bluuebunny/crossref_metadata_2025_split huggingface dataset:
Parallel induction strategies for cat‐86: separating chloramphenicol induction from protein synthesis inhibition
Authors: Rogers Elizabeth J. , Ambulos Nicholas P. , Gu Zhiping , Lovett Paul S. 
Published: October 2006

SummaryInduction of cat‐86 translation results from the stalling of a ribosome at a discrete location in the leader region of the transcript. Stalling destabilizes an adjacent region of secondary structure that sequesters the cat‐86 ribosome binding site, thereby activating cat‐86 translation. Two well characterized antibiotics, chloramphenicol and erythromycin, induce cat‐86 by stalling a ribosome at the appropriate leader site. Here we demonstrate differences between the two antibiotics with respect to induction. First, induction by chloramphenicol is dependent on nucleotides in the leader sequence that are different from those necessary for erythromycin induction. Second, variants of Bacillus subtilis that are chloramphenicol resistant because of chromosome mutations permit cat‐86 induction by chloramphenicol, whereas erythromycin‐resistance host mutations block or greatly reduce cat‐86 induction by erythromycin. Third, selected strains of B. subtilis bearing alterations in proteins of the 50S ribosomal subunit interfere with cat‐86 induction by chloramphenicol, yet these strains are chloramphenicol sensitive. Lastly, induction by chloramphenicol is not reversed by removal of the antibiotic whereas erythromycin induction is reversible. The data indicate that chloramphenicol induction results from an effect of the drug that is not identical to its role as a general inhibitor of ribosome elongation. Induction by erythromycin, on the other hand, could not be distinguished from its antibiotic activity.
The presence of a novel type of surface polysaccharide in Rhizobium meliloti requires a new fatty acid synthase‐like gene cluster involved in symbiotic nodule development
Authors: Petrovics György , Putnoky Peter , Reuhs Bradley , Kim John , Thorp Tina A. , Noel K. Dale , Carlson Russell W. , Kondorosi Adam 
Published: October 2006

SummaryBacterial exopolysaccharide (EPS) and lipopolysaccharide (LPS) molecules have been shown to play important roles in plant‐bacterium interactions. Here we have demonstrated that the fix‐23 loci, which compensate for exo mutations during symbiotic nodule development, are involved in the production of a novel polysaccharide that is rich in 3‐deoxy‐D manno‐2‐octulosonic acid (Kdo) but is not the classical LPS. This molecule is likely to be a surface antigen since antiserum to whole Rhizobium meliloti cells reacts strongly with it, and since mutations in fix‐23 result in an inability to produce this polysaccharide and to bind bacteriophage 16‐3. It is likely that this Kdo‐rich polysaccharide is analogous to certain Escherichia coli K‐antigens which are anchored to the membrane via a phospholipid moiety. DNA sequence analysis of one gene cluster of this region revealed that the predicted protein products of six genes exhibit a high degree of homology and similar organization to those of the rat fatty acid synthase multifunctional enzyme domains.
Expression of the ferric enterobactin receptor (PfeA) of Pseudomonas aeruginosa: involvement of a two‐component regulatory system
Authors: Dean Charles R. , Poole Keith 
Published: October 2006

SummaryExpression of the ferric enterobactin receptor in Pseudomonas aeruginosa is inducible by enterobactin and requires sequences upstream of the structural gene (pfeA). Nucleotide sequencing of a 2.5 kilo‐base pair (kb) region of DNA immediately upstream of pfeA revealed two open reading frames (ORFs), pfeR and pfeS, which appeared to comprise an operon. The predicted products of pfeR and pfeS (molecular weight 26 796 and 50 597, respectively) exhibited a high degree of homology to response‐regulator and sensor components, respectively, of the superfamily of prokaryotic environmentally responsive protein pairs. Consistent with an apparent role in regulating expression of pfeA in response to enterobactin, introduction of pfeR/pfeS into P. aeruginosa on a high‐copy‐number vector enhanced enterobactin‐dependent expression of pfeA. Furthermore, a pfeR mutant obtained by in vitro mutagenesis and gene replacement failed to express PfeA despite the presence of enterobactin in the culture medium. Analysis of the hydropathy profiles of PfeR and PfeS supported a cytoplasmic location for PfeR and a cytoplasmic membrane location for PfeS.
Isolation and analysis of a linear plasmid‐located gene of Borrelia burgdorferi B29 encoding a 27 kDa surface lipoprotein (P27) and its overexpression in Escherichia coli
Authors: Reindl Markus , Redl Bernhard , Stöffler Georg 
Published: October 2006

SummaryUsing an antiserum of a patient with cutaneous manifestations of Lyme borreliosis we have isolated the gene encoding a 27 kDa protein antigen (P27) of Borrelia burgdorferi B29 from a λ‐gt11 expression library. Nucleotide sequence analysis revealed that it is a basic protein of 248 amino acids with a typical prokaryotic leader sequence of 17 amino acid residues at the N‐terminus of the proposed translation product. Biochemical investigations showed that P27 is a surface‐exposed lipoprotein. From pulsed‐field gel electrophoresis and subsequent Southern blot analysis it is evident that the p27 gene is located on a linear plasmid of a size of approximately 55 kb. It was over‐expressed in Escherichia coli and the purified recombinant protein was used for biochemical and serological studies. Northern and Western blot analysis demonstrated that p27 is expressed in the European B. burgdorferi strain B29, but not in the American strain B31.
Preparation and analysis of isogenic mutants in the transferrin receptor protein genes, tbpA and tbpB, from Neisseria meningitidis
Authors: Irwin Sean W. , Averil Nuzhat , Cheng Cindy Y. , Schryvers Anthony B. 
Published: October 2006

SummaryIsogenic mutants were constructed in the tbpA and tbpB genes from Neisseria meningitidis strain B16B6, which code for the transferrin receptor proteins, Tbp1 and Tbp2. Insertion mutants of the tbpA and tbpB genes were obtained by shuttle mutagenesis and by in vitro cassette mutagenesis, respectively. The Isogenic mutants were verified by Southern blot and Western blot analysis. Isogenic mutants deficient in Tbp1 or Tbp2 demonstrated a reduced transferrin binding activity in intact cells and total membranes but were incapable of utilizing transferrin iron for growth. Tbp1 could be isolated by affinity methods from the mutant lacking Tbp2 but isolation of Tbp2 from the mutant lacking Tbp1 required the presence of exogenous Tbp1.
The region of human transferrin involved in binding to bacterial transferrin receptors is iocalized in the C‐lobe
Authors: Alcantara J. , Yu R. H. , Schryvers A. B. 
Published: October 2006

SummaryIron‐saturated human transferrin was digested with either chymotrypsin or trypsin to produce C‐lobe and N‐lobe protein fragments. Individual protein fragments were purified by a combination of gel filtration and Concanavalin A affinity chromatographic procedures. The C‐lobe and N‐lobe fragments of human transferrin were then used in binding assays to assess their ability in binding to the bacterial transferrin receptors. Competitive binding assays demonstrated that the C‐lobe fragment of human transferrin binds as well as intact human transferrin to bacterial transterrin receptors from Neisseria meningitidis, Neisseria gonorrhoeae and Haemophlius influenzae. Using isogenic mutants of N. meningitidis deficient in either of the transferrin‐binding proteins (Tbps), we demonstrated that both transferrin‐binding proteins were able to bind to the C‐lobe fragment of human transferrin.
Identification of an operon involved in the assimilatory nitrate‐reducing system of Azotobacter vineiandii
Authors: Ramos Francisco , Blanco Gonzalo , Gutiérrez Juan C. , Luque Francisco , Tortolero Maria 
Published: October 2006

SummaryA number of mutants lacking nitrate reductase (Nas−) or nitrite reductase (Nis−) activities have been isolated and characterized. An operon including two new genes (nasA and nasB) has been defined and cloned from an Azotobacter vinelandii gene bank. nasA encodes for nitrite reductase apoenzyme, whereas nasB is specific for nitrate reductase activity. Nitrate reductase exerts a regulatory effect on nasAB.
Lactococcus lactis: high‐level expression of tetanus toxin fragment C and protection against lethal challenge
Authors: Wells Jeremy M. , Wilson Peter W. , Norton Pamela M. , Gasson Michael J. , Le Page Richard W. F. 
Published: October 2006

SummaryTo determine if the food‐grade bacterium Lactococcus lactis holds promise as a vaccine antigen delivery vector we have investigated whether this bacterium can be made to produce high levels of a heterologous protein antigen. A regulated expression system has been developed which may be generally suitable for the expression of foreign antigens (and other proteins) In L. lactis. The system utilizes the fast‐acting T7 RNA polymerase to transcribe target genes, and provides the first example of the successful use of this polymerase in a Gram‐positive bacterium. When the performance of the expression system was characterized using tetanus toxin fragment C (TTFC) up to 22% of soluble cell protein was routinely obtained as TTFC. Mice immunized subcutaneously with L. lactis expressing TTFC were protected from lethal challenge with tetanus toxin. These results show for the first time that L. lactis is able to express substantial quantities of a heterologous protein antigen and that this organism can present this antigen to the Immune system in an immunogenic form.
Yeast DNA recombination and repair proteins Rad 1 and Radio constitute a complex in vivo mediated by localized hydrophobic domains
Authors: Bardwell A. Jane , Bardwell Lee , Johnson Douglas K. , Friedberg Errol C. 
Published: October 2006

SummaryThe Saccharomyces cerevisiae Rad 1 and Rad 10 proteins are required for damage‐specific incision during nucleotide excision repair and also for certain mitotic recombination events between repeated sequences. Previously we have demonstrated that Rad1 and Rad10 form a specific complex in vitro. Using the ‘two‐hybrid’ genetic assay system we now report that Rad1 and Rad10 proteins are subunits of a specific complex in the cell nucleus. The Rad10‐binding domain of Rad1 protein maps to a localized region between amino acids 809–997. The Rad1 ‐binding domain of Radio protein maps between amino acids 90–210. These domains are evolutionarily conserved and are hydrophobic in character. Although significant homology exists between Rad10 and the human‐DNA‐repair protein Ercc1 in this region, we were unable to detect any interaction between Ercc1 and Rad1 proteins. We conclude that Rad1 and Rad10 operate in DNA repair and mitotic recombination as a constitutive complex.
Regulation of epidermin biosynthetic genes by EpiQ
Authors: Peschel Andreas , Augustin Johannes , Kupke Thomas , Stevanovic Stefan , Götz Friedrich 
Published: October 2006

SummaryWe investigated the role of epiQ in the biosynthesis of the lantibiotic epidermin. epiQ was essential for epidermin production. It was shown that EpiQ controls epidermin production by transcriptionally activating the epiA promoter, used for transcription of most of the epidermin biosynthetic genes. Additional copies of epiQ increased epidermin production in the epidermin‐producing wild‐type strain Staphylococcus epidermidis Tü3298. The epiA promoter region was characterized by primer extension analysis. Two inverted repeats, putative operator sites for EpiQ binding, are located upstream of the −35 region and one is localized downstream of the‐10 region. Crude protein extracts from S. epidermidis Tü3298 and epiQ expressing Escherichia coli cells led to gel mobility shifts of a DNA fragment bearing the inverted repeat which is located immediately upstream of the ‐35 region. DNA fragments bearing the other two inverted repeats were not shifted. The epiQ gene product could be detected by overexpression in the E. coli T7 system using antiserum raised against synthetic pep‐tides of EpiQ. Furthermore, EpiQ, like other DNA‐binding proteins, was shown to bind strongly to heparin sepharose.
Site‐specific insertion of gene cassettes into integrons
Authors: Collis Christina M. , Grammaticopoulos Georgia , Briton Jayne , Stokes H.W. , Hall Ruth M. 
Published: October 2006

SummarySite‐specific insertion of gene cassettes into the insert region of integrons has been demonstrated. Insertion was only observed if the integron DNA integrase was expressed in the recipient cell and if the cassette DNA was ligated prior to transformation. The essential ligation products were resistant to treatment with exonuclease III, indicating that they were closed circular molecules. Insertion of cassettes into integron fragments containing either no insert (one recombination site), or one gene cassette (two recombination sites), was demonstrated. In the latter case, insertion occurred predominantly at the core site located 5′ to the resident cassette, which corresponds to the only site available when no insert is present in the recipient. When DNA molecules including two gene cassettes were used, insertion of only one of the gene cassettes was generally observed, suggesting that resolution of the circular molecule to generate two independent circular cassettes occurred more rapidly than insertion into the recipient integron.
Iron and oxygen regulation of Escherichia coli MnSOD expression: competition between the global regulators Fur and ArcA for binding to DNA
Authors: Tardat Brigitte , Touati Danièle 
Published: October 2006

SummaryManganese superoxide dismutase (the sod A gene product) in Escherichia coli, is negatively regulated by two global regulators, ArcA (aerobic respiration control) and Fur (ferric uptake regulation), coupling its expression to aerobic metabolism and the intracellular iron pool. Footprinting analyses were carried out on the sodA promoter region with purified Fur protein and with ArcA protein overproduced in crude extracts. ArcA is able to bind in vitro in the absence of the in vivo triggering signal. The binding occurs in one step and study of contacts within the operator sequence reveals binding on one side of the double helix. The DNA protection extends to a much larger region (about 65 bp) than would be expected for a 27 kDa protein, suggesting polymerization. Both Fur and ArcA footprints encompass the –35 and –10 promoter region and there is considerable overlap of their binding sequences, in agreement with in vivo results suggesting that either regulator alone can block sodA transcription. Furthermore, competition experiments show that Fur and ArcA binding to the sodA promoter are mutually exclusive and that ArcA can easily displace Fur, but not vice versa. The biological significance of this in vitro behaviour is discussed.
Spatial expression and autoregulation of hetR, a gene involved in the control of heterocyst development in Anabaena
Authors: Black Todd A. , Cai Yuping , Wolk C. Peter 
Published: October 2006

SummaryThe spatially patterned differentiation of hetero‐cysts in the filamentous cyanobacterium Anabaena requires a functional hetR gene. Transcriptional fusions to luxAB show that hetR is transcribed at a low level throughout the filament when Anabaena is grown with combined nitrogen, and that induction of the gene begins within 2 h following nitrogen deprivation. By 3.5 h, induction is localized to spaced foci. By 6h, there is an overall induction of at least threefold in whole cultures, reflecting at least a 20‐fold increase within spatially separated cells. The induction requires the presence of a functional hetR gene, indicating that hetR is autoregulatory. Full induction of a heterocyst structural gene, hepA, also requires a functional hetR locus.
comF, a Bacillus subtilis late competence locus, encodes a protein similar to ATP‐dependent RNA/DNA helicases
Authors: Londoño‐Vallejo J. A. , Dubnau D. 
Published: October 2006

SummaryWe have sequenced and genetically characterized comF, a Bacillus subtilis competence locus, previously identified by Tn917 transposon insertion mutagenesis. Expression of the locus, in which three open reading frames (ORFs) were found, is driven by a single σ;A‐like promoter in front of comFORF1 and is dependent on early regulatory competence genes and only expressed in competence medium. The predicted amino acid sequences of two of the ORFs showed similarities to known proteins in the Gen‐Bank and SwissProt databases: ComFORF1 is similar to an extensive family of ATP‐dependent RNA/DNA helicases with closer similarity to the DEAD protein subfamily and to the PriA protein in Escherichia coli. The latter is a DNA translocase/helicase required for primosome assembly at the replication fork of phage φX174. ComFORF3 is 22% identical to Com101, a protein required for genetic competence in Haemophilus influenzae, a naturally competent Gram‐negative bacterium. In‐frame comFORF1 deletions were 1000‐fold deficient in transformability compared to the wild type, whereas disruptions of the other two ORFs were only five‐ to 10‐fold lower. These observations allow us to hypothesize that the ComFORF1 late gene product plays an essential role during the binding and uptake events involved in Bacillus subtilis transformation.
Identification of nodSUIJ genes in Nod locus 1 of Azorhizobium caulinodans: evidence that nodS encodes a methyltransferase involved in Nod factor modification
Authors: Geelen Danny , Mergaert Peter , Geremia Roberto A. , Goormachtig Sofie , Van Montagu Marc , Holsters Marcelle 
Published: October 2006

SummaryThe Azorhizobium caulinodans strain ORS571 nodu‐lation genes nodSUIJ were located downstream from nodABC. Complementation data and transcriptional analysis suggest that nodABCSUIJ form a single operon. Mutants with Tn5 insertions in the genes nodS, nodU, and nodJ were delayed in nodulation of Sesbania rostrata roots and stems. The NodS amino acid sequences of ORS571, Bradyrhizobium japonicum, and Rhizobium sp. strain NGR234, contain a consensus with similarity to 5‐adenosylmethionine (SAM)‐utilizing methyltransferases. A naringenin‐inducible nodS‐dependent protein of approximately 25kDa could be cross‐linked to radiolabelled SAM. By applying L‐[methyl‐3H]‐methionine in vivo. Nod factors of ORS571, known to be N‐methylated, could be labelled in wild type and nodU mutants but not in nodS mutants. Therefore, we propose that NodS is a SAM‐utilizing methyltransferase involved in Nod factor synthesis.
Co‐dependent positive regulation of the ansBF promoter of Escherichia coli by CRP and the FNR protein: a molecular analysis
Authors: Jennings Michael P. , Beacham Ifor R. 
Published: October 2006

SummaryTranscription of the ansB gene, encoding l‐asparaginase II, is positively regulated by cAMP receptor protein (CRP) and by the product of the fnr gene, the FNR protein. These global regulatory proteins mediate the expression of ansB in Escherichia coli in response to carbon source and to anaerobiosis, respectively, and are required concurrently for optimal ansB expression. The mechanism whereby CRP and FNR interact co‐operatively with the ansB promoter to achieve transcription has not previously been established. We have utilized an ansB‘‐’1lacZ fusion, in conjunction with deletion analysis and site‐directed mutagenesis, to identify two sites which interact with these regulatory proteins In the ansB promoter. The first is an FNR site, centred 41.5 bp upstream of the major transcriptional start site. The second site, located 28 bp upstream of the FNR site, is the site of CRP regulation. This site is homologous to both the CRP and FNR binding‐site consensus sequences and may respond to both CRP and FNR. The concurrent requirement for CRP and FNR for optimal expression of ansB may be explained if, first, essentially no transcription occurs unless the FNR is bound at the downstream site, and, second, the level of transcription when FNR alone is present is enhanced when CRP binds at the upstream site.
Cloning and characterization of the psaE gene of the cyanobacterium Synechococcus sp. PCC 7002: characterization of a psaE mutant and overproduction of the protein in Escherichia coli
Authors: Zhao J. , Snyder W. B. , Mühlenhoff U. , Rhiel E. , Warren P. V. , Golbeck J. H. , Bryant D. A. 
Published: October 2006

SummaryThe psaE gene, encoding a 7.5 kDa peripheral protein of the photosystem I complex, has been cloned and characterized from the cyanobacterium Synechococcus sp. PCC 7002. The gene is transcribed as an abundant monocistronic transcript of approximately 325 nt. The PsaE protein has been overproduced in Escherichia coli, purified to homogeneity, and used to raise polyclonal antibodies. Mutant strains, in which the psaE gene was insertionally inactivated by interposon mutagenesis, were constructed and characterized. Although the PS I complexes of these strains were similar to those of the wild type, the strains grew more slowly under conditions which favour cyclic electron transport and could not grow at all under photoheterotrophic conditions. The results suggest that PsaE plays a role in cyclic electron transport in cyanobacteria.
Molecular analysis of two ScrR repressors and of a ScrR–FruR hybrid repressor for sucrose and D‐fructose specific regulons from enteric bacteria
Authors: Jahrels K. , Lengeler J. W. 
Published: October 2006

SummaryThe scr regulon of pUR400 and the chromosomally encoded scr regulon of Klebsiella pneumoniae KAY2026 are both negatively controlled by a specific repressor (ScrR). As deduced from the nucleotide sequences, both scrR genes encode polypeptides of 334 residues (85.5% identical base pairs, 91.3% identical amino acids), containing an N‐terminal helix‐turn‐helix motif. Comparison with other regulatory proteins revealed 30.6% identical amino acids to FruR, 27.0% to Lacl and 28.1% to GaIR. Six scrRs super‐repressor mutations define the inducer‐binding domain. The scr operator sequences were identified by in vivo titration tests of the sucrose repressor and by in vitro electrophoretic mobility shift assays. D‐fructose, an intracellular product of sucrose transport and hydrolysis, and D‐fructose 1‐phosphate were shown to be molecular inducers of both scr regulons. An active ScrR–FruR hybrid repressor protein was constructed with the N‐terminal part of the sucrose repressor of K. pneumoniae and the C‐terminal part of the fructose repressor of Salmonella typhimurium, LT2. Gel retardation assays showed that the hybrid protein bound to scr‐specific operators, and that D‐fructose 1‐phosphate, the inducer for FruR, was the only inducer. In vivo, neither the operators of the fru operon nor of the pps, operon, the natural targets for FruR, were recognized, but the scr operators were. These data and the data obtained from the super‐repressor alleles confirm previous models on the binding of repressors of the Lacl family to their operators.
A family of IS1031 elements in the genome of Acetobacter xylinum: nucleotide sequences and strain distribution
Authors: Coucheron Dag H. 
Published: October 2006

SummaryAn insertion sequence (here called IS 1031A) from Acetobacter xylinum ATCC 23769 has recently been isolated. This study describes the complete nucleotide sequence of IS 1031A as well as the sequences of two novel iso‐IS 1031 elements, IS1031C and IS1031D, from A. xylinum ATCC 23769. The three ISs are all exactly 930 bp long, have imperfect terminal inverted repeats of 24 bp for IS1031A and 21 bp for IS1031C and IS1031D, are flanked by three base pair direct repeats, and contain an open reading frame encoding a putative basic protein of 278 amino acids. Because of nucleotide substitutions, IS1031C and IS1031D differ from IS 1031A by 12.9% while IS1031C differs from IS1031D by only 0.6%. Hybridization analyses of total DNA from nine A. xylinum strains showed that all strains contained IS 1031‐like elements varying in copy number from three to at least 16. None of three Acetobacter aceti strains examined contained IS1031‐like elements. Taken together, the results suggest that A. xylinum contains a family of IS 1031 elements with considerably diversified nucleotide sequences.