bluuebunny/crossref_metadata_2025_split huggingface dataset:
Exploring the Mycoplasma capricolum genome: a minimal cell reveals its physiology
Authors: Bork Peer , Ouzounis Christos , Casari Georg , Schneider Reinhard , Sander Chris , Dolan Maureen , Gilbert Walter , Gillevet Pat M. 
Published: October 2006

SummaryWe report on the analysis of 214 kb of the parasitic eubacterium Mycoplasma capricolum sequenced by genomic walking techniques. The 287 putative proteins detected to date represent about half of the estimated total number of 500 predicted for this organism. A large fraction of these (75%) can be assigned a likely function as a result of similarity searches. Several important features of the functional organization of this small genome are already apparent. Among these are (i) the expected relatively large number of enzymes involved in metabolic transport and activation, for efficient use of host cell nutrients; (ii) the presence of anabolic enzymes; (iii) the unexpected diversity of enzymes involved in DNA replication and repair: and (iv) a sizeable number of orthologues (82 so far) in Escherichia coli. This survey is beginning to provide a detailed view of how M. capricolum manages to maintain essential cellular processes with a genome much smaller than that of its bacterial relatives.
Molecular characterization of the eps gene cluster of Pseudomonas solanacearum and its transcriptional regulation at a single promoter
Authors: Huang Jianzhong , Schell Mark 
Published: October 2006

SummaryProduction of EPS I, an unusual exopolysaccharide virulence factor of the phytopathogen Pseudomonas solanacearum, requires the 18 kb eps gene cluster. DNA sequence analysis of the first seven genes of eps (epsAPBCDEF), subcellular localization of their products in maxicells, and phoA fusion analysis showed that: (i) epsA, epsB, epsE, and epsF encode exported or membrane‐associated proteins probably involved in polymerization and/or export of EPS I; (ii) epsC and epsD encode soluble enzymes probably involved in synthesis of sugar components of EPS I (N‐acetylgalactosaminuronic acid and possibly N‐acetyitrideoxygalactose, respectively); and (iii) epsP probably encodes a phosphatase involved in EPS I production in an unknown way. Non‐polar insertional mutagenesis showed that most, if not all, of these eps genes are absolutely required for production of EPS I. Using random eps::lacZ fusions and primer extension we located a transcription start site and promoter upstream of epsA. Analysis of a plasmid with this promoter fused to lacZ showed that a 140 bp regulatory region upstream of the eps transcription start site was sufficient for normal regulation of eps transcription by the multicomponent virulence gene regulatory network of P. solanacearum. Deletion of this eps promoter from a plasmid‐borne epsAPBCDE::lacZ fusion reduced its expression 10‐fold, indicating that this promoter alone is responsible for regulated transcription of an eps operon composed of at least epsAPBCDE. Analysis of genomic and plasmid‐borne eps::lacZ fusions suggested that most remaining eps genes are part of this same operon or, and this is less likely, comprise a second co‐ordinately regulated eps operon.
Importance of lipooligosaccharide structure in determining gonococcal resistance to hydrophobic antimicrobial agents resulting from the mtr efflux system
Authors: Lucas Claressa E. , Hagman Kayla E. , Levin James C. , Stein Daniel C. , Shafer William M. 
Published: October 2006

SummaryLevels of gonococcal resistance to antimicrobial hydrophobic agents (HAs) are controlled by the mtr (multiple transferrable resistance) system, composed of the mtrRCDE genes. The mtrR gene encodes a transcriptional repressor that appears to regulate expression of the upstream and divergent mtrCDE operon. The mtrCDE genes encode membrane proteins analogous to the MexABOprK proteins of Pseudomonas aeruginosa that mediate export of structurally diverse antimicrobial agents. In this study we found that a single base pair deletion in a 13bp inverted repeat sequence within the mtrR promoter resulted in increased resistance of gonococci to both crystal violet (CV) and erythromycin (ERY) as well as to the more lipophilic non‐ionic detergent Triton X‐100 (TX‐100). However, this cross‐resistance was contingent on the production of a full‐length lipooligosaccharide (LOS) by the recipient strain used in transformation experiments. Introduction of this mutation (mtrR‐171) into three chemically distinct deep‐rough LOS mutants by transformation resulted in a fourfold increase in resistance to TX‐100 compared with a 160‐fold increase in an isogenic strain producing a full‐length LOS. However, both wild‐type and deep‐rough LOS strains exhibited an eightfold increase in resistance to CV and ERY as a result of the mtrR‐171 mutation. This suggests that gonococci have different LOS structural requirements for mtr‐mediated resistance to HAs that differ in their lipophilic properties. Evidence is presented that gonococci exclude HAs by an energy‐dependent efflux process mediated by the mtr system.
Structural requirements for the glycolipid receptor of human uropathogenic Escherichia coli
Authors: Striker Robert , Nilsson Ulf , Stonecipher Andrea , Magnusson Göran , Hultgren Scott J. 
Published: October 2006

SummaryThe binding of uropathogenic Escherichia coli to the globo series of glycolipids via P pili is a critical step in the infectious process that is mediated by a human‐specific PapG adhesin. Three classes of PapG adhesins exist with different binding specificities to Galα4Gal‐containing glycolipids. The structural basis for PapG recognition of the human glycolipid receptor globoside was investigated by using soluble saccharide analogues as inhibitors of bacterial haemagglutination. The minimum binding epitope was confirmed as the Galα4Gal moiety, but parts of the GalNAcβ and glucose residues, which flank the Galα4Gal in globoside (GbO4), were also shown to be important for strong binding. Furthermore, the same five hydroxyl groups of Galα4Gal in globotriasyl ceramide that were recognized by a previously characterized PapG variant were also recognized by the human‐specific PapG in binding the GbO4 that dominates In the human kidney. Saccharide analogues that blocked haemagglutination also blocked the adherence of human uropathogenic E. coli to human kidney sections. Knowledge of the molecular details of the PapG‐GbO4 interaction will make it possible to design antiadherence therapeutics.
Screening of stable proteins in an extreme thermophile, Thermus thermophilus
Authors: Tamakoshi Masatada , Yamagishi Akihiko , Oshima Tairo 
Published: October 2006

SummaryThe leuB gene codes for 3‐isopropylmalate dehydrogenase of the leucine biosynthetic pathway in an extreme thermophile, Thermus thermophilus. The leuB gene of the thermophile was replaced with a temperature‐sensitive chimeric leuB gene. The resultant transformant was adapted to high temperature, a thermostable mutant strain being obtained. A single base substitution that replaces isoleucine at 93 with leucine was found in the chimeric leuB gene of the thermostable mutant. The resultant amino acid residue coincided with the corresponding residue of the T. thermophilus enzyme. It was confirmed that the mutant enzyme is more stable than the original chimeric enzyme. This system can be used to produce stabilized mutants of other enzymes without structural knowledge of them.
Hypothesis: chromosome separation in Escherichia coli involves autocatalytic gene expression, transertion and membrane‐domain formation
Authors: Norris Vic 
Published: October 2006

SummaryTo explain how daughter chromosomes are separated into discrete nucleoids and why chromosomes are partitioned with pole preferences, I propose that differential gene expression occurs during DNA replication in Escherichia coli. This differential gene expression means that the daughter chromosomes have different patterns of gene expression and that cell division is not a simple process of binary fission. Differential gene expression arises from autocatalytic gene expression and creates a separate proteolipid domain around each developing chromosome via the coupled transcription‐translation–insertion of proteins into membranes (transertion). As these domains are immiscible, daughter chromosomes are simultaneously replicated and separated into discrete nucleoids. I also propose that the partitioning relationship between chromosome age and cell age arises because the poles of cells have a proteolipid composition that favours transertion from one nucleoid rather than from the other. This hypothesis forms part of an ensemble of related hypotheses which attempt to explain cell division, differentiation and wall growth in bacteria in terms of the physical properties and interactions of the principal constituents of cells.
Colicin import and pore formation: a system for studying protein transport across membranes?
Authors: Lazdunski Claude J. 
Published: October 2006

SummaryPore‐forming colicins are a family of protein toxins (Mr40–70kDa) produced by Escherichia coli and related bacteria. They are bactericidal by virtue of their ability to form ion channels in the inner membrane of target cells. They provide a useful means of studying questions such as toxin action, polypeptide translocation across and into membranes, voltage‐gated channels and receptor function. These colicins bind to a receptor in the outer membrane before being translocated across the cell envelope with the aid of helper proteins that belong to nutrient‐uptake systems and the so‐called‘Tol’proteins, the function of which has not yet been properly defined. A distinct domain appears to be associated with each of three steps (receptor binding, translocation and formation of voltage‐gated channels). The Tol‐dependent uptake pathway is described here. The structures and interactions of TolA, B, Q and R have by now been quite clearly defined. Transmembrane α‐helix interactions are required for the functional assembly of the E. coli Tol complex, which is preferentially located at contact sites between the inner and outer membranes. The number of colicin translocation sites is about 1000 per cell. The role and the involvement of the OmpF porin (with colicins A and N) have been described in a recent study on the structural and functional interactions of a colicin‐resistant mutant of OmpF. The X‐ray crystal structure of the channel‐forming fragment of colicin A and that of the entire colicin la have provided the basis for biophysical and site‐directed muta‐genesis studies. Thanks to this powerful combination, it has been established that the interaction with the receptor in the outer membrane leads to a very substantial conformational change, as a result of which the N‐terminal domains of colicins interact with the lumen of the OmpF pore and then with the C‐terminal domain of TolA. A molten globular conformation of colicins probably constitutes the intermediate translocation/insertion competent state. Once the pore has formed, the polypeptide chain spans the whole cell envelope. Three distinct steps occur in the last stage of the process: (i) fast binding of the C‐terminal domain to the outer face of the cytoplasmic membrane; (ii) a slow insertion of the polypeptide chain into the outer face of the inner membrane in the absence of Δψ and (iii) a profound reorganization of the helix association, triggered by the transmembrane potential and resulting in the formation of the colicin channel.
Virus DNA packaging: the strategy used by phage λ
Authors: Catalano Carlos E. , Cue David , Feiss Michael 
Published: October 2006

SummaryPhage λ, like a number of other large DNA bacterio‐phages and the herpesviruses, produces concatemeric DNA during DNA replication. The concatemeric DNA is processed to produce unit‐length, virion DNA by cutting at specific sites along the concatemer. DNA cutting is coordinated with DNA packaging, the process of translocation of the cut DNA into the preformed capsid precursor, the prohead. A key player in the λ DNA packaging process is the phage‐encoded enzyme terminase, which is involved in (i) recognition of the concatemeric λ DNA; (ii) initiation of packaging, which includes the introduction of staggered nicks at cosN to generate the cohesive ends of virion DNA and the binding of the prohead; (iii) DNA packaging, possibly including the ATP‐driven DNA translocation; and (iv) following translocation, the cutting of the terminal cosN lo complete DNA packaging. To one side of cosN is the site cosB, which plays a role in the initiation of packaging; along with ATP, cosB stimulates the efficiency and adds fidelity to the endo‐nuclease activity of terminase in cutting cosN. cosB is essential for the formation of a post‐cleavage complex with terminase, complex I, that binds the prohead, forming a ternary assembly, complex II. Terminase interacts with cosN through its large subunit, gpA, and the small terminase subunit, gpNul, interacts with cosB. Packaging follows complex II formation. cosN is flanked on the other side by the site cosQ, which is needed for termination, but not initiation, of DNA packaging. cosQ is required for cutting of the second cosN, i.e. the cosN at which termination occurs. DNA packaging in λ has aspects that differ from other λ DNA transactions. Unlike the site‐specific recombination system of λ, for DNA packaging the initial site‐specific protein assemblage gives way to a mobile, translocating complete, and unlike the DNA replication system of λ, the same protein machinery is used for both initiation and translocation during λ DNA packaging.
Functional implications of the expression of PilC proteins in meningococci
Authors: Virji Mumtaz , Makepeace Katherine , Peak Ian , Payne Gail , Saunders Jon R. , Ferguson David J. P. , Moxon H. Richard 
Published: October 2006

SummaryMultiple forms of PilC were found in Neisseria meningitidis (Nm) strains isolated from the oropharynx, blood or cerebrospinal fluid expressing either Class I or Class II pili. PilC expression was observed less frequently in case as opposed to carrier isolates. Moreover, PilC and pili were not always co‐expressed. Several heavily piliated strains had no detectable PilC protein as determined by Western blotting using an antiserum previously used to detect such proteins in adhesive variants (Nassif et al., 1994). Serogroup B strain MC58 produced large numbers of pili, but expressed barely detectable amounts of PilC. A clonal variant of this strain with increased expression of PilC concurrently exhibited increased adherence to Chang conjunctival epithelial cells and human umbilical vein endothelial cells (Huvecs), but with more rapid binding to the former. No alteration in pilin sequence occurred in this variant, suggesting the involvement of PilC in increased adhesion. A Pil‐ backswitcher isolated from the hyper‐adherent variant was PilC+ but was non‐adherent, indicating that any PilC adherence function requires pilus expression. Parental variant (low PilC) produced pili in bundles that were easily detached from the bacterial surface and were frequently associated with Huvec surfaces after bacteria had been sheared off, but pili infrequently replaced bacteria during infection with the PilC‐expressing variant. The hyper‐adherent variant, which appeared to produce morphologically distinct pilus bundles, was able to withstand considerable shearing force and remained firmly attached to Huvecs. This raises the possibility that the observed hyper‐adherence may arise from better anchorage of pili to the bacterial surface in addition to increased adhesion to some host cell surfaces.
The Cryptococcus neoformans GAL7 gene and its use as an inducible promoter
Authors: Wickes Brian L. , Edman Jeffrey C. 
Published: October 2006

SummaryA Cryptococcus neoformans galactose auxotroph was created by ultraviolet light mutagenesis and complemented with a C. neoformans genomic library. The translated sequence of the complementing DNA revealed a high degree of simlarity to a number of UDP glucose‐D‐gatactose‐1‐phosphate uridylyitransferases. Expression of C. neoformans GAL7 mRNA followed a pattern similar to Saccharomyces cerevisiae expression; it was first observed within 2.5 min of induction and fully induced by 30 min. The gene was completely repressed in the presence of glucose. The GAL7 promoter was isolated and used to construct a promoter cassette. Two genes were tested in this cassette for galactose regulation by creating GAL7 promoter fusions with their coding regions. MFα, which encodes a pheromone, was found to produce filaments only in transformants that were induced by galactose. A second gene, β‐glucuronidase (gusA), which is a commonly used reporter gene, was tested and also found to be expressed. When the GAL7 p::GUS fusion was used to quantify inducibitity of the GAL7 promoter, the level of enzyme activity was at least 500‐fold greater for cells grown in galactose than for cells grown in glucose. The GAL7 promoter is the first inducible promoter characterized in C neoformans and the GUS gene is the first heterologous gene shown to be expressed in this yeast pathogen.
Regulation of ptsH and ptsl gene expression in Streptococcus salivarius ATCC 25975
Authors: Gagnon Guy , Vadeboncoeur Christian , Gauthier Lucie , Frenette Michel 
Published: October 2006

SummaryThe transcriptional regulation of the Streptococcus salivarius ptsH and ptsl genes coding for the general energy‐coupling proteins HPr and enzyme I of the phosphoenolpyruvate:sugar phosphotransferase system were investigated. These genes form an operon with the gene order ptsH–ptsl. Three distinct mRNA species were detected: a 0.5 kb transcript specific for ptsH, and two long transcripts (2.2 arid 2.4 kb) covering the whole pts operon. Transcription of all these mRNAs initiated at the same nucleotide located 9 bp downstream from a promoter located immediately upstream from the ptsH gene. The presence of a high‐energy stem–loop structure (T0) located at the begining of ptsl was responsible for the premature transcrrption termination generating the 0.5 kb ptsH‐specific transcript. The long transcripts ended in the poly(U) region of two rho‐independent‐like terminators (T1 and T2) at the 3′ end of ptsl. Studies with a 2‐deoxyglucose‐resistant spontaneous mutant of S. salivarius (L26) that produces an HPr–El fusion protein suggest that the regulation of HPr and El expression involves transcriptional as well as translational mechanisms.
A central domain of Rhizobium NodE protein mediates host specificity by determining the hydrophobicity of fatty acyl moieties of nodulation factors
Authors: Bloemberg Guido V. , Kamst Eric , Harteveld Marga , van der Drift Koen M. G. M. , Haverkamp Johan , Thomas‐Oates Jane E. , Lugtenberg Ben J. J. , Spaink Herman P. 
Published: October 2006

SummaryPreviously, we have shown that the nodE gene is a major determinant of the difference in host range between Rhizobium leguminosarum biovars viciae and trifolii. A new genetic test system for stringent functional analysis of nodE genes was constructed. By testing chimeric nodE genes constructed by the exchange of poiymerase chain reaction (PCR)‐generated restriction cassettes, we show that a central domain, containing only 44 non‐conserved amino acid residues, determines the host specificity of the NodE protein (401 amino acid residues). Mass spectrometric analysis of the lipo‐chitin oligosaccharides (LCOs) produced by the new test strain containing the biovar viciae nodE gene shows that molecules containing a polyunsaturated C18:4 (trans‐2. trans‐4. trans‐6. cis‐11‐octadecatetraenoic) fatty acyl moiety are produced, as is the case for wild‐type R. leguminosarum bv. viciae. The LCOs determined by the biovar trifolii nodE gene, which was overproduced in our test strain, carry C1 8:2 and C18:3 fatty acyl chains containing two or three conjugated trans double bonds, respectively. Therefore, the main difference between the nodE‐determined LCOs of biovar viviae and trifolii in this system is the presence or absence of one cis double bond, resulting in the very different hydrophobicity of the LCOs. Using a newly developed spot application assay, we show that the 18:2‐ and C18:3‐containing LCOs are able to induce the formation of nodule primordia on roots of Trifolium pratense. On the basis of these and other recent results, we propose that the host range of nodulation of the R. leguminosarum biovars viciae and trifolii is determined by the degree of hydrophobicity of the poly‐unsaturated fatty acyl moieties of their LCOs, which is mediated by the host‐specific central domain of the NodE protein.
Molecular characterization of the lincomycin‐production gene cluster of Streptomyces lincolnensis 78‐11
Authors: Peschke Ursula , Schmidt Heike , Zhang Hui‐Zhan , Piepersberg Wolfgang 
Published: October 2006

SummaryThe lincomycin (LM)‐production gene cluster of the overproducing strain Streptomyces iincolnensis 78‐11 was cloned, analysed by hybridization, as well as by DNA sequencing, and compared with the respective genome segments of other lincomycin producers. The lmb/lmr gene cluster is composed of 27 open reading frames with putative biosynthetic or regulatory functions (lmb genes) and three resistance (lmr) genes, two of which, lmrA and lmrC, flank the cluster. A very similar overall organization of the lmb/lmr cluster seems to be conserved in four other LM producers, although the clusters are embedded in non‐homologous genomic surroundings, in the wild‐type strain (S. lincolnensis NRRL2936), the lmb/lmr‐cluster apparently is present only in single copy. However, in the industrial strain S. lincolnensis 78‐11 the non‐adjacent gene clusters for the production of LM and melanin (melC) both are duplicated on a large (0.45‐0.5 Mb) fragment, accompanied by deletion events. This indicates that enhanced gene dosage is one of the factors for the overproduction of LM and demonstrates that large‐scale genome rearrangements can be a result of classical strain improvement by mutagenesis. Only a minority of the putative Lmb proteins belong to known protein families. These include members of the γ‐glutamyl transferases (LmbA), amino acid acylases (LmbC), aromatic amino acid aminotransferases (LmbF), imidazoleglycerolphosphate dehydratases (LmbK), dTDP‐glucose synthases (LmbO), dTDP‐glucose 4,6‐dehydratases (LmbM) and (NDP‐) ketohexose (or ketocyclitol) aminotransferases (LmbS). In contrast to earlier proposals on the biosynthetic pathway of the C‐8 sugar moiety (methylthiolincosaminide), this branch of the LM pathway actually seems to be based on nucleotide‐activated sugars as precursors.