bluuebunny/crossref_metadata_2025_split huggingface dataset:
Replication mutants of Staphylococcus aureus macrolide‐lincosamide‐streptogramin B resistance plasmid pT48
Authors: Catchpole I. , Dyke K. G. H. 
Published: October 2006

SummaryCopy‐number mutants of Staphylococcus aureus macrolide‐lincosamide‐streptogramin B (MLS) resistance plasmid pT48 were isolated by their resistance to the non‐inducing macrolide, tylosin. One mutant plasmid, pcopD3, showed a three‐ to five‐fold cis‐dominant increase in copy number, and nucleotide sequence analysis revealed that the mutant had a single base change within the replication region. All other pT48 mutants examined had the unusual pheno‐type of increased plasmid multimerization and elevated copy number. These mutants were effective in trans and DNA sequencing showed that plasmids with this phenotype were deleted in one of two ways. The deletions caused similar alterations to the C‐ter‐minus of the wild‐type pT48 Rep protein. The two types of mutant Rep proteins terminate with the same pentapeptide sequence: Ala‐Asn‐Glu‐lle‐Asp. The multimerization phenotype of these mutants can be explained by defective termination of rolling‐circle type replication.
The ArsR protein is a trans‐acting regulatory protein
Authors: Wu J. , Rosen B. P. 
Published: October 2006

SummaryThe arsR gene encodes the regulatory protein of the plasmid‐encoded arsenical resistance operon. A series of in‐frame fusions was constructed between the C‐terminally truncated arsR gene and the coding region for the mature form of β‐lactamase (blaM). Fusions containing most of the arsR gene were still inducible by arsenicals. Fusions containing less than 102 residues of the 117‐residue ArsR protein were constitutive. When a wild‐type arsR gene was placed in trans, the constitutive constructs were again inducible. The results demonstrate that the ArsR protein is a trans‐acting regulatory protein which controls its own expression.
Mitochondrial gene sequences show fungal homology for Pneumocystis carinii
Authors: Pixley F. J. , Wakefield A. E. , Banerji S. , Hopkin J. M. 
Published: October 2006

SummaryA 6.8 kjlobase fragment of mJtochondrial DNA from Pneumocystis carinii encodes for apocytochrome b, NADH dehydrogenase subunits 1, 2, 3, and 6, cytochrome oxidase subunit II, and the small subunit of ribosomal RNA. Comparative sequence analysis with a series of organisms representative of the fungal and protozoan groups shows that R carinii has, consistently, an average similarity of 60% with the fungi but only 20% with the protozoa. The data indicate homology with the fungi for this opportunistic pathogen.
Characterization of a polygalacturonase gene of Aspergillus niger RH5344
Authors: Ruttkowski E. , Khanh N. Q. , Wientjes F.‐J. , Gottschalk M. 
Published: October 2006

SummaryWe have cloned a gene encoding a polygalacturonase (PG) in the filamentous fungus Aspergillus niger RH5344. The structural gene comprises 1141 bp codIng for 362 amino acids and the open reading frame is disrupted by one intron of 52 bp. Eukaryotic consensus sequences for transcription regulation are found only in deviated forms. The biological functionality of the isolated PG gene was established by retransformation in A. niger and Aspergillus awamori. In addition, we have found that the PG protein of A. niger shares significant similarities with PG proteins from tomato and Erwinia carotovora. Comparison of the three enzymes revealed a highly conserved region in their C‐terminal region probably comprising the elements of substrate binding and the catalytic centre.
A chimeric nucleotide‐binding protein, encoded by a hisP–malK hybrid gene, is functional in maltose transport in Salmonella typhimurium
Authors: Schneider E. , Walter C. 
Published: October 2006

SummaryWe have isolated a hybrid gene, composed of the first 455 nucleotides of hisP and nucieotides 275–1107 of malK, the genes coding for the nucleotide‐binding components of the high‐affinity transport systems for histidine and maltose in Salmonella typhimurium, respectively. The fusion had occurred by recombination within 11 homologous base pairs located between the two DNA fragments. In the chimeric protein peptidic motifs A and B, proposed to be part of the nucleotide‐binding fold, originate from HisP and MalK, respectively. Plasmid pES42‐39, harbouring the hybrid gene, was shown to complement only a malK mutation but failed to complement a hisP deletion mutation. The chimeric protein was identified by immunoblotting as a protein with an apparent molecular mass of 49kDa. Removal of the C‐terminal 77 amino acid residues from the chimeric protein resulted in the loss of function in transport. In contrast, 51 amino acid residues could be removed from the C‐terminus of wild‐type MalK without any effect. Upon overproduction the chimeric protein, as wild‐type MalK, inhibited expression of the malB regulon. However, both truncated proteins, when overproduced, did not exhibit this activity. Based on these results, a tentative model of the functional domains of MalK is presented.
Organization of genes encoding membrane proteins of the Escherichia coli ferrienterobactin permease
Authors: Chenault S. S. , Earhart C. F. 
Published: October 2006

SummaryTransposon mutagenesis and plasmid complementation studies have identified two genes, fepD and fepG, which are essential for ferrienterobactin transport in Escherichia coli. These genes mapped In the enterobactin gene cluster and genetic evidence indicated that they are transcribed as part of an operon (fepD, fepG, fepC). The nucleotide sequence of fepD was determined; it could encode a hydrophobic 33.8kDa protein with sequence homologies to other iron and vitamin B12 transport proteins. Also identified, between fepD and fepB, was an open reading frame (ORF43) with no detectable function; its 43 kDa protein product (P43) was seen on polyacrylamide gels. The fepD–C operon and ORF43 were divergently transcribed from a 110bp region containing a binding site for the repressor protein Fur.
Nucleotide sequence and genetic organization of the ferric enterobactin transport system: homology to other peripiasmic binding protein‐dependent systems in Escherichia coli
Authors: Shea C. M. , McIntosh M. A. 
Published: October 2006

SummaryThe nucleotide sequence of the Escherichia coli fep genomic region has been determined. Three new loci were identified. One of these, P43, encodes a membrane protein that is not essential for ferric enterobactin transport. Two others, fepD and fepG, were found to be essential for transport and their translational products showed extensive homology to other integral membrane proteins involved in TonB‐dependent transport processes. The FepC amino acid sequence suggested a peripheral membrane location and revealed conserved ATP‐binding domains. Together these data indicate that ferric enterobactin is transported through a typical periplasmic binding protein‐dependent system, in addition, the transcriptional organization of these genes was examined and primer extension analysis identified a single iron‐regulated bidirectional promoter between the P43 gene and the fepDGC operon.
Pro‐peptide as an intermolecular chaperone: renaturation of denatured subtilisin E with a synthetic pro‐peptide
Authors: Ohta Y. , Hojo H. , Aimoto S. , Kobayashi T. , Zhu X. , Jordan F. , Inouye M. 
Published: October 2006

SummaryThe ammo‐terminal pro‐sequence consisting of 77 amino acid residues is required to guide the folding of secreted subtilisin E, a serine protease, into active, mature enzyme (Ikemura et al., 1987). Furthermore, denatured subtilisin E can be folded to active enzyme in an intermolecular process with the aid of an exogenously added pro‐subtilisin E, the active site of which was mutated (Zhu et al., 1989). In this report, we have synthesized the pro‐peptide of 77 residues (corresponding to ‐1 to ‐77 in the sequence, where residue +1 is the N‐terminal amino acid residue of the mature protein), and have found that it could intermolecularly complement the folding of denatured subtilisin E to active enzyme. Furthermore, we have found that the synthetic pro‐peptide exhibits specific strong binding to the active mature enzyme by inhibiting it competitively at its active centre with an upper limit to a Ki of 5.4 × 10−7. In contrast, synthetic pro‐peptides corresponding to ‐44 to ‐77, ‐1 to 64 and ‐1 to ‐43 inhibited the enzyme with Ki values weaker by two orders of magnitude. The results indicate that the sequence extending from ‐1 to ‐77 is essential for specificity of interaction, perhaps generating a conformation that accounts for both roles found hitherto, i.e. specific binding to the active centre, and guiding of the refolding to active enzyme. Thus these results suggest that the pro‐peptide functions as an intermolecular chaperone.
The role of pili in the interactions of pathogenic Neisseria with cultured human endothelial cells
Authors: Virji M. , Kayhty H. , Ferguson D. J. P. , Alexandrescu C. , Heckels J. E. , Moxon E. R. 
Published: October 2006

SummaryThe influence of the two surface structures of Neisseria meningitidis, capsule and pili, in bacterial interactions with human endothelial cells was investigated. Increased association correlated with the presence of pili on bacteria while capsule type had no apparent effect. Strains expressing both Class I and Class II pili associated with endothelial cells in significantly larger numbers compared with the non‐piliated variants of the same strains (>10×). Variants of Neisseria gonorrhoeae strain P9 expressing antigenically distinct pili also associated with endothelial cells in larger numbers (>30×) compared with the non‐piliated variant. Electron microscopic studies confirmed these data and showed that gonococci were internalized more frequently compared with meningococci. One consequence of increased association was an increase in the cytopathic effect of bacteria on the target cells.
Site‐specific insertion of genes into integrons: role of the 59‐base element and determination of the recombination cross‐over point
Authors: Hall R. M. , Brookes D. E. , Stokes H. W. 
Published: October 2006

SummaryFrom examination of published DNA sequences of genes found inserted at a specific site in integrons, all genes are shown to be associated, at their 3′ ends, with a short imperfect inverted repeat sequence, a 59‐base element or relative of this element. The similarity of the arrangement of gene inserts in the integron and in the Tn7 transposon family is described. A refined consensus for the 59‐base element is reported. Members of this family are highly diverged and the relationship of a group of longer elements to the 59‐base elements is demonstrated. The ability of 59‐base elements of different length and sequence to act as sites for recombination catalysed by the integron‐encoded DNA integrase is demonstrated, confirming that elements of this family have a common function. The ability of elements located between gene pairs to act as recombination sites has also been demonstrated. The recombination cross‐over point has been localized to the GTT triplet which is conserved in the core sites, GTTRRRY, found at the 3′ end of 59‐base elements. Recombination at the core site found in inverse orientation at the 5′ end of the 59‐base elements was not detected, and the sequences responsible for orientation of the recombination event appear to reside within the 59‐base element.A model for site‐specific insertion of genes into integrons and Tn 7‐iike transposons is proposed. Circular units consisting of a gene associated with a 59‐base element are inserted into an ancestral element which contains neither a gene nor a 59‐base element. Further genes can be inserted at one or both of the composite sites generated by the first gene‐insertion
The rifampicin‐inducible genes srn6 from F and pnd from R483 are regulated by antisense RNAs and mediate plasmid maintenance by kiiling of plasmid‐free segregants
Authors: Nielsen A. K. , Thorsted P. , Thisted T. , Wagner E. G. H. , Gerdes K. 
Published: October 2006

SummaryThe gene systems srnB of plasmid F and pnd of plasmid R483 were discovered because of their induction by rifampicin. Induction caused membrane damage, RNase I influx, degradation of stable RNA and, consequently, cell killing. We show here that the srnB and pnd systems mediate efficient stabilization of a mini‐R1 test‐plasmid. We also show that the killer genes srnB′ and pndA are regulated by antisense RNAs, and that the srnC‐ and pndB‐encoded antisense RNAs, denoted SrnC‐ and PndB‐RNAs, are unstable molecules of approximately 60 nucleotides. The srnB and pndA mRNAs were found to be very stable. The differential decay rates of the inhibitory antisense RNAs and the killer‐gene‐encoding mRNAs explain the induction of these gene systems by rifampicin. Furthermore, the observed plasmid‐stabilization phenotype associated with the srnB and pnd systems is a consequence of this differential RNA decay: the newborn plasmid‐free cells inherit the stable mRNAs, which, after decay of the unstable antisense RNAs, are translated Into killer proteins, thus leading to selective killing of the plasmid‐free segregants. Thus our observations lead us to conclude that the F srnB and R483 pnd systems are phenotypically indistinguishable from the R1 hok/sok system, despite a 50% dissimilarity at the level of DNA sequence.
Simultaneous expression of a bacteriophage Mu transposase and repressor: a way of preventing killing due to mini‐Mu replication
Authors: Toussaint A. , Expert D. , Desmet L. 
Published: October 2006

Summary In vitro studies of bacteriophage Mu transposition have shown that the phage‐encoded transposase and repressor bind the same sequences on the phage genome. We attempted to test that prediction in vivo and found that Mu repressor directly inhibits transposition. We also found that, in the absence of repressor, constitutive expression of Mu transposition functions pA and pB is lethal in Escherichia coli strains lysogenic for a mini‐Mu and that this is the result of intensive replication of the mini‐Mu. These findings have important consequences where such mini‐Mus are used as genetic tools. We also tested whether in Erwinia chrysanthemi the effect of transposition functions on a resident mini‐Mu was the same as in E. coli. We observed that expression of pA alone was lethal in E. chrysanthemi and that a large fraction of the survivors underwent precise excision of the mini‐Mu.
The molecular basis of carbon‐starvation‐induced general resistance in Escherichia coli
Authors: Matin A. 
Published: October 2006

SummaryAt the onset of starvation Escherichia coli undergoes a temporally ordered program of starvation gene expression involving 40–80 genes which some four hours later yields cells possessing an enhanced general resistance. Two classes of genes are induced upon carbon starvation: the cst genes, requiring cyclic AMP, and the pex genes, not requiring this nucleotide for induction. The cst genes are not involved in the development of the resistant state and are concerned with escape from starvation, while the pex gene induction appears to be associated with resistance. Many of the latter are induced in response to a variety of starvation conditions. They include heat shock and oxidation resistance genes, and some utilize minor, stationary‐phase‐specific sigma factors for induction during starvation. The protective role of stress proteins may be due to their ability to rescue misfolded macromolecules. The starvation promoters can be potentially useful for selective expression of desired genes in metabolically sluggish populations, e.g. in high‐density industrial fermentations and in situ bioremediation.
The regulation of genetic competence in Bacillus subtilis
Authors: Dubnau D. 
Published: October 2006

SummaryGenetic competence develops as a global response of Bacillus subtilis to the onset of stationary phase, in glucose‐minimal salts‐based media. The onset of competence is accompanied by the expression of several late gene products that are required for the binding, processing and uptake of transforming DNA. A number of regulatory genes have been identified that are needed for the appropriate synthesis of the late gene products. The regulatory gene products include a number of known transcription factors, as well as several members of the bacterial two‐component regulatory system. Genetic analysis has suggested a scheme for the flow of regulatory information signaling the onset of competence. Most of these regulatory products appear to be involved in the response to nutritional status, while the components responsible for growth stage and cell‐type‐specific control remain unknown. The general implications of this scheme for post‐exponential expression are discussed.
Evolution of the MIP family of integral membrane transport proteins
Authors: Pao G. M. , Wu L.‐F. , Johnson K. D. , Höfte H. , Chrispeels M. J. , Sweet G. , Sandal N. N. , Saier M. H. 
Published: October 2006

SummarySix integral membrane proteins of bacterial, animal, and plant origin, which are believed to function in solute transport, share sequence identity and are grouped together as members of the MIP family. These include the Escherichia coli glycerol facilitator, the major intrinsic protein from bovine lens fibre junction membranes, a plant tonoplast membrane protein, a soybean protein from the peribacteroid membrane, and a Drosophila neurogenic protein. These proteins, each of which appears to consist of six transmembrane helical segments per subunit, apparently arose by internal duplication of a three‐transmembrane segment. Phylogenetic‘trees’interrelating these proteins and segments are presented.
Molecular characterization of an operon (hyp) necessary for the activity of the three hydrogenase isoenzymes in Escherichia coli
Authors: Lutz S. , Jacobi A. , Schlensog V. , Böhm R. , Sawers G. , Böck A. 
Published: October 2006

SummaryThe 58/59 min region of the Escherichia coli chromosome contains two divergently oriented gene clusters coding for proteins with a function in hydrogenase formation. One cluster (the hyc operon), transcribed counterclockwise with respect to the E. coli chromosome, codes for gene products with a structural role in hydrogenase 3 formation (Böhm et al., 1990). The nucleotide sequence of the divergently transcribed operon (hyp) has been determined. It contains five genes, all of which are expressed in vivo in a T7 promoter/polymerase system, and the sizes of the synthesized products correspond with those predicted from the amino acid sequence. Complementation analysis of previously characterized mutants showed that the hypB, hypC and hypD genes have a function in the formation of all three hydrogenase isoenzymes, lesions in hypB being complemented by high nickel ion concentration in the medium. Prevention of hypBCDE gene expression led to an altered electrophoretic pattern of hydrogense 1 and 2 constituent subunits, indicating increased chemical or proteolytic susceptibility. Under fermentative growth conditions, operon expression was governed by an NtrA‐dependent promoter lying upstream of hypA working together with an fnr gene product‐dependent promoter which was localized within the hyp A gene. The latter (operon‐internal) promoter is responsible for hypBCDE transcription under non‐fermentative conditions when the ‐24/‐12 NtrA‐dependent promoter upstream of hyp A is silent.
Purification and characterization of two‐subunit cytochrome aa3 from Bacillus cereus
Authors: Garcia‐Horsman J. A. , Barquera B. , Gonzalez‐Halphen D. , Escamilla J. E. 
Published: October 2006

SummaryCytochrome c‐oxidase type aa3 (EC 1.9.3.1.) was purified to homogeneity from vegetative Bacillus cereus by ion‐exchange and hydroxylapatite chromatography in the presence of Triton X–100. Gel filtration analysis suggested a dimeric structure apparently 172kDa in size; however, only a monomer of 81kDa was detected when analysed by non‐denaturing gel electrophoresis. Denaturing gel electrophoresis analysis of the protein showed the presence of two subunits (51 and 30kDa), Atomic absorption and visible spectroscopy showed typical aa3 redox centres with haem a iron and copper in a ratio of 22 nmol and 35ng‐atom per mg protein, respectively. No haem c was found associated with the purified enzyme in the conditions reported here. Oxidase activity was fully reconstituted by phospholipids in the presence of N, N, N′ N′‐tetramethyl‐p‐phenylenediamine or reduced yeast cytochrome c (but not horse cytochrome c) as electron donors. This activity was abolished by cyanide and carbon monoxide.
Characterization of the yellow‐pigment genes of Erwinia herbicola
Authors: Lee L.‐Y. , Liu S.‐T. 
Published: October 2006

SummaryA 6.7kb DNA fragment containing the pigment genes of Erwinia herbicola Eho13 has been cloned into Escherichia coli. These genes were chromosomally encoded in E. herbicola. The entire DNA fragment could be divided into at least three regions. Deletions in Region I resulted in a non‐pigmented phenotype, a deletion in Region II resulted in a pink/yellow phenotype, deletions in Region III resulted in either a pink or a non‐pigmented phenotype. Tn 1000 insertions in the same regions, however, gave different phenotypes. Insertions in Region U produced a pink phenotype. Insertions in Region III resulted in either a light‐yellow or a non‐pigmented phenotype. Minicell studies showed that the 6.7kb DNA fragment encoded at least five proteins (50kDa, 42kDa, 36kDa, 35kDa and 34kDa). A 2.7kb Hin dIII deletion in Region I caused the disappearance of these proteins, suggesting that this 2.7 kb fragment may play a regulatory role in pigment synthesis. Our results also showed that a 4.1kb Eco RV fragment consisted of Region I and a part of Region II complemented a pink/yellow clone of Eho10 (pHL545), suggesting that the pigments of Eho13 and Eho10 were probably similar or identical.
The use of a rare codon specifically during development?
Authors: Leskiw B. K. , Bibb M. J. , Chater K. F. 
Published: October 2006

SummaryA range of circumstantial evidence suggests that in Streptomyces spp., genes required for vegetative growth do not contain the leucine codon TTA. Instead, the codon seems to be confined to a few genes necessary during differentiation, when the colonies begin to produce aerial hyphae and antibiotics. Thus, mutations in bldA, the structural gene for tRNALeuTTA, do not retard vegetative growth, but they prevent normal aerial mycelium and antibiotic production. Most of the known TTA‐containing genes specify regulatory or resistance proteins associated with antibiotic‐production clusters. Possibly the ability to translate the UUA codons in mRNA from such genes is confined to late stages of colony development. Factors that might have contributed to the evolution of this unusual situation are discussed.