bluuebunny/crossref_metadata_2025_split huggingface dataset:
Nucleotide sequence and organization of the upstream region of the Corynebacterium glutamicum lysA gene
Authors: Marcel T. , Archer J. A. C. , Mengin‐Lecreulx D. , Sinskey A. J. 
Published: October 2006

SummaryMaximum expression of the Corynebacterium glutamicum lysA gene is dependent upon the presence of a 2.3 kb region immediately 5’of the lysA reading frame. Subcloning and functional analysis of the upstream region implied that this region contained the lysA promoter. Sequence determination of the upstream region revealed a single open reading frame, orfX, in the same orientation as lysA. The orfX coding sequence exhibited all the sequence characteristics of a gene with the potential for a 550‐amino‐acid polypeptide product. Expression of lysA is coupled to that of orfX via a common promoter located immediately 5 of orfX. The RNA start site has been determined by S1 nuclease mapping. Both the orfX and the lysA gene are expressed as a single 3.0kb RNA transcript. These data indicate that orfX and lysA are genes within a two‐gene operon. Expression of the lysA gene is not subject to regulation by lysine. The orfX gene product was shown not to be directly linked to the lysine biosynthetic pathway, nor is it the enzyme incorporating DAP into the peptidoglycan precursor.
Instability of bacteriophage Mu transposase and the role of host Hfl protein
Authors: Gama M.‐J. , Toussaint A. , Pato M. L. 
Published: October 2006

SummaryThe activity of the transposase of bacteriophage Mu is unstable, requiring the protein to be synthesized throughout the lytic cycle (Pato and Reich, 1982). Using Western blot analysis, we analysed the stability of the transposase protein during the lytic cycle and found that it, too, is unstable. The instability of the protein is observed both in the presence and the absence of Mu ONA replication, and is independent of other Mu‐encoded proteins and the transposase binding sites at the Mu genome ends. Stability of the protein is enhanced in host strains mutated at the hfl locus; however, stability of the transposase activity is not enhanced in these strains, suggesting that functional inactivation of the protein is not simply a result of its proteolysis.
Role of the nodD and syrM genes in the activation of the regulatory gene nodD3, and of the common and host‐specific nod genes of Rhizobium meliloti
Authors: Maillet F. , Debellé F. , Dénarié J. 
Published: October 2006

SummaryTo analyse the regulation of the nodulation (nod) genes of Rhizobium meliloti RCR2011 we have isolated lacZ gene fusions to a number of common, host‐range and regulatory nod genes, using the mini‐MU‐lac bacteriophage transposon Mud II1734. Common (nodA, nodC, nod region IIa) and host‐range (nodE, nodG, nodH) genes were found to be regulated similarly. They were activated (i) by the regulatory nodD1 gene in the presence of flavones such as chrysoeriol, luteolin and 7,3′,4′‐trihydroxyflavone, (ii) by nodD2 in the presence of alfalfa root exudate but not with the NodD1‐activating flavones, and (iii) by the regulatory genes syrM‐nodD3 even in the absence of plant inducers. Thus common and host‐range nod genes belong to the same regulon. In contrast to the nodD1 gene, the regulatory nodD3 gene was not expressed constitutively and exhibited a complex regulation. It required syrM for expression, was activated by nodD1 in the presence of luteolin and was positively autoregulated.
Towards an understanding of the structural basis of ‘forbidden’ transport pathways in the Escherichia coli lactose carrier: mutations probing the energy barriers to uncoupled transport
Authors: King S. C. , Wilson T. H. 
Published: October 2006

SummaryRecent progress in the analysis of mutants of the Eschenchia coli lactose carrier function is reviewed, with special emphasis on the structural basis for energy barriers which prevent ‘forbidden’ con‐formational changes. Mutations which break down the barriers to forbidden isomerizations involving the binary carrier:sugar (CS) and carrier:proton (CH) complexes have been obtained in several laboratories. These mutants allow uncoupled transport of H+ or galactoside in the lactose carrier which normally couples cation and sugar movement in a 1:1 stoichiometry. These uncoupled mutants appear to be associated with changes in both sugar and cation recognition, suggesting that the physical interactions forming the basis for co‐substrate recognition and uncoupling are not independently variable.By postulating that translocation involves transformation of the stable intermediate of the co‐transport cycle to unstable transition state conformations of the carrier, it is possible to consider the consequences of mutagenesis in terms of transition state theory. Consistent with several experimental observations, the analysis predicts in each mutant the occurrence of more than one abnormality in the transport cycle (such as changes in sugar recognition, cation recognition or the coupling reaction). We have called the general phenomenon a ‘mutational double‐effect’ because any mutation which alters the Gibbs free energy change of one reaction in the transport
How Escherichia coli RNA polymerase can negatively regulate transcription from a constitutive promoter
Authors: Duval‐Valentin G. , Reiss C. 
Published: October 2006

SummaryWe previously described the structures and functions of specific complexes between the bla promoter from Tn3(present in pBR322) and RNA polymerase (RNAP), showing that, at excess RNAP, complexes can form in which one or two RNAPs bind to the same promoter (1:1 and 2:1 complexes) (Duval‐Valentin and Ehrlich, 1988). We report here that the 2:1 complex cannot be detected below 25° C; above that temperature, a 1:1 complex forms at a rate one order of magnitude faster than that of the 2:1 complex, and above 30° C, the amounts of both species become equal for RNAP/promoter ratio r30≤r≤70. The 2:1 complex decays back to a 1:1 complex losing the last RNAP at a rate about three times that of the 1:1 complex decay. Functional assays of the complexes formed at excess RNAP show that both 1:1 and 2:1 complexes are immediately and permanently inhibited, even when the promoters are pre‐incubated with ribonucleotide selections potentially enabling entrance into abortive cycling or formation of a stressed complex. We conclude that the inhibition step probably takes place in the complex formation pathway between RPi and RPo, at a novel stable intermediate isomer, RPj, formed above 25° C. A possible mechanism of formation of the 2:1 complex is outlined. In vivo studies, in which r was modified by varying the bacterial growth rate, show a reduction of bla expression as r values are upshifted, specific to the bla promoter from Tn3.
Import‐defective colicin B derivatives mutated in the TonB box
Authors: Mende J. , Braun V. 
Published: October 2006

SummaryThe pore‐forming colicin B is taken up into Escherichia coli by a receptor and TonB‐dependent process. The receptor and colicin B both contain a similar amino acid sequence, close to the N‐terminal end, termed the TonB box. Point mutations were introduced into the TonB‐box region of the colicin B structural gene cba resulting in colicin B derivatives which were partially or totally Inactive against E. coli cells. All derivatives still bound to the receptor. An inactive derivative killed cells when translocated across the outer membrane by osmotic shock treatment, and formed pores in planar lipid bilayer membranes identical to the wild‐type colicin. Some of the mutations were partially suppressed by mutations in the tonB structural gene. It was concluded that the TonB‐box mutations define a region that is involved in the uptake of colicin B across the outer membrane.
Pore‐forming cytolysins of Gram‐negative bacteria
Authors: Welch R. A. 
Published: October 2006

SummaryA great deal is known about the structure, function and metabolic effects of enzymatic bacterial toxins such as the diphtheria, pertussis and cholera toxins. By comparison, our understanding of the pore‐forming, cytolytic toxins, particularly those produced by Gram‐negative bacterial pathogens, is far less complete. The genetics and biochemistry of a large, newly discovered family of calcium‐dependent, pore‐forming cytotoxins (RTX toxins) produced by different genera of the Enterobacteriaceae and Pasteurellaceae are discussed in this review. This toxin family is especially noteworthy because the individual toxins often exhibit different cell‐ and host‐specificity. A brief review is also included of two ancestrally unrelated groups of calcium‐independent, pore‐forming toxins, the haemolysins produced by Proteus mirabilis and Serratia marcescens and the aerolysins secreted by species of Aeromonas. Their structure and function are contrasted with those of the RTX family members. Emerging questions about the role of cytolysins in pathogenesis are presented. Perhaps the most important issue raised is whether or not less attention should be paid to the the lytic capacity of these cytotoxins, with more energy being devoted to the understanding of their non‐lytic Inhibitory activities against host cells.
Organization of the fimbrial gene region of Bacteroides nodosus: class I and class II strains
Authors: Hobbs M. , Dalrymple B. P. , Cox P. T. , Livingstone S. P. , Delaney S. F. , Mattick J. S. 
Published: October 2006

SummaryThe fimbrial subunit genes of Bacteroides nodosus may be divided into two distinct classes, based on the sequence of the major subunit gene fimA (accompanying paper — Mattick et al., 1991). The genetic organization of the fimbrial gene region in these two classes is also distinct. Upstream of fimA in both classes in opposite transcriptional orientation is the gene aroA which encodes amino acid biosynthetic enzyme 5‐enolpyruvylshikimate‐3‐phosphate synthase. However, downstream of fimA the two classes are quite different until homology is restored at a bidirectional transcription termination signal separating the fimbrial operon from a gene clpB, which appears to encode the regulatory subunit of an ATP‐dependent protease. Between aroA and clpB class I strains contain, apart from fimA, only one other gene (fimB). Sequence and polymerase chain reaction analyses indicate that fimB does not have a separate promoter but rather is co‐transcribed with fimA at a level attenuated by the strength of the transcription termination signal in the intergenic region. In class II strains fimA is followed by a more extended region containing three genes, which appear to have the same transcriptional arrangement as fimB. The second of these genes (fimD) may represent a functional analogue of fimB although there is no close sequence homology. The first gene (fimC) has no obvious similarity to either fimB or fimD. Beyond fimO, at the 3 end of the class ll‐specific region, is a variant fimbrial subunit gene (fimZ) which is virtually identical in serogroups 0 and H and which appears to represent a duplicate, possibly redundant, gene closely related to the progenitor of the more divergent structural subunit fimA gene found in these strains. Comparisons of the predicted ftmZ product with those of fimA in class I and class II strains, as well as of the boundaries of the class‐specific regions, suggest that the class II sequences evolved in another type 4 fimbriate species and were subsequently substituted in the 8. nodosus genome by lateral transfer. Analysis of the sequences flanking fimA in different strains indicates that recombinational exchange of both fimA and the entire operon has also occurred between strains, and is possibly a mechanism for disseminating structural diversity in the population.
Ionizing groups in lipopolysaccharides of Pseudomonas cepacia in relation to antibiotic resistance
Authors: Cox A. D. , Wilkinson S. G. 
Published: October 2006

SummaryContrary to previous reports, lipopolysaccharides from Pseudomonas cepacia contain a 3‐deoxyoct‐2‐ulosonic acid (probably a single residue). The lipopolysaccharides contain only two phosphate residues, one of which apparently forms a phosphodiester bridge between 4‐amino‐4‐deoxyarabinose and a glucosamine residue in lipid A. The second, unlocated phosphate residue occurs mainly as a monoester in some lipopolysaccharides, and mainly as a diester in others. All lipopolysaccharides lack pyrophosphate residues. The results support the view that the resistance of P. cepacia to cationic antibiotics stems from ineffective binding to the outer membrane, as a consequence of the low number of phosphate and carboxylate groups in the lipopolysaccharide, and the presence of the protonated aminodeoxypentose.
Involvement of fixLJ in the regulation of nitrogen fixation in Azorhizobium caulinodans
Authors: Kaminski P. A. , Elmerich C. 
Published: October 2006

SummaryA gene bank of Azorhizobium caulinodans DNA constructed in the bacteriophage λGEM11 was screened with Rhizobium meliloti fixL and fixJ genes as probes. One positive recombinant phage, ORSλL, was isolated. The nucleotide sequence of a 3.7 kb fragment was established. Two open reading frames of 1512 bp and 613bp were identified as fixL and fixJ. Kanamycin cartridges were inserted into the cloned fixL and fixJ genes and recombined into the host genome. The resulting mutants were Nif− Fix−, suggesting that the two genes were required for symbiotic nitrogen fixation and for nitrogen fixation in the free‐living state. Using pnifH–lacZ and pnifA–lacZ fusions, it was shown that the FixLJ products controlled the expression of nifH and nifA in bacteria grown in the free‐living state.
Comparison of the class 1 outer membrane proteins of eight serological reference strains of Neisseria meningitidis
Authors: Maiden M. C. J. , Suker J. , McKenna A. J. , Bygraves J. A. , Feavers I. M. 
Published: October 2006

SummaryPrimers suitable for the amplification of the gene encoding the class 1 outer membrane protein of Neisseria meningitidis by the polymerase chain reaction (PCR) were designed from published DNA sequences and used to study the gene in eight meningococcal strains of different serogroup, serotype and subtype. At high annealing stringency one product, shown to correspond to the class 1 protein gene, was amplified from each strain. For three strains an additional smaller product, provisionally identified as the gene encoding the class 3 outer membrane protein, was amplified at lower annealing stringencies. Nucleotide sequence analysis of the PCR products corresponding to the class 1 proteins established the differences in the primary structure of the proteins between each of the subtypes and other outer‐membrane proteins from Neisseria spp. These differences impose constraints on possible structural models of these proteins. Most amino acid sequence variation occurred in two domains of between 8 and 17 amino acids; there was an additional region which varied mainly between classes of outer membrane protein and there were nine conserved regions. Using appropriate primers it was possible to distinguish between class 1 outer membrane protein genes from strains of different subtypes by the PCR.
Local DNA topology and gene expression: the case of the leu‐500 promoter
Authors: Lilley D. M. J. , Higgins C. F. 
Published: October 2006

SummaryMany promoters are sensitive to DNA supercoiling, and it is becoming apparent that this may play an important rote in gene regulation. The twin super ‐coiled‐domain hypothesis (Liu and Wang, 1987) proposes that transcription can lead to local variation in supercoiling. The mutant leu‐500 promoter has presented a long‐standing problem to the understanding of the control of promoter function by DNA supercoiling. This promoter is activated by mutations in the gene encoding topoisomerase I, but is apparently unaffected by mutations in the genes encoding DNA gyrase. We propose a model to explain the anomalous regulation of this promoter, based on the twin super‐coiled‐domain model. This allows us to account for the unusual properties of the leu‐500 promoter, and confirms the biological importance of the twin super‐coiled‐domain model. We suggest that such topological coupling between promoters may be general, leading to co‐operativity and anti‐co‐operativity between divergent promoter pairs.
A model for asymmetric septum formation during sporulation in Bacillus subtilis
Authors: Errington J. 
Published: October 2006

SummaryMany differentiation processes in both prokaryotes and eukaryotes begin with an asymmetric division, producing ‘daughter’ cells that differ in size and developmental fate. This is particularly obvious in the well‐studied prokaryotic lif cycles of Caulobacter and Bacillus. In no system, however, is the mechanism of asymmentric division understood. Here I propose a model for the mechanism o asymmetric division during sporulation in Bacillus subtilis. The model explains bth the timing and asymmetric localization of spore‐septum formation. It also explains the morphological phenotypes of various asporogenous (spo) mutants.
Characterization of the RNA polymerases of Crithidia fascicuiata
Authors: Köck J. , Cornelissen A. W. C. A. 
Published: October 2006

SummaryWe have characterized the crithidial RNA polymerases (RNAPs). The RNAPs from this organism were resolved by chromatography and could be classified into RNAP I, II and III because of their differential sensitivity to class‐specific RNA polymerase inhibitors, such as alpha‐amanitin (ama) and tagetitoxin. The three RNAP classes were subsequently characterized in nuclear run‐ons using class‐specific DNA templates and the inhibitory effect of ama on RNA chain elongation. These experiments showed that Crithidia fasciculata contains the normal set of eukaryotic RNAPs. However, RNAP II was found to be relatively resistant to ama, which seems to be a general feature of kinetoplastid parasites. Tagetitoxin, a potent inhibitor of purified crithidial RNAP III, does not inhibit RNAP lit in nuclear run‐on experiments, suggesting that the inhibitory effect of tagetitoxin depends on the enzyme conformation, i.e.‘naked’enzyme versus that in a transcription complex. Finally, the role of RNAP Ml in the transcription of the mini‐exon genes, which provide the 5′ end of each mRNA, is discussed.
Two distinct steps in pullulanase secretion by Escherichia coli K12
Authors: Pugsley A. P. , Poquet I. , Kornacker M. G. 
Published: October 2006

SummaryTwo distinct steps in the secretion of the extracellular, cell‐surface‐anchored lipoprotein pullulanase by Escherichia coli were uncoupled by allowing export of the enzyme to the cytoplasmic membrane via the signal peptide sec‐gene‐dependent general export pathway, and then inducing the pulC—0 operon of genes required for translocation to the cell surface. The secretion intermediate cofractionated mainly with intermediate‐density vesicles when cells were gently lysed and the resulting vesicles were separated by isopycnic sucrose density centrifugation. Cytoplasmic forms of pullulanase (which are not exported because they lack a functional signal peptide) are more sensitive to heat inactivation, denaturation by sodium dodecyl sulphate and carboxymethylation than the intermediate and cell‐surface forms. The latter are distinguished only by the fact that the secretion intermediate is less susceptible to proteinase K and trypsin, and is partially inaccessible to substrate or in an inactive conformation in sphaeroplasts. These and other results indicate that the secretion intermediate can acquire considerable higher‐ordered structure, including disulphide bridges, before it is transported to the cell surface; this seems to rule out the possibility that it is threaded through this membrane as a locally unfolded polypeptide.
A novel function of the cAMP‐CRP complex in Escherichia coli: cAMP‐CRP functions as an adaptor for the CytR repressor in the deo operon
Authors: Søgaard‐Andersen L. , Pedersen H. , Hoist B. , Valentin‐Hansen P. 
Published: October 2006

SummaryUnlike classical bacterial repressors, the CytR repressor of Escherichia coli cannot independently regulate gene expression. Here we show that CytR binding to the deoP2 promoter relies on interaction with the master gene regulatory protein, CRP, and, furthermore, that cAMP‐CRP and CytR bind co‐operatively to deoP2. Using mutant promoters we show that tandem, properly spaced DNA‐bound cAMP‐CRP complexes are required for this co‐operative binding. These data suggest that CytR forms a bridge between tandem cAMP‐CRP complexes, and that cAMP‐CRP ftjnctions as an adaptor for CytR. The implications of this new version of negative control in E. coli on bacterial gene expression and on combinatorial gene regulation in higher organisms are discussed.
Construction and characterization of Bordetella pertussis mutants lacking the vir‐regulated P.69 outer membrane protein
Authors: Roberts M. , Fairweather N. F. , Leininger E. , Pickard D. , Hewlett E. L. , Robinson A. , Hayward C. , Dougan G. , Charles L. G. 
Published: October 2006

SummaryThe Bordetella pertussis P.69 protein is an immunogen with vaccine potential. The role of this protein in pathogenesis is unclear; it has been associated with the toxic adenylate cyclase and adhesion to eukaryotic cells. For further analysis of the role of P.69 in the biology of B. pertussis, we have constructed strains which specifically lack P.69.The cloned P.69 (Prn) gene of S. pertussis was insertionally inactivated with a kanamycin‐resistance cassette. This inactivated gene was used to construct P.69− mutants of B. pertussis by allelic exchange using plasmid pRTP1. B. pertussis P.69 strains produced normal levels of other vir‐regulated factors, including adenylate cyclase. The serotype of B. pertussis, determined by Eldering and Preston typing sera and monoclonal antibodies, was also unaffected by the presence or absence of P.69.The ability of a prn mutant to adhere to and invade HEp2 cells was not significantly different from that of its parent strain. A strain containing a mutation in fhaB was significantly less adhesive and invasive than its parent, and a prn fhaB double mutant exhibited an even greater reduction in adhesiveness and invasiveness down to levels comparable with a Vir− strain.
Organization of the genes coding for the reaction‐centre L and M subunits and B870 antenna polypeptides α and β from the aerobic photosynthetic bacterium Erythrobacter species OCH114
Authors: Liebetanz R. , Hornberger U. , Drews G. 
Published: October 2006

SummaryIn the aerobic photosynthetic bacterium Erythrobacter species OCH114 the structural genes coding for the light‐harvesting (LH) complex B870 and the reaction‐centre (RC) polypeptides (the gene products of the pufB, pufA, pufL and pufM genes) are mapped on a 2.728kbp Eco RI fragment. Sequencing of this fragment revealed that the deduced amino acid sequences contain 50 (B870 β), 52 (B850 α), 283 (RCL) and 331 (RCM) residues with the corresponding molecular weights of 5592, 5814, 31364, and 37671, respectively. In the corresponding mRNA a ‘hairpin’ structure (ΔG°= ‐26.6 kcal) is predicted to be located immediately downstream of pufA. The RC and LH polypeptides are highly homologous to those of the purple photosynthetic bacteria Rhodobacter capsulatus, Rhodo‐bacter sphaeroides and Rhodopseudomonas viridis. Directly downstream of pufM there is an open reading frame (ORF) of unknown size. Partial sequencing indicates that this ORF is highly homologous to the cytochrome subunit of the photosynthetic reaction centre from R. viridis. In the puf operon no pufQ or pufX genes could be found, but the bchA gene is located upstream of that operon. Plasmid pESS8.9 containing the 2.728kbp Eco RI fragment reconstituted a photo‐inactive mutant of Erythrobacter species OCH114. Comparative analysis of the DNA region upstream of the puf operon and of bacteriochlorophyll (Bchl) synthesis indicated that Bchl synthesis and puf gene expression are regulated differently in Erythrobacter and purple bacteria, respectively.