
60 kDa chaperonin (groEL)
Only in NCS
putative tellurium resistance protein C
similar to GroES protein
outer membrane protein II
Only in NCS
small heat shock protein
Proteins in a spot that appeared at a higher intensity in gels of samples prepared from cells grown in the absence of L-cysteine
Spot 3, which is identified by a purple arrow (Fig 3), appeared at a higher intensity in the gels of samples prepared from cells grown in the absence of L-cysteine (NCNS and NCS). This spot contained peptides that matched to a conserved hypothetical lipobinding protein from Erwinia pyrifoliae with a Mascot score of 153 and a sequence coverage of 7% (Table 1). Spot 3 from the NCS gel also contained peptides that matched to two other proteins: a branched-chain amino-acid aminotransferase from Yersinia pestis with a Mascot score of 277 and a sequence coverage of 25% and an outer membrane protein II from Enterobacter aerogenes (E. aerogenes) with a Mascot score of 104 and a sequence coverage of 9%.
Proteins in spots that appeared at higher intensities in gels of samples from cells grown in the absence of selenite
Three spots, which are identified by dark red arrows (Fig 3), appeared at higher intensities on gels that contained samples prepared from cells grown in the absence of selenite (NCNS and CNS). Spot 4 from the NCNS and CNS gels was unique. This spot contained peptides that matched to an EF-Tu protein from S. typhimurium with Mascot scores of 1143 and 1467 and sequence coverages of 64% and 52%, respectively (Table 1). In addition, spots 5 and 6 were more intense in gels containing samples prepared from cells grown in the absence of selenite (NCNS and CNS). They contained peptides that matched to EF-Tu from S. typhimurium with Mascot scores of 611 and 666 and sequence coverages of 25% and 30%, respectively. Spot 6 also contained peptides that matched to 5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase (MetE) from Erwinia carotovora with a Mascot score of 173 and a sequence coverage of 10%.
Proteins in spots that appeared at higher intensities in gels of samples prepared from cells grown in the presence of selenite
Five spots, which are identified by red arrows (Fig 3), appeared at higher intensities in gels of samples prepared from cells grown in the presence of selenite (NCS and CS). Spot 7 was the largest in the CS gels. In the no L-cysteine, selenite gel (NCS), it contained peptides that matched to 4 proteins (Table 1): an outer membrane protein II from E. aerogenes with a Mascot score of 921 and a sequence coverage of 86%, OmpA from E. sakazakii with a Mascot score of 700 and a sequence coverage of 40%, a putative membrane component hydrogenase from S. typhimurium with a Mascot score of 561 and a sequence coverage of 25% and a CysK protein from E. coli with a Mascot score of 245 and a sequence coverage of 16%. In the CS gels, it contained a similar set of peptides for 3 proteins: an outer membrane protein II from E. aerogenes with a Mascot score of 960 and a sequence coverage of 73%, OmpA from E. sakazakii with a Mascot score of 696 and a sequence coverage of 38% and a putative membrane component hydrogenase from S. typhimurium with a Mascot score of 549 and a sequence coverage of 25%. It lacked peptides that matched to CysK.
Under selenite sensitive condition (NCS), spots 8 and 9 were unique (Fig 3). They did not appear in gels of samples prepared from cells grown under the other three conditions. Spot 8 contained peptides that matched to 4 proteins (Table 1): a putative tellurium resistance protein C from E. coli  with a Mascot score of 516 and a sequence coverage of 52%, a protein similar to GroES from Klebsiella pneumoniae with a Mascot score of 184 and a sequence coverage of 46%, an outer membrane protein II from E. aerogenes with a Mascot score of 191 and a sequence coverage of 18% and OmpA from E. sakazakii with a Mascot score of 165 and a sequence coverage of 8%. Spot 9 contained peptides that matched to a small heat shock protein from S. typhimurium with a Mascot score of 299 and a sequence coverage of 32%.
Spots 10 and 11 were unique on the gels containing protein samples prepared from cells treated with selenite (NCS and CS). Spot 10 contained peptides that matched to 4 proteins: an outer membrane protein II from E. aerogenes with a Mascot score of 917 and a sequence coverage of 73%, OmpA from E. sakazakii with a Mascot score of 633 and a sequence coverage of 35%, a putative membrane component hydrogenase from S. typhimurium with a Mascot score of 501 and a sequence coverage of 22% and GroEL from Pantoea agglomerans with a Mascot score of 333 and a sequence coverage of 15%. Spot 11 contained peptides that matched to 3 proteins: outer membrane protein II from E. aerogenes with a Mascot score of 641 and a sequence coverage of 52%, OmpA from E. sakazakii with a Mascot score of 434 and a sequence coverage of 37% and a putative membrane component hydrogenase from S. typhimurium with a Mascot score of 362 and a sequence coverage of 20%. It lacked peptides for GroEL.
Detection of cysK, cysE and sulfate permease transcripts in cells grown in the presence and absence of L-cysteine
A previous proteomic study on selenite tolerant E. coli cells used M-9 minimal medium that lacked L-cysteine . When two derivative K12 strains, MC4100 and GC4468, growing at logarithmic phase were pretreated with 0.25 mM selenite for 60 minutes and then exposed to 25 mM selenite, they were able to tolerate the increased selenite concentrations and were even able to grow for one hour. After this point, the cells entered into a stationary phase and began to die. This increased tolerance was attributed to higher expression levels of superoxide dismutase in response to selenite induced oxidative stress. The current work carried these E. coli studies one step further by demonstrating that Enterobacter sp. YSU was also sensitive to selenite in M-9 minimal medium, but the selenite inhibition was relieved by supplementing the medium with L-cysteine (Figs 1 and 2).
The role of L-cysteine in the observed selenite resistance/sensitivity phenotypes was investigated using proteomics. Interestingly, spot 7 from cells grown under the selenite sensitive conditions (NCS) contained peptides for CysK (Fig 3 and Table 1), whereas the same spot from cells grown under selenite resistant conditions (CS) did not. This result supported the hypothesis that L-cysteine conferred selenite resistance to Enterobacter sp. YSU by preventing selenite uptake through the non-specific pathway or sulfate permease system. The presence of L-cysteine probably caused feedback inhibition of CysE, which converted serine to O-acetylserine [30, 35]. Then, lower levels of the O-acetylserine, the derivative of N-acetylserine, which acted as an inducer for the L-cysteine synthesis genes, resulted in decreased expression of cysK and the sulfate transport genes. Although the RT-PCR experiments did not conclusively show that the addition of L-cysteine to the growth medium reduced the level of cysK transcripts, they did suggest that L-cysteine indirectly repressed the expression of the sulfate transport genes. By decreasing the level of uptake through the sulfate permease system, L-cysteine allowed the cells to grow even when the selenite concentrations were 40 times higher than the sulfate concentrations.
Under the selenite sensitive conditions (NCS), Enterobacter sp. YSU appeared to respond to selenite by expressing a putative tellurium resistance protein C [5, 34] in spot 8. Since tellurium is located directly under selenium in the periodic table of elements and is chemically similar to selenium , this protein may have been expressed non-specifically in response to selenite but was not successful in conferring resistance. The association of a GroES-like protein with the tellurium resistance protein and the appearance of the small heat shock protein in unique spot 9 suggested that these cells were probably experiencing selenite-induced oxidative stress .
The function of other identified proteins in spots 7, 10 and 11, which were present at higher intensities in cultures grown in the presence of selenite (NCS and CS) than in cultures grown in the absence of selenite (NCNS and CNS), was not clear. They all contained peptides for 3 proteins: outer membrane protein II (OmpA) from E. aerogenes , OmpA from E. sakazakii and a putative component membrane hydrogenase of OmpA. Basic Local Alignment Search Tool (BLAST) analysis and the references associated with the BLAST results did not provide any additional information about these polypeptides. However, an anaerobic strain of Clostridium pasteurianum reduced selenite using a hydrogenase during anaerobic respiration , and other Gram negative bacteria such as Stenotrophomonas maltophilia and Enterobacter sp. SLD1a-1 reduced selenite and selenate and deposited elemental selenium just inside or outside the cell surface [21, 39]. Since the three proteins in spots 7, 10 and 11 shared many of the same peptides (Table 1), they may form a single OmpA-like protein that reduced selenite to elemental selenium using the hydrogenase component. Further studies are needed to understand the role that they played in selenite resistance.
Viable cell count and turbidometric growth curves in M-9 minimal medium showed that Enterobacter sp. YSU required L-cysteine to be resistant to 40 mM selenite. Selenite can enter E. coli through a specific, undefined pathway and a non-specific sulfate permease pathway . Proteomic and RT-PCR analysis of Enterobacter sp. YSU cultures grown in the absence of L-cysteine and presence of selenite (NCS) and in the presence of L-cysteine and selenite (CS) suggested that L-cysteine conferred selenite resistance by feedback inhibition of the synthesis of N-acetylserine. This intermediate in L-cysteine synthesis acted as an inducer for cysK and the sulfate permease genes, cysA, cysT, and cysW. The lower levels of inducer decreased the expression of sulfate permease and may have limited selenite transport into the cells through the non-specific pathway, allowing the bacteria survive. This work linked studies on selenite tolerance in M-9 medium lacking L-cysteine  with research on selenite transport in E. coli .
Bacterial strain and media
M-9 minimal medium  was described previously, and 5X M-9 Salts were obtained from Becton, Dickinson and Company (Sparks, MD). When required, M-9 medium was supplemented with 0.04 mg/ml L-cysteine (Fisher Scientific, Fair Lawn, NJ) and 40 mM sodium selenite (MP Biomedicals, Aurora, OH). Luria-Bertani (LB) medium  and agar were obtained from Fisher Scientific. Enterobacter sp. YSU was described previously .
Viable cell count growth curves
Four Enterobacter sp. YSU cultures, two containing L-cysteine and two lacking L-cysteine, were grown in M-9 minimal overnight at 37°C and diluted 1:20 into corresponding fresh M-9 minimal medium containing or lacking L-cysteine. These four new cultures were grown at 37°C with shaking. After 1.5 hours of growth, selenite or an equal volume of water was added to give the following culture conditions: no L-cysteine and no selenite (NCNS), no L-cysteine and selenite (NCS), L-cysteine and no selenite (CNS), and L-cysteine and selenite (CS). Samples from each culture were removed every 45 minutes, serially diluted and plated on LB-agar medium in triplicate. Plates were incubated overnight at 37°C, and colony forming units (CFUs) were counted.
Proteomic analysis growth curves
Two Enterobacter sp. YSU cultures, one containing L-cysteine and the other lacking L-cysteine, were grown in M-9 minimal overnight at 37°C and diluted 1:20 into corresponding fresh M-9 minimal medium containing or lacking L-cysteine. These new cultures were grown at 37°C with shaking, and turbidity was measured every 0.5 hour using a Klett Colorimeter with a KS-54 filter. After 2.5 hours of growth, the two cultures were divided into equal volumes. Sodium selenite or an equal volume of water was added to give the four NCNS, NCS, CNS and SC growth conditions. Immediately before and one hour after the addition of sodium selenite, samples were harvested by centrifugation at 5,000 × g and 4°C for 10 minutes, and cell pellets were stored at -80°C.
Cells were thawed and resuspended in lysing buffer containing 8 M urea, 2 M thiourea, 2% (w/v) 3-[(3-cholamidopropyl)dimethy-ammonio]-1-propanesulfonate (CHAPS), 2% (w/v) SB 3-10, 40 mM tris(hydroxymethyl)aminomethane (Tris) (Amresco, Solon, OH), 0.2% (v/v) Bio-Lyte® 3/10 ampholyte (Bio Rad), 1% (v/v) tributylphosphine (TBP) (Bio Rad) and 1% (v/v) Halt Protease Inhibitor Cocktail (Pierce, Rockford, IL). Cells were lysed using a MiniBeadbeater-8 (BioSpec, Bartlesville, OK) and mixed to final concentrations of 0.2 μg/ml RNase (Amresco) and 200 U/ml DNase I (Pierce). After centrifuging the lysate for 10 minutes at 16,000 × g, the supernatant was treated with Bio-Rad's 2D Clean Up Kit™ (Bio-Rad) and the final pellet was resuspended in rehydration buffer containing 8 M urea, 1% (w/v) CHAPS, 15 mM dithiothreitol (DTT), trace Bromophenol Blue (Amresco), and 0.2% (w/v) Bio-Lyte® 3/10 ampholyte (Amresco). A modified Bradford assay [41, 42] was used to determine protein concentrations before 2DGE analysis.
Two-dimensional gel electrophoresis (2DGE)
Isoelectric focusing  was carried out using a Bio-Rad Protean IEF Cell (Bio-Rad). A total of 150 μg of protein was separated using an 11 cm IPG Ready Strip (Bio-Rad) with a fixed pH range of 4-7. After active rehydration at 50 V and 20°C for 12 hours, the sample was focused, starting at 0 V and ending at 8,000 V for a total of 40,000 volt hours.
After focusing, the IPG Ready Strip was washed with Equilibration Buffer I containing 6 M urea, 2% (w/v) SDS, 0.375 M Tris-HCl pH 8.8, 20% (v/v) glycerol and 130 mM DTT for 10 minutes and with Equilibration Buffer II containing 6 M urea, 2% (w/v) SDS, 0.375 M Tris-HCl pH 8.8, 20% (v/v) glycerol, and 135 mM iodoacetamide for an additional 10 minutes. The IPG strip was then submerged in 1× tris glycine SDS (TGS) buffer containing 0.025 M tris base, 0.192 M glycine and 0.1% (w/v) sodium dodecyl sulfate (SDS) (Amresco) before being placed in a 10.5%-14% Criterion precast gel (Bio-Rad) for protein size separation at 100 V.
Staining, imaging, spot selection and protein identification
After size separation, gels were stained with SYPRO® Ruby Protein Stain (Bio-Rad), imaged using the Bio-Rad Gel Chemidoc™ XRS Gel Documentation System and analyzed for matchsets using the Bio-Rad PD Quest 2-D Image Analysis Software . The matchsets were used to select and excise protein spots which were sent to The Ohio State University Proteomics Facility to be digested with trypsin and analyzed by capillary-liquid chromatography-nanospray tandem mass spectrometry (Nano-LC/MS/MS). The sequences of the peptide fragments were analyzed by the Mascot software package (Matrix Science Inc., Boston, MA) to determine the potential identity of the proteins in each excised spot .
Cells were grown and harvested as in Fig 2. RNA was extracted using Bio-Rad's Aurum™ Total RNA Mini Kit, and cDNA from 0.5 μg of purified RNA was synthesized using Bio-Rad's iScript™ Select cDNA Synthesis Kit. PCR reactions containing 1.5 μl of cDNA, GoTaq® Green Master Mix (Promega, Madison, WI) and primers for cysK (5'-CTCGCTGACTATCGGTCA-3' and 5'-GATACCCGCAAGAATACC-3'), cysA (5'-ATGAGCATTGAGATTGCC-3' and 5'-ACGACTAATTGGGTGTAG-3'), cysT (5'-GCTGTTTGTGTGCCTGAT-3' and 5'-CGACTTTGCAGAGTGTTA-3'), cysW (5'-CGTGCAGGCGTTCAGCAA-3' and 5'-CCTGTTGCGCGCGTTTTT-3') or cysE (5'-ATGCCGTGTGAAGAACTG-3' and 5'-CTCGAAGGTATGGTGAAT-3') were performed for 30 cycles of 95°C for 1 minute, 50°C for 1 minute and 72°C for 1 minute. Equal volumes of the resulting PCR reactions were analyzed using a 1% agarose (Amresco) gel.
This study was funded by the National Science Foundation through grant number 0542178 and the University Research Council at the Youngstown State University (YSU) School of Graduate Studies. We thank Dr. Kari Green-Church of the Ohio State University for analyzing our spot samples. We thank Dr. David Asch and Dr. Gary Walker from YSU for critiquing the manuscript.
- Burk RF: Molecular biology of selenium with implications for its metabolism. FASEB J 1991, 5: 2274–2279.PubMedGoogle Scholar
- Foster CB: Selenoproteins and the metabolic features of the archaeal ancestor of eukaryotes. Mol Biol Evol 2004, 22: 383–386. 10.1093/molbev/msi007PubMedView ArticleGoogle Scholar
- Heider J, Bock A: Selenium metabolism in micro-organisms. Adv Microb Physiol 1993, 35: 71–109. full_textPubMedView ArticleGoogle Scholar
- Stadtman TC: Selenocysteine. Annu Rev Biochem 1996, 65: 83–100. 10.1146/annurev.bi.65.070196.000503PubMedView ArticleGoogle Scholar
- Zannoni D, Borsetti F, Harrison JJ, Turner RJ: The bacterial response to the chalcogen metalloids Se and Te. Adv Microb Physiol 2008, 53: 1–71. full_textPubMedView ArticleGoogle Scholar
- Forstrom JW, Zakowski JJ, Tappel AL: Identification of the catalytic site of rat liver glutathione peroxidase as selenocysteine. Biochemistry (N.Y.) 1978, 17: 2639–2644. 10.1021/bi00606a028View ArticleGoogle Scholar
- Zinoni F, Birkmann A, Stadtman TC, Bock A: Nucleotide sequence and expression of the selenocysteine-containing polypeptide of formate dehydrogenase (formate-hydrogen-lyase-linked) from Escherichia coli . Proc Natl Acad Sci USA 1986, 83: 4650–4654. 10.1073/pnas.83.13.4650PubMed CentralPubMedView ArticleGoogle Scholar
- Turner RJ, Weiner JH, Taylor DE: Selenium metabolism in Escherichia coli . Biometals 1998, 11: 223–227. 10.1023/A:1009290213301PubMedView ArticleGoogle Scholar
- Takahata M, Tamura T, Abe K, Mihara H, Kurokawa S, Yamamoto Y, Nakano R, Esaki N, Inagaki K: Selenite assimilation into formate dehydrogenase H depends on thioredoxin reductase in Escherichia coli . J Biochem 2008, 143: 467–473. 10.1093/jb/mvm247PubMedView ArticleGoogle Scholar
- Kim IY, Veres Z, Stadtman TC: Escherichia coli mutant SELD enzymes. The cysteine 17 residue is essential for selenophosphate formation from ATP and selenide. J Biol Chem 1992, 267: 19650–19664.PubMedGoogle Scholar
- Leinfelder W, Forchhammer K, Veprek B, Zehelein E, Bock A: In vitro synthesis of selenocysteinyl-tRNA(UCA) from seryl-tRNA(UCA): involvement and characterization of the selD gene product. Proc Natl Acad Sci USA 1990, 87: 543–557. 10.1073/pnas.87.2.543PubMed CentralPubMedView ArticleGoogle Scholar
- Forchhammer K, Bock A: Selenocysteine synthase from Escherichia coli . Analysis of the reaction sequence. J Biol Chem 1991, 266: 6324–6338.PubMedGoogle Scholar
- Leinfelder W, Zehelein E, Mandrand-Berthelot MA, Bock A: Gene for a novel tRNA species that accepts L-serine and cotranslationally inserts selenocysteine. Nature 1988, 331: 723–725. 10.1038/331723a0PubMedView ArticleGoogle Scholar
- Li C, Reches M, Engelberg-Kulka H: The bulged nucleotide in the Escherichia coli minimal selenocysteine insertion sequence participates in interaction with SelB: a genetic approach. J Bacteriol 2000, 182: 6302–6307. 10.1128/JB.182.22.6302-6307.2000PubMed CentralPubMedView ArticleGoogle Scholar
- Sandman KE, Tardiff DF, Neely LA, Noren CJ: Revised Escherichia coli selenocysteine insertion requirements determined by in vivo screening of combinatorial libraries of SECIS variants. Nucleic Acids Res 2003, 31: 2234–241. 10.1093/nar/gkg304PubMed CentralPubMedView ArticleGoogle Scholar
- Terada A, Yoshida M, Seko Y, Kobayashi T, Yoshida K, Nakada M, Nakada K, Echizen H, Ogata H, Rikihisa T: Active oxygen species generation and cellular damage by additives of parenteral preparations: selenium and sulfhydryl compounds. Nutrition 1999, 15: 651–655. 10.1016/S0899-9007(99)00119-7PubMedView ArticleGoogle Scholar
- Bebien M, Lagniel G, Garin J, Touati D, Vermeglio A, Labarre J: Involvement of superoxide dismutases in the response of Escherichia coli to selenium oxides. J Bacteriol 2002, 184: 1556–1564. 10.1128/JB.184.6.1556-1564.2002PubMed CentralPubMedView ArticleGoogle Scholar
- Seko Y, Imura N: Active oxygen generation as a possible mechanism of selenium toxicity. Biomed Environ Sci 1997, 10: 333–339.PubMedGoogle Scholar
- Spallholz JE, Hoffman DJ: Selenium toxicity: cause and effects in aquatic birds. Aquat Toxicol 2002, 57: 27–37. 10.1016/S0166-445X(01)00268-5PubMedView ArticleGoogle Scholar
- Shamberger RJ: The genotoxicity of selenium. Mutat Res 1985, 154: 29–48.PubMedView ArticleGoogle Scholar
- Dungan RS, Yates SR, Frankenberger WT Jr: Transformations of selenate and selenite by Stenotrophomonas maltophilia isolated from a seleniferous agricultural drainage pond sediment. Environ Microbiol 2003, 5: 287–295. 10.1046/j.1462-2920.2003.00410.xPubMedView ArticleGoogle Scholar
- Mergeay M, Nies D, Schlegel HG, Gerits J, Charles P, Van Gijsegem F: Alcaligenes eutrophus CH34 is a facultative chemolithotroph with plasmid-bound resistance to heavy metals. J Bacteriol 1985, 162: 328–334.PubMed CentralPubMedGoogle Scholar
- Roux M, Sarret G, Pignot-Paintrand I, Fontecave M, Coves J: Mobilization of selenite by Ralstonia metallidurans CH34. Appl Environ Microbiol 2001, 67: 769–773. 10.1128/AEM.67.2.769-773.2001PubMed CentralPubMedView ArticleGoogle Scholar
- Sarret G, Avoscan L, Carriere M, Collins R, Geoffroy N, Carrot F, Coves J, Gouget B: Chemical forms of selenium in the metal-resistant bacterium Ralstonia metallidurans CH34 exposed to selenite and selenate. Appl Environ Microbiol 2005, 71: 2331–2337. 10.1128/AEM.71.5.2331-2337.2005PubMed CentralPubMedView ArticleGoogle Scholar
- Ledgham F, Quest B, Vallaeys T, Mergeay M, Coves J: A probable link between the DedA protein and resistance to selenite. Res Microbiol 2005, 156: 367–374. 10.1016/j.resmic.2004.11.003PubMedView ArticleGoogle Scholar
- Bebien M, Kirsch J, Mejean V, Vermeglio A: Involvement of a putative molybdenum enzyme in the reduction of selenate by Escherichia coli . Microbiology 2002, 148: 3865–3872.PubMedGoogle Scholar
- Ranjard L, Nazaret S, Cournoyer B: Freshwater bacteria can methylate selenium through the thiopurine methyltransferase pathway. Appl Environ Microbiol 2003, 69: 3784–3790. 10.1128/AEM.69.7.3784-3790.2003PubMed CentralPubMedView ArticleGoogle Scholar
- Ranjard L, Prigent-Combaret C, Nazaret S, Cournoyer B: Methylation of inorganic and organic selenium by the bacterial thiopurine methyltransferase. J Bacteriol 2002, 184: 3146–3149. 10.1128/JB.184.11.3146-3149.2002PubMed CentralPubMedView ArticleGoogle Scholar
- Lindblow-Kull C, Kull FJ, Shrift A: Single transporter for sulfate, selenate, and selenite in Escherichia coli K-12. J Bacteriol 1985, 163: 1267–1269.PubMed CentralPubMedGoogle Scholar
- Muller S, Heider J, Bock A: The path of unspecific incorporation of selenium in Escherichia coli . Arch Microbiol 1997, 168: 421–427. 10.1007/s002030050517PubMedView ArticleGoogle Scholar
- Lacourciere GM, Levine RL, Stadtman TC: Direct detection of potential selenium delivery proteins by using an Escherichia coli strain unable to incorporate selenium from selenite into proteins. Proc Natl Acad Sci USA 2002, 99: 9150–9153. 10.1073/pnas.142291199PubMed CentralPubMedView ArticleGoogle Scholar
- Holmes A, Vinayak A, Benton C, Esbenshade A, Heinselman C, Frankland D, Kulkarni S, Kurtanich A, Caguiat J: Comparison of two multimetal resistant bacterial strains: Enterobacter sp. YSU and Stenotrophomonas maltophilia ORO2. Curr Microbiol 2009.Google Scholar
- Perkins DN, Pappin DJ, Creasy DM, Cottrell JS: Probability-based protein identification by searching sequence databases using mass spectrometry data. Electrophoresis 1999, 20: 3551–3567. 10.1002/(SICI)1522-2683(19991201)20:18<3551::AID-ELPS3551>3.0.CO;2-2PubMedView ArticleGoogle Scholar
- Tarr PI, Bilge SS, Vary JC Jr, Jelacic S, Habeeb RL, Ward TR, Baylor MR, Besser TE: Iha: a novel Escherichia coli O157:H7 adherence-conferring molecule encoded on a recently acquired chromosomal island of conserved structure. Infect Immun 2000, 68: 1400–1407. 10.1128/IAI.68.3.1400-1407.2000PubMed CentralPubMedView ArticleGoogle Scholar
- Kredich NM: Biosynthesis of cysteine. In Escherichia coli and Salmonella Cellular and Molecular Biology. ASM Press, Washington, D.C.; 1996:514–527.Google Scholar
- Emsley J: The Elements. Second edition. Oxford University Press, Oxford, England; 1995.Google Scholar
- Chen R, Schmidmayr W, Kramer C, Chen-Schmeisser U, Henning U: Primary structure of major outer membrane protein II (OmpA protein) of Escherichia coli K-12. Proc Natl Acad Sci USA 1980, 77: 4592–4596. 10.1073/pnas.77.8.4592PubMed CentralPubMedView ArticleGoogle Scholar
- Yanke LJ, Bryant RD, Laishley EJ: Hydrogenase I of Clostridium pasteurianum functions as a novel selenite reductase. Anaerobe 1995, 1: 61–67. 10.1016/S1075-9964(95)80457-9PubMedView ArticleGoogle Scholar
- Dungan RS, Frankenberger WT Jr: Reduction of selenite to elemental selenium by Enterobacter cloacae SLD1a-1. J Environ Qual 1998, 27: 1301–1306.View ArticleGoogle Scholar
- Ausubel F, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K: Short Protocols in Molecular Biology. Third edition. John Wiley & Sons, Inc, New York, NY; 1997.Google Scholar
- Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 1976, 72: 248–254. 10.1016/0003-2697(76)90527-3PubMedView ArticleGoogle Scholar
- Chandler JM, Treece ER, Trenary HR, Brenneman JL, Flickner TJ, Frommelt JL, Oo ZM, Patterson MM, Rundle WT, Valle OV, Kim TD, Walker GR, Cooper CR Jr: Protein profiling of the dimorphic, pathogenic fungus, Penicillium marneffei . Proteome Sci 2008, 6: 17. 10.1186/1477-5956-6-17PubMed CentralPubMedView ArticleGoogle Scholar
- O'Farrell PH: High resolution two-dimensional electrophoresis of proteins. J Biol Chem 1975, 250: 4007–4021.PubMed CentralPubMedGoogle Scholar
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