bluuebunny/crossref_metadata_2025_split huggingface dataset:
Internal deformation of the human crystalline lens during accommodation
Authors: Weeber Henk A. , Van Der Heijde Rob G. L. 
Published: February 2008

Abstract.Purpose:  To describe the internal deformations in the crystalline lens that occur during accommodation.Methods:  A computer‐based mechanical model of accommodation was created using the finite element method. The lens geometry of the model was based on in vivo measurements of human lenses in the accommodated state. The mechanical properties of the lens were based on ex vivo measurements of human lenses. To achieve a state of disaccommodation, the lens equator was stretched by 7%. The internal strains and displacements were calculated for a young accommodating lens, a lens of pre‐presbyopic age and a lens of presbyopic age (20, 40 and 60 years old, respectively).Results:  The model showed that the radial strain was maximal in the nucleus for the young accommodating lens and minimal in the nucleus for the oldest non‐accommodating lens. In the young lens the deformations occurred throughout the entire lens, whereas in the older non‐accommodating lens the deformations were concentrated in the equatorial region.Conclusions:  The model predicted that during accommodation, changes in lens thickness are mainly caused by deformation of the nucleus. In the older, non‐accommodating lens, the deformations occur predominantly in the equatorial region and do not affect the central curvatures of the lens.
Identification of differentially expressed genes in spontaneously regressing melanoma using the MeLiM Swine Model
Authors: Rambow F. , Malek O. , Geffrotin C. , Leplat J.‐J. , Bouet S. , Piton G. , Hugot K. , Bevilacqua C. , Horak V. , Vincent‐Naulleau S. 
Published: February 2008

SummaryPartial and some few cases of complete spontaneous regression have been observed in cutaneous melanoma patients but little is known about the molecular mechanisms involved. The Melanoblastoma‐bearing Libechov Minipig (MeLiM) is a suitable animal model to study the phenomenon of spontaneous regression because MeLiM pigs exhibit naturally occurring melanomas which regress completely 6 months after birth. In this study, we used suppression subtractive hybridization (SSH) to identify molecular determinants of melanoma regression within swine melanoma tissues and melanoma cell cultures. Several markers involved in cell‐adhesion, ‐communication, ‐motility, signal transduction, negative regulation of cell proliferation, transport and immune response were identified that correlated with melanoma regression whereas the main genes involved in melanin synthesis showed a strong downregulation. For the most differentially expressed genes, we validated the results obtained by SSH with qRT‐PCR and with immunohistochemistry for some of them (CD9, MITF, RARRES1). Most notable, for the first time in melanoma, we identified the retinoic acid responder 1 gene (RARRES1) as a main actor of the regression process in melanoma. This first gene expression study in swine melanoma regression, may contribute to the finding of new therapeutic targets for human melanoma treatment.
Alignment of Genetic and Physical Maps of
Gibberella zeae
Authors: Lee Jungkwan , Jurgenson James E. , Leslie John F. , Bowden Robert L. 
Published: February 2008

ABSTRACT

We previously published a genetic map of
Gibberella zeae
(
Fusarium graminearum
sensu lato) based on a cross between Kansas strain Z-3639 (lineage 7) and Japanese strain R-5470 (lineage 6). In this study, that genetic map was aligned with the third assembly of the genomic sequence of
G. zeae
strain PH-1 (lineage 7) using seven structural genes and 108 sequenced amplified fragment length polymorphism markers. Several linkage groups were combined based on the alignments, the nine original linkage groups were reduced to six groups, and the total size of the genetic map was reduced from 1,286 to 1,140 centimorgans. Nine supercontigs, comprising 99.2% of the genomic sequence assembly, were anchored to the genetic map. Eight markers (four markers from each parent) were not found in the genome assembly, and four of these markers were closely linked, suggesting that >150 kb of DNA sequence is missing from the PH-1 genome assembly. The alignments of the linkage groups and supercontigs yielded four independent sets, which is consistent with the four chromosomes reported for this fungus. Two proposed heterozygous inversions were confirmed by the alignments; otherwise, the colinearity of the genetic and physical maps was high. Two of four regions with segregation distortion were explained by the two selectable markers employed in making the cross. The average recombination rates for each chromosome were similar to those previously reported for
G. zeae
. Despite an inferred history of genetic isolation of lineage 6 and lineage 7, the chromosomes of these lineages remain homologous and are capable of recombination along their entire lengths, even within the inversions. This genetic map can now be used in conjunction with the physical sequence to study phenotypes (e.g., fertility and fitness) and genetic features (e.g., centromeres and recombination frequency) that do not have a known molecular signature in the genome.
Inhibition of Species of the
Aspergillus
Section
Nigri
and Ochratoxin A Production in Grapes by Fusapyrone
Authors: Favilla Mara , Pascale Michelangelo , Ricelli Alessandra , Evidente Antonio , Amalfitano Carmine , Altomare Claudio 
Published: February 2008

ABSTRACT

Fusapyrone (FP), an antifungal natural compound, was tested against the three main ochratoxigenic species of the
Aspergillus
section
Nigri
. The MICs at 24 h were 6.0, 11.6, and 9.9 μg/ml for
Aspergillus carbonarius
,
Aspergillus tubingensis
, and
Aspergillus niger
, respectively. Strong inhibition of growth and morphological changes were still observed at half the MIC after 7 days. The application of a 100 μg/ml FP solution in a laboratory assay on artificially inoculated grapes resulted in a significant reduction (up to 6 orders of magnitude) of
A. carbonarius
CFU counts. Dramatic reductions of the ochratoxin A (OTA) content, compared to the content of the positive control (average amount of OTA, 112.5 ng/g of grape; three experiments), were obtained with the application of either 100 or 50 μg/ml of FP (0.6 or 5.1 ng/g of grape, respectively).
Differences in Carotenoid Composition among
Hymenobacter
and Related Strains Support a Tree-Like Model of Carotenoid Evolution
Authors: Klassen Jonathan L. , Foght Julia M. 
Published: February 2008

ABSTRACT

Carotenoids are structurally diverse pigments of biotechnological interest as natural colorants and in the prevention of human disease. The carotenoids present in 19 strains taxonomically related to the poorly described, nonphotosynthetic bacterial genus
Hymenobacter
, including 10 novel isolates cultivated from Victoria Upper Glacier, Antarctica, were characterized using high-performance liquid chromatography (HPLC). Nine chemically distinct carotenoids, present in various combinations irresolvable by conventional crude spectrophotometric analyses, were purified by preparative HPLC and characterized using UV-visible light absorption spectroscopy and high-resolution mass spectrometry. All major
Hymenobacter
carotenoids appear to be derived from a common backbone of 2′-hydroxyflexixanthin and include previously unreported presumptive hexosyl, pentosyl, and methyl derivatives. Their distribution does not, however, correlate perfectly with 16S rRNA gene phylogeny. Carotenoid composition, therefore, may be strain specific and does not follow a strictly homogeneous pattern of vertical evolutionary descent.
Markerless Multiple-Gene-Deletion System for
Streptococcus mutans
Authors: Banerjee Anirban , Biswas Indranil 
Published: February 2008

ABSTRACT

Inactivation or selective modification is essential to elucidate the putative function of a gene. The present study describes an improved Cre-
loxP
-based method for markerless multiple gene deletion in
Streptococcus mutans
, the principal etiological agent of dental caries. This modified method uses two mutant
loxP
sites, which after recombination creates a double-mutant
loxP
site that is poorly recognized by Cre recombinase, facilitating multiple gene deletions in a single genetic background. The effectiveness of this modified strategy was demonstrated by the construction of both single and double gene deletions at the
htrA
and
clpP
loci on the chromosome of
Streptococcus mutans
. HtrA and ClpP play key roles in the processing and maturation of several important proteins, including many virulence factors. Deletion of these genes resulted in reducing the organism's ability to withstand exposure to low pH and oxidative agents. The method described here is simple and efficient and can be easily implemented for multiple gene deletions with
S. mutans
and other streptococci.
Solvent Stress Response of the Denitrifying Bacterium “
Aromatoleum aromaticum
” Strain EbN1
Authors: Trautwein Kathleen , Kühner Simon , Wöhlbrand Lars , Halder Thomas , Kuchta Kenny , Steinbüchel Alexander , Rabus Ralf 
Published: February 2008

ABSTRACT

The denitrifying betaproteobacterium “
Aromatoleum aromaticum
” strain EbN1 degrades several aromatic compounds, including ethylbenzene, toluene,
p
-cresol, and phenol, under anoxic conditions. The hydrophobicity of these aromatic solvents determines their toxic properties. Here, we investigated the response of strain EbN1 to aromatic substrates at semi-inhibitory (about 50% growth inhibition) concentrations under two different conditions: first, during anaerobic growth with ethylbenzene (0.32 mM) or toluene (0.74 mM); and second, when anaerobic succinate-utilizing cultures were shocked with ethylbenzene (0.5 mM), toluene (1.2 mM),
p
-cresol (3.0 mM), and phenol (6.5 mM) as single stressors or as a mixture (total solvent concentration, 2.7 mM). Under all tested conditions impaired growth was paralleled by decelerated nitrate-nitrite consumption. Additionally, alkylbenzene-utilizing cultures accumulated poly(3-hydroxybutyrate) (PHB) up to 10% of the cell dry weight. These physiological responses were also reflected on the proteomic level (as determined by two-dimensional difference gel electrophoresis), e.g., up-regulation of PHB granule-associated phasins, cytochrome

cd
1

nitrite reductase of denitrification, and several proteins involved in oxidative (e.g., SodB) and general (e.g., ClpB) stress responses.
Codon-Optimized Bacterial Genes Improve
l
-Arabinose Fermentation in Recombinant
Saccharomyces cerevisiae
Authors: Wiedemann Beate , Boles Eckhard 
Published: February 2008

ABSTRACT

Bioethanol produced by microbial fermentations of plant biomass hydrolysates consisting of hexose and pentose mixtures is an excellent alternative to fossil transportation fuels. However, the yeast
Saccharomyces cerevisiae
, commonly used in bioethanol production, can utilize pentose sugars like
l
-arabinose or
d
-xylose only after heterologous expression of corresponding metabolic pathways from other organisms. Here we report the improvement of a bacterial
l
-arabinose utilization pathway consisting of
l
-arabinose isomerase from
Bacillus subtilis
and
l
-ribulokinase and
l
-ribulose-5-P 4-epimerase from
Escherichia coli
after expression of the corresponding genes in
S. cerevisiae
.
l
-Arabinose isomerase from
B. subtilis
turned out to be the limiting step for growth on
l
-arabinose as the sole carbon source. The corresponding enzyme could be effectively replaced by the enzyme from
Bacillus licheniformis
, leading to a considerably decreased lag phase. Subsequently, the codon usage of all the genes involved in the
l
-arabinose pathway was adapted to that of the highly expressed genes encoding glycolytic enzymes in
S. cerevisiae
. Yeast transformants expressing the codon-optimized genes showed strongly improved
l
-arabinose conversion rates. With this rational approach, the ethanol production rate from
l
-arabinose could be increased more than 2.5-fold from 0.014 g ethanol h
−1
(g dry weight)
−1
to 0.036 g ethanol h
−1
(g dry weight)
−1
and the ethanol yield could be increased from 0.24 g ethanol (g consumed
l
-arabinose)
−1
to 0.39 g ethanol (g consumed
l
-arabinose)
−1
. These improvements make up a new starting point for the construction of more-efficient industrial
l
-arabinose-fermenting yeast strains by evolutionary engineering.
Cultivation-Independent Characterization of
Methylobacterium
Populations in the Plant Phyllosphere by Automated Ribosomal Intergenic Spacer Analysis
Authors: Knief Claudia , Frances Lisa , Cantet Franck , Vorholt Julia A. 
Published: February 2008

ABSTRACT

Bacteria of the genus
Methylobacterium
are widespread in the environment, but their ecological role in ecosystems, such as the plant phyllosphere, is not very well understood. To gain better insight into the distribution of different
Methylobacterium
species in diverse ecosystems, a rapid and specific cultivation-independent method for detection of these organisms and analysis of their community structure is needed. Therefore, 16S rRNA gene-targeted primers specific for this genus were designed and evaluated. These primers were used in PCR in combination with a reverse primer that binds to the tRNA
Ala
gene, which is located upstream of the 23S rRNA gene in the 16S-23S intergenic spacer (IGS). PCR products that were of different lengths were obtained due to the length heterogeneity of the IGS of different
Methylobacterium
species. This length variation allowed generation of fingerprints of
Methylobacterium
communities in environmental samples by automated ribosomal intergenic spacer analysis. The
Methylobacterium
communities on leaves of different plant species in a natural field were compared using this method. The new method allows rapid comparisons of
Methylobacterium
communities and is thus a useful tool to study
Methylobacterium
communities in different ecosystems.
Solid-Phase Capture of Pathogenic Bacteria by Using Gangliosides and Detection with Real-Time PCR
Authors: Desai Prerak T. , Walsh Marie K. , Weimer Bart C. 
Published: February 2008

ABSTRACT
We developed a method for concentrating pathogens from samples without enrichment. Immobilized gangliosides concentrated bacteria for detection with real-time PCR. A sensitivity of ∼4 CFU/ml (3 h) in samples without competing microflora was achieved. Samples with competing microflora had a sensitivity of 40,000 CFU/ml. The variance was less than one cycle.
Heterologous Expression of theTreponema pallidumLaminin-Binding Adhesin Tp0751 in the Culturable SpirocheteTreponema phagedenis
Authors: Cameron Caroline E. , Kuroiwa Janelle M. Y. , Yamada Mitsunori , Francescutti Teresa , Chi Bo , Kuramitsu Howard K. 
Published: February 2008

ABSTRACTTreponema pallidumsubsp.pallidum, the causative agent of syphilis, is an unculturable, genetically intractable bacterium. Here we report the use of the shuttle vector pKMR4PEMCS for the expression of a previously identifiedT.pallidumlaminin-binding adhesin, Tp0751, in the nonadherent, culturable spirocheteTreponema phagedenis. Heterologous expression of Tp0751 inT.phagedeniswas confirmed via reverse transcriptase PCR analysis with tp0751 gene-specific primers and immunofluorescence analysis with Tp0751-specific antibodies; the latter assay verified the expression of the laminin-binding adhesin on the treponemal surface. Expression of Tp0751 withinT.phagedeniswas functionally confirmed via laminin attachment assays, in which heterologous Tp0751 expression conferred uponT.phagedenisthe capacity to attach to laminin. Further, specific inhibition of the attachment ofT.phagedenisheterologously expressing Tp0751 to laminin was achieved by using purified antibodies raised against recombinantT.pallidumTp0751. This is the first report of heterologous expression of a gene from an unculturable treponeme inT.phagedenis. This novel methodology will significantly advance the field of syphilis research by allowing targeted investigations ofT.pallidumproteins purported to play a role in pathogenesis, and specifically host cell attachment, in the nonadherent spirocheteT.phagedenis.
Naturally Occurring Nonpathogenic Isolates of the Plant Pathogen
Pseudomonas syringae
Lack a Type III Secretion System and Effector Gene Orthologues
Authors: Mohr Toni J. , Liu Haijie , Yan Shuangchun , Morris Cindy E. , Castillo José A. , Jelenska Joanna , Vinatzer Boris A. 
Published: February 2008

ABSTRACT

Pseudomonas syringae
causes plant diseases, and the main virulence mechanism is a type III secretion system (T3SS) that translocates dozens of effector proteins into plant cells. Here we report the existence of a subgroup of
P. syringae
isolates that do not cause disease on any plant species tested. This group is monophyletic and most likely evolved from a pathogenic
P. syringae
ancestor through loss of the T3SS. In the nonpathogenic isolate
P. syringae
508 the genomic region that in pathogenic
P. syringae
strains contains the
hrp-hrc
cluster coding for the T3SS and flanking effector genes is absent.
P. syringae
508 was also surveyed for the presence of effector orthologues from the closely related pathogenic strain
P. syringae
pv. syringae B728a, but none were detected. The absence of the
hrp-hrc
cluster and effector orthologues was confirmed for other nonpathogenic isolates. Using the AvrRpt2 effector as reporter revealed the inability of
P. syringae
508 to translocate effectors into plant cells. Adding a plasmid-encoded T3SS and the
P. syringae
pv. syringae 61 effector gene
hopA1
increased in planta growth almost 10-fold. This suggests that
P. syringae
508 supplemented with a T3SS could be used to determine functions of individual effectors in the context of a plant infection, avoiding the confounding effect of other effectors with similar functions present in effector mutants of pathogenic isolates.
A Novel Chromate Reductase from
Thermus scotoductus
SA-01 Related to Old Yellow Enzyme
Authors: Opperman Diederik Johannes , Piater Lizelle Ann , van Heerden Esta 
Published: February 2008

ABSTRACT

Bacteria can reduce toxic and carcinogenic Cr(VI) to insoluble and less toxic Cr(III).
Thermus scotoductus
SA-01, a South African gold mine isolate, has been shown to be able to reduce a variety of metals, including Cr(VI). Here we report the purification to homogeneity and characterization of a novel chromate reductase. The oxidoreductase is a homodimeric protein, with a monomer molecular mass of approximately 36 kDa, containing a noncovalently bound flavin mononucleotide cofactor. The chromate reductase is optimally active at a pH of 6.3 and at 65°C and requires Ca
2+
or Mg
2+
for activity. Enzyme activity was also dependent on NADH or NADPH, with a preference for NADPH, coupling the oxidation of approximately 2 and 1.5 mol NAD(P)H to the reduction of 1 mol Cr(VI) under aerobic and anaerobic conditions, respectively. The

K
m

values for Cr(VI) reduction were 3.5 and 8.4 μM for utilizing NADH and NADPH as electron donors, respectively, with corresponding
V
max
values of 6.2 and 16.0 μmol min
−1
mg
−1
. The catalytic efficiency (
k
cat
/

K
m

) of chromate reduction was 1.14 × 10
6
M
−1
s
−1
, which was >50-fold more efficient than that of the quinone reductases and >180-fold more efficient than that of the nitroreductases able to reduce Cr(VI). The chromate reductase was identified to be encoded by an open reading frame of 1,050 bp, encoding a single protein of 38 kDa under the regulation of an
Escherichia coli
σ
70
-like promoter. Sequence analysis shows the chromate reductase to be related to the old yellow enzyme family, in particular the xenobiotic reductases involved in the oxidative stress response.
Secretome Analysis Uncovers an Hcp-Family Protein Secreted via a Type VI Secretion System inAgrobacterium tumefaciens
Authors: Wu Hung-Yi , Chung Pei-Che , Shih Hsiao-Wei , Wen Sy-Ray , Lai Erh-Min 
Published: February 2008

ABSTRACTAgrobacterium tumefaciensis a plant-pathogenic bacterium capable of secreting several virulence factors into extracellular space or the host cell. In this study, we used shotgun proteomics analysis to investigate the secretome ofA. tumefaciens, which resulted in identification of 12 proteins, including 1 known secretory protein (VirB1*) and 11 potential secretory proteins. Interestingly, one unknown protein, which we designated hemolysin-coregulated protein (Hcp), is a predicted soluble protein without a recognizable N-terminal signal peptide. Western blot analysis revealed thatA. tumefaciensHcp is expressed and secreted when cells are grown in both minimal and rich media. Further biochemical and immunoelectron microscopy analysis demonstrated that intracellular Hcp is localized mainly in the cytosol, with a small portion in the membrane system. To investigate the mechanism of secretion of Hcp inA. tumefaciens, we generated mutants with deletions of a conserved gene,icmF, or the entire putative operon encoding a recently identified type VI secretion system (T6SS). Western blot analysis indicated that Hcp was expressed but not secreted into the culture medium in mutants with deletions oficmFor the t6ss operon. The secretion deficiency of Hcp in theicmFmutant was complemented by heterologoustransexpression oficmF, suggesting thaticmFis required for Hcp secretion. In tumor assays with potato tuber disks, deletion ofhcpresulted in approximately 20 to 30% reductions in tumorigenesis efficiency, while no consistent difference was observed whenicmFor the t6ss operon was deleted. These results increase our understanding of the conserved T6SS used by both plant- and animal-pathogenic bacteria.
Basis of Arginine Sensitivity of Microbial
N
-Acetyl-
l
-Glutamate Kinases: Mutagenesis and Protein Engineering Study with the
Pseudomonas aeruginosa
and
Escherichia coli
Enzymes
Authors: Fernández-Murga M. Leonor , Rubio Vicente 
Published: February 2008

ABSTRACT

N
-Acetylglutamate kinase (NAGK) catalyzes the second step of arginine biosynthesis. In
Pseudomonas aeruginosa
, but not in
Escherichia coli
, this step is rate limiting and feedback and sigmoidally inhibited by arginine. Crystal structures revealed that arginine-insensitive
E. coli
NAGK (EcNAGK) is homodimeric, whereas arginine-inhibitable NAGKs, including
P. aeruginosa
NAGK (PaNAGK), are hexamers in which an extra N-terminal kinked helix (N-helix) interlinks three dimers. By introducing single amino acid replacements in PaNAGK, we prove the functionality of the structurally identified arginine site, as arginine site mutations selectively decreased the apparent affinity for arginine. N-helix mutations affecting R24 and E17 increased and decreased, respectively, the apparent affinity of PaNAGK for arginine, as predicted from enzyme structures that revealed the respective formation by these residues of bonds favoring inaccessible and accessible arginine site conformations. N-helix N-terminal deletions spanning ≥16 residues dissociated PaNAGK to active dimers, those of ≤20 residues decreased the apparent affinity for arginine, and complete N-helix deletion (26 residues) abolished arginine inhibition. Upon attachment of the PaNAGK N-terminal extension to the EcNAGK N terminus, EcNAGK remained dimeric and arginine insensitive. We concluded that the N-helix and its C-terminal portion after the kink are essential but not sufficient for hexamer formation and arginine inhibition, respectively; that the N-helix modulates NAGK affinity for arginine and mediates signal transmission between arginine sites, thus establishing sigmoidal arginine inhibition kinetics; that the mobile αH-β16 loop of the arginine site is the modulatory signal receiver; and that the hexameric architecture is not essential for arginine inhibition but is functionally essential for physiologically relevant arginine control of NAGK.