bluuebunny/crossref_metadata_2025_split huggingface dataset:
Analysis of the subcellular location of pullulanase produced by Escherichia coli carrying the pulA gene from Klebsiella pneumoniae strain UNF5023
Authors: Pugsley A. P. , Kornacker M. G. , Ryter A. 
Published: October 2006

SummaryThree different techniques, protease accessibility, ceil fractionation and in situ immunocytochemistry, were used to study the location of the lipoprotein pullulanase produced by Escherichia coli K12 carrying the cloned pullulanase structural gene (pulA) from Klebsiella pneumoniae, with or without the K. pneumoniae genes required to transport pullulanase to the cell surface (secretion‐competent and secretion‐incompetent, respectively). Pullulanase produced by secretion‐competent strains could be slowly but quantitatively released into the medium by growing the cells in medium containing pronase. The released pullulanase lacked the N‐terminal fatty‐acylated cystelne residue (and probably also a short N‐terminal segment of the pullulanase polypeptide), confirming that the N‐terminus is the sole membrane anchor in the protein. Pullulanase produced by secretion‐incompetent strains was not affected by proteases, confirming that it is not exposed on the cell surface. Pullulanase cofractionated with both outer and inner membrane vesicles upon isopycnic sucrose gradient centrifugation, irrespective of the secretion competence of the strain. Examination by electron‐microscopy of vesicles labelled with antipullulanase serum and protein A‐gold confirmed that pullulanase was associated with both types of vesicles. When thin‐sectioned cells were examined by the same technique, pullulanase was found to be located mainly on the cell surface of the secretion‐competent cells and mainly in the proximity of the inner membrane in the secretion‐incompetent cells. Thus, while the results from three independent techniques (substrate accessibility, protease accessibility and in situ immunocytochemistry) show that pullulanase is transported to the cell surface of secretion‐competent cells, this could not be confirmed by cell‐fractionation techniques. Possible explanations for this discrepancy are discussed.
Processes of genome evolution reflected by base frequency differences among Serratia marcescens genes
Authors: Sharp P. M. 
Published: October 2006

SummaryThe G + C content of silent sites in codons varies greatly among Serratia marcescens genes; the value in any one gene seems to reflect a balance between mutation pressure towards high G + C content and natural selection constraining choice among synonymous codons. Interestingly, non‐coding sequences have substantially lower G + C content than silent sites thought to be under little selective constraint.
Two cyclic AMP‐regulated genes from Dictyostelium discoideum encode homologous proteins
Authors: Ramji D. P. , Richards A. J. , Jagger P. , Bleasby A. , Hames B. D. 
Published: October 2006

SummaryExpression of the 7E and 2C genes late in Dictyostelium development ceases upon cell disaggregation but, in contrast to many other genes we have studied, expression is fully restored by exogenous cAMP (A. J. Richards et af., submitted). The 7E and 2C genes encode polypeptides of similar size (9220 and 10573 Daltons, respectively), each of which contains an unusually high proportion of serine plus glycine residues (41% and 59%, respectively). Each protein possesses a relatively serine‐rich N‐terminus and glycine‐rich C‐terminus and contains the conserved sequence S(X)SSS(X2)SS(X)SS(X2)SFGS. These data suggest that genes 7E and 2C may have arisen by duplication of a common ancestor. Computer analysis indicates that both gene products are probably intra‐cellular structural proteins that form extended coil structures.
Site‐directed mutagenesis of the ‘zinc finger’ DNA‐binding domain of the nitrogen‐regulatory protein NIT2 of Neurospora
Authors: Fu Y.H. , Marzluf G.A. 
Published: October 2006

SummaryThe major nitrogen‐regulatory gene nit‐2 of Neurospora crassa activates the expression of numerous unlinked structural genes which specify nitrogen‐catabolic enzymes during conditions of nitrogen limitation. The nit‐2 gene encodes a regulatory protein of 1036 amino acid residues with a single ‘zinc finger’ and a downstream basic region, which together may constitute a DNA‐binding domain. The zinc finger domain of the NIT2 protein was synthesized nit‐3, the nitrate reductase structural gene. A series of NIT2 mutant proteins obtained by site‐directed mutagenesis was expressed and tested for functional activity. The results demonstrate that both the single zinc‐finger motif and the downstream basic region of NIT2 are critical for its trans‐activating function in vivo and specific DNA‐binding in vitro.
Rates of assembly and degradation of bacterial ice nuclei
Authors: Watanabe N. M. , Southworth M. W. , Warren G. J. , Wolber P. K. 
Published: October 2006

SummaryThe kinetics of ice‐nucleus assembly from newly synthesized nucleation protein were observed following induction of nucleation gene expression in the heterologous host Escherichia coli. Assembly was significantly slower for the small proportion of ice nuclei active above ‐4.4°C; this was consistent with the belief that these nuclei comprise the largest aggregates of nucleation protein. The kinetics of nucleus degradation were followed after inhibiting protein synthesis. Nucleation activity and protein showed a concerted decay, indicating that most of the functional ice nuclei are in equilibrium with a single cellular pool of nucleation protein. A minority of the ice nuclei decayed much more slowly than the majority; presumably their nucleation protein was distinct either by virtue of different structure or different subcellular compartmentalization, or because of its presence in a metabolically distinct subpopulation of cells.
The importance of the 5′‐region in regulating the stability of sdh mRNA in Bacillus subtilis
Authors: Melin L. , Friden H. , Dehlin E. , Rutberg L. , von Gabain A. 
Published: October 2006

SummaryThe decay of the polycistronic Bacillus subtilis sdh mRNA was analysed using probes specific for each of the component cistrons, sdhC, sdhA and sdhB. In exponentially growing cells, the entire sdh mRNA seems to decay with an ‘all or nothing’ mechanism and with a uniform half‐life of 2–3 min for all cistrons. In stationary‐phase cells, the half‐life of the 5′‐part had dropped to about 0.6 min whereas that of the 3′‐part was about 1.2 min. Decay of sdh mRNA was also measured in exponentially growing cells containing a ‘down‐mutation’ in the ribosomal binding site preceding sdhC which decreases the expression of sdhC by about 90%. The mutation has a moderate effect on expression of the downstream cistron sdhA. In this mutant, the half‐life of the 5′‐part of sdh mRNA was about 0.5 min (i.e. the same as in stationary phase wild‐type cells) and the half‐life of the 3′‐part about 1.3 min. Also, analysis of the decay of an sdh‐cat fusion transcript revealed that the sdh (5′) part decayed more rapidly than the cat part and this difference was more pronounced in stationary‐phase cells compared to exponentially growing cells. The results of these experiments demonstrate the importance of the 5‐ segment of sdh mRNA in controlling the stability of the transcript under different growth conditions.
Cryptic α‐toxin gene in toxic shock syndrome and septicaemia strains of Staphylococcus aureus
Authors: O'Reilly M. , Kreiswirth B. , Foster T. J. 
Published: October 2006

SummaryThe majority of clinical isolates of Staphylococcus aureus that produce toxic shock syndrome toxin‐1 (TSST‐1) fail to express α‐toxin, despite having a copy of the hla gene in the chromosome. The hla gene was cloned from an Hla‐ TSST‐1+ strain, Todd 555, which had been isolated from a case of toxic shock syndrome in the USA. Of the 630 bases of the Todd 555 gene sequenced, 46 differed from the hla gene sequence of strain Wood 46. The defect in α‐toxin expression was shown to be due to a nonsense mutation which converted a CAG glutamine codon in the equivalent position in the functional Wood 46 sequence to a TAG stop codon. The same mutation was present in the hla gene cloned from a human septicaemia strain (V37) isolated in Dublin. The nonsense mutation of Todd 555 was suppressed by the supE 44 mutation in Escherichia coli resulting in haemolytic activity in cell lysates. Hybrid hla genes were formed by splicing fragments of hla from Todd 555 and Wood 46. Expression of one such chimaeric hla gene in S. aureus demonstrated that the Todd 555 hla gene has a functional agr‐regulated promoter. The silent hla gene may be a cryptic gene in S. aureus.
Analysis of site‐directed mutations in the α‐and β‐subunits of Klebsiella pneumoniae nitrogenase
Authors: Kent H. M. , Baines M. , Gormal C. , Smith B. E. , Buck M. 
Published: October 2006

SummaryUsing directed mutagenesis, amino acid substitutions have been made in the α‐ and β‐subunits of the Klebsiella pneumoniae nitrogenase component 1 at positions normally occupied by conserves cysteine or tyrosine residues. Nif', Nif and intermediate pheno‐types have been obtained. To extend our earlier biochemical characterization (Kent et al, 1989) the electrophoretic mobiliy of component 1 of the mutant and wild‐type nitrogenases has been analysed by non‐denaturing gel electrophoresis. The major and minor forms of component 1 separated by this methodology have been probed for by using both polyclonal and monoclonal antibodies. All Nif mutants exhibited a distribution of electrophoretic forms of component 1 comparable to the wild type, and the abundance of the major from found in purified nitrogennase correlated approximately with the specific activity of the extract. In contrast, after electrophoresis, component 1 from Nif mutants exhibited either a major low‐mobility from or a fast‐moving from. Analysis of co‐factor (FeMoco) allowed us to conclude that changing cysteine 275 to alanine in the α‐submit produces component 1 defective in its interaction with FeMoco. Substitution of other con‐served cysteine residues by alanine appears to prevent early steps in nitrogebnase assembly or to promote degradation. Two single mutations (cysteine89 to alanine in the β‐subunit) which are tightly Nif can be combined to produce a weakly active nitrogenase, indicating regions involved in the interaction between subunits.
Mutagenesis, cloning and complementation analysis of C4‐dicarboxylate transport genes from Rhodobacter capsulatus
Authors: Hamblin M. J. , Shaw J. G. , Curson J. P. , Kelly D. J. 
Published: October 2006

SummaryTransposon mutagenesis was used to isolate insertion mutants of the photosynthetic bacterium Rhodobacter capsulatus which were unable to grow under aerobic conditions in the dark on malate, succinate or fumarate as sole carbon sources. Of five mutants isolated, all were deficient in C4‐dicarboxylate transport. However, these mutants were still capable of photoheterotrophic growth, although at a slower rate than the wild type, on malate and succinate (but not fumarate). The mutated locus (designated dct) was complemented in trans using a cosmid gene bank. Subcloning and complementation analysis indicated that at least three closely linked genes essential for aerobic dicarboxylate transport were contained within an 8.3kb region of the Rhodobacter capsulatus chromosome.
Gene sequences and comparison of the fimbrial subunits representative of Bacteroides nodosus serotypes A to I: class I and class II strains
Authors: Mattick J. S. , Anderson B. J. , Cox P. T. , Dalrymple B. P. , Bills M. M. , Hobbs M. , Egerton J. R. 
Published: October 2006

SummaryWe have determined the nucleotide sequences of the genes encoding the fimbrial subunits representative of the known Bacteroides nodosus serogroups. All of the genes are preceded by a highly conserved region which includes the likely promoter and transcriptional regulator sites as well as the ribosome‐biding site, and are followed within a short but variable distance by a sequence with the characteristics of a transcription termination or attenuation signal. Based on sequence and organization, the subunits can be divided into two major classes called I (serogroups A, B, C., E, F, G, and I) and II (serogroups D and H). All contain the same seven‐amino‐acid positively charged leader sequence and conserved hydrophobic amino‐terminal sequence typical of type 4 fimbriae. Beyond this point the class II subunits are quite different from class I and share features more in common with those from other type 4 fimbriate bacteria, such as Moraxella bovis and Pseudomonas aeruginosa. The larger class I may be further subdivided into two subsets: (i) ((A, E, F)(B, I)) and (ii) (C, G). These proteins exhibit three major clusters of variation, at either end of the presumptive disulphide loop which spans the central third of the protein, and near the carboxy‐terimus, with dispersed changes in between. The length of the mature subunits varies from 152–156 amino acids, and the variation Includes small insertions or deletions in the variable clusters between more conserved domains. The class II subunits are 149 amino acids in length and contain two pairs of cysteine residues: one is at the end of the amino‐terminal conserved region, and the other is at the end of the protein. The major variation occurs in the central region of the molecule, and again small insertions or deletions are required to align adjacent conserved domains. There is also a striking absence of silent codon changes in the 5′ coding region of all of these genes, indicating that these sequences have a secondary genetic function, probably in recombi‐national exchange.
Catabolite repression of α amylase gene expression in Bacillus subtilis involves a trans‐acting gene product homologous to the Escherichia coli lacl and galR repressors
Authors: Henkin T. M. , Grundy F. J. , Nicholson W. L. , Chambliss G. H. 
Published: October 2006

SummaryExpression of the α‐amylase gene of Bacillus subtilis is controlled at the transcriptional level, and responds to the growth state of the cell as well as the availability of rapidly metabolizable carbon sources. Glucose‐mediated repression has previously been shown to involve a site near the transcriptional start‐point of the amyE gene. In this study, a transposon insertion mutation was characterized which resulted in loss of glucose repression of amyE gene expression. The gene affected by this mutation, which was localized near 263° on the B. subtilis chromosomal map, was isolated and its DNA sequence was determined. This gene, designated ccpA, exhibited striking homology to repressor genes of the lac and gal repressor family. The ccpA gene was found to be allelic to alsA, previously identified as a regulator of acetoin biosynthesis, and may be involved in catabolite regulation of other systems as well.
Intervening sequences (IVSs) in the 23S ribosomal RNA genes of pathogenic Yersinia enterocolitica strains. The IVSs in Y enterocolitica and Salmonella typhimurium have a common origin
Authors: Skurnik M. , Toivanen P. 
Published: October 2006

SummaryThe 23S ribosomal RNA (rRNA) was shown to be in two fragments in pathogenic Yersinia enterocolitica. The cleavage site in the structural gene of the 23S rRNA was occupied by an intervening sequence (IVS) of about 100 nucleotides, analogous to IVSs found in salmonellae (Burgin et al., 1990). Nucleotide sequences of IVSs of several Y. enterocolitica strains revealed that the IVSs of the highly virulent Y. enterocolitica serotypes strains, and the IVS of Salmonella typhimurium were about 90% similar. On the other hand, the IVSs of the highly and the poorly virulent Y. enterocolitica serotypes were only about 60% similar. These results give the impression that at some point during the IVS evolution, the highly virulent Y. enterocolitica and S. typhimurium both received their IVSs at about the same time from the same source, and that the poorly virulent serotypes received their IVSs earlier. We also found that strain LB5010, derived by extended mutagenization of S. typhimurium LT2, had lost the IVSs originally present in LT2, and that this loss had created a new ‘hairpin loop’ which substituted for the original ‘hairpin loop’.
The ferric‐pseudobactin receptor PupA of Pseudomonas putida WCS358: homology to TonB‐dependent Escherichia coli receptors and specificity of the protein
Authors: Bitter W. , Marugg J. D. , de Weger L. A. , Tommassen J. , Weisbeek P. J. 
Published: October 2006

SummaryThe initial step in the uptake of iron via ferric pseudobactin by the plant‐growth‐promoting Pseudomonas putida strain WCS358 is binding to a specific outer‐membrane protein. The nucleotide sequence of the pupA structural gene, which codes for a ferric pseudobactin receptor, was determined. It contains a single open reading frame which potentially encodes a polypeptide of 819 amino acids, including a putative N‐terminal signal sequence of 47 amino acids. Significant homology, concentrated in four boxes, was found with the TonB‐dependent receptor proteins of Escherichia coli. The pupA mutant MH100 showed a residual efficiency of 30% in the uptake of 55HFe3+ complexed to pseudobactin 358, whereas the iron uptake of four other pseudobactins was not reduced at all. Cells of strain WCS374 supplemented with the pupA gene of strain WCS358 could transport ferric pseudobactin 358 but showed no affinity for three other pseudobactins. It is concluded that PupA is a specific receptor for ferric pseudobactin 358, and that strain WCS358 produces at least one other receptor for other pseudobactins.
Transcriptional analysis of the restriction and modification genes of bacteriophage P1
Authors: Sharrocks A. D. , Hornby D. P. 
Published: October 2006

SummaryBacteriophage P1 res and mod genes encode the restriction and modification polypeptides of the Type III restriction enzyme EcoP1. Northern blot analysis using res‐ and mod‐specific probes revealed the presence of two separate transcripts in strains harbouring the Eco P1 restriction and modification genes. Furthermore, by constructing a series of fusions with a promoterless lacZ gene, we show that both the res and mod genes are transcribed from separate promoters. A more detailed investigation of the mod promoter region revealed two promoters located some 70 and 140 bp upstream from the translational start codon. In addition, another pair of promoters and a further separate promoter are located more than 500 bp upstream from this start codon. Two short open reading frames are located between these distal and proximal promoter clusters.Transcription of the res gene is initiated from within the mod open reading frame from two adjacent promoters. In addition a functional promoter is located on the antisense strand close to the res promoter region. The relationship between the transcription units of the res and mod genes is discussed.
Structure and sequence of the rfb (O antigen) gene cluster of Salmonella serovar typhimurium (strain LT2)
Authors: Jiang X.‐M. , Neal B. , Santiago F. , Lee S. J. , Romana L. K. , Reeves P. R. 
Published: October 2006

SummaryThe rfb gene cluster of Salmonella LT2 has been cloned and sequenced. The genes rfbA, rfbB, rfbD, rfbF, rfbG, rfbK, rfbM and rfbP were located individually and the gene rfbL was located outside the cluster. Approximately 16 open reading frames were found in the region which is essential for the expression of O antigen. The gene products of rfbB and rfbG were found to have homology with the group of dehydrogenase and related enzymes described previously. Analysis of the G+C ratio of the rfb cluster extended the area of low‐G+C composition previously found in the sequence of rfbJ to the whole rfb gene cluster. Three to five segments with discrete G+C contents and codon adaptation indices are present in the rfb region, indicating a heterogeneous origin of these segments. Potential promoters were found near the start of the rfb region, supporting the possibility that the rfb gene cluster is an operon.
The role of σF in prespore‐specific transcription in Bacillus subtilis
Authors: Partridge S. R. , Foulger D. , Errington J. 
Published: October 2006

SummarySporulation in Bacillus subtilis is a simple developmental system in which a single cell undergoes differentiation to two ‘sister’ cells, namely the prespore and the sporangium. Prespore‐specific gene expression is largely dependent on the synthesis of a transcription factor, σG. Transcription of spolllG, the gene encoding σG, is under precise temporal and spatial control, requiring the products of at least eight genes that are expressed in the pre‐divisional cell. Here we show that the product of one of these genes, another sigma factor, σF, is by itself sufficient to direct transcription of spolllG in non‐sporulating cells. The results indicate that the cell‐specificity of prespore gene expression is determined by a mechanism that exerts temporal and spatial control over the activity of σF.
Calcium in bacteria: a solution to which problem?
Authors: Norris V. , Chen M. , Goldberg M. , Voskuil J. , McGurk G , Holland B. 
Published: October 2006

SummaryCalcium and calcium‐binding proteins including those resembling calmodulin are implicated in numerous diverse processes in bacteria. These processes include chemotaxis, sporulation, virulence, the transport of sugars and proteins, phosphorylation, heat shock, the initiation of DNAS replication, septation, nucleoid structure, nuclease activity and recombination, the stability of the envelope, and phospholipids synthesis and configuration. That such varied processes should have a common factor, calcium, suggests major underlying principles of calcium metabolism metabolism which have yet to be discovered.
ERIC sequences: a novel family of repetitive elements in the genomes of Escherichia coli, Salmonella typhimurium and other enterobacteria
Authors: Hulton C. S. J. , Higgins C. F. , Sharp P. M. 
Published: October 2006

SummaryWe describe a family of highly conserved, Enterobacterial Repetitive Intergenic Consensus (ERIC) sequences, 14 of which have been identified in Escherichia coli and Salmonella typhimurium and a further three in other enterobacterial species (Yersinia pseudotubercuiosis, Kiebsiella pneumoniae and Vibrio cholerae). ERIC sequences are 126 bp long and appear to be restricted to transcribed regions of the genome, either in intergenic regions of polycistronic operons or in untranslated regions upstream or downstream of open reading frames. ERIC sequences are highly conserved at the nucleotide sequence level but their chromosomal locations differ between species. Several features of ERIC sequences resemble those of REP sequences (Stern et al., 1984) although the nucleotide sequence is entirely different. The question of whether ERICs have a specific function, or represent a form of 'selfish’DNA, is discussed.
Formation of Rhizobium phaseoli symbiotic plasmids by genetic recombination
Authors: Soberón‐Chavez G. , Nájera R. , Espín G. , Moreno S. 
Published: October 2006

SummaryWe report here the formation of symbiotic plasmids (pSyms), by genetic recombination between rearranged pSyms, which lack symbiotic information, and resistance plasmids carrying parts of different symbiotic plasmids (R's). This recombination was found to occur both between plasmids derived from different Rhizobium phaseoli isolates, and between plasmids derived from strains obtained from the same original isolate.We also present evidence on the formation of a functional symbiotic plasmid by recombination of an R, carrying nif and nod genes from strain CFN42, and an indigenous plasmid present in this strain (pCFN42e), which was thought to be unrelated to its symbiotic plasmid (pCFN42d). These data are discussed with respect to the stability and transfer of Rhizobium symbiotic information.