bluuebunny/crossref_metadata_2025_split huggingface dataset:
General Stress Transcription Factor ς
B
and Its Role in Acid Tolerance and Virulence of
Listeria monocytogenes
Authors: Wiedmann Martin , Arvik Torey J. , Hurley Richard J. , Boor Kathryn J. 
Published: December 2019

ABSTRACT

The gene encoding the general stress transcription factor ς
B
in the gram-positive bacterium
Listeria monocytogenes
was isolated with degenerate PCR primers followed by inverse PCR amplification. Evidence for gene identification includes the following: (i) phylogenetic analyses of reported amino acid sequences for ς
B
and the closely related ς
F
proteins grouped
L. monocytogenes
ς
B
in the same cluster with the ς
B
proteins from
Bacillus subtilis
and
Staphylococcus aureus
, (ii) the gene order in the 2,668-bp portion of the
L. monocytogenes sigB
operon is
rsbU-rsbV-rsbW-sigB-rsbX
and is therefore identical to the order of the last five genes of the
B. subtilis sigB
operon, and (iii) an
L. monocytogenes
ς
B
mutant had reduced resistance to acid stress in comparison with its isogenic parent strain. The
sigB
mutant was further characterized in mouse models of listeriosis by determining recovery rates of the wild-type and mutant strains from livers and spleens following intragastric or intraperitoneal infection. Our results suggest that ς
B
-directed genes do not appear to be essential for the spread of
L. monocytogenes
to mouse liver or spleen at 2 and 4 days following intragastric or intraperitoneal infection.
ftsE
(Ts) Affects Translocation of K
+
-Pump Proteins into the Cytoplasmic Membrane of
Escherichia coli
Authors: Ukai Hideki , Matsuzawa Hiroshi , Ito Koreaki , Yamada Mamoru , Nishimura Akiko 
Published: December 2019

ABSTRACT

The
ftsE
(Ts) mutation of
Escherichia coli
causes defects in cell division and cell growth. We expressed alkaline phosphatase (PhoA) fusion proteins of KdpA, Kup, and TrkH, all of which proved functional in vivo as K
+
ion pumps, in the mutant cells. During growth at 41°C, these proteins were progressively lost from the membrane fraction. The reduction in the abundance of these proteins inversely correlated with cell growth, but the preformed proteins in the membrane were stable at 41°C, indicating that the molecules synthesized at the permissive temperature were diluted in a growth-dependent manner at a high temperature. Pulse-chase experiments showed that KdpA-PhoA was synthesized, but the synthesized protein did not translocate into the membrane of the ftsE(Ts) cells at 41°C and degraded very rapidly. The loss of KdpA-PhoA from the membrane fractions of ftsE(Ts) cells was suppressed by a multicopy plasmid carrying the
ftsE
+
gene. While cell growth stopped when the abundance of these proteins decreased 15-fold, the addition of a high concentration of K
+
ions specifically alleviated the growth defect of ftsE(Ts) cells but not cell division, and the cells elongated more than 100-fold. We conclude that one of the causes of growth cessation in the ftsE(Ts) mutants is a defect in the translocation of K
+
-pump proteins into the cytoplasmic membrane.
The Regulated Outer Membrane Protein Omp21 from
Comamonas acidovorans
Is Identified as a Member of a New Family of Eight-Stranded β-Sheet Proteins by Its Sequence and Properties
Authors: Baldermann C. , Lupas A. , Lubieniecki J. , Engelhardt H. 
Published: December 2019

ABSTRACT

Omp21, a minor outer membrane protein of the soil bacterium
Comamonas acidovorans
, was purified from a spontaneous mutant lacking a surface layer and long-chain lipopolysaccharide. Omp21 synthesis is enhanced by oxygen depletion, and the protein has a variable electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gel electrophoresis due to its heat-modifiable behavior. The structural gene
omp21
encodes a precursor of 204 amino acids with a putative signal peptide of 21 amino acids. Mature Omp21 is a typical outer membrane protein with a high content of β structure as determined by infrared spectroscopy. Sequence comparisons show that it belongs to a new outer membrane protein family, characterized by eight amphipathic β strands, which includes virulence proteins, such as the neisserial opacity proteins,
Salmonella typhimurium
Rck, and
Yersinia enterocolitica
Ail, as well as the major outer membrane proteins OmpA from
Escherichia coli
and OprF from
Pseudomonas aeruginosa
.
Escherichia coli
Strains Lacking Protein HU Are UV Sensitive due to a Role for HU in Homologous Recombination
Authors: Li Shisheng , Waters Raymond 
Published: December 2019

ABSTRACT

hupA
and
hupB
encode the α and β subunits of the
Escherichia coli
histone-like protein HU. Here we show that
E. coli hup
mutants are sensitive to UV in the

rec
+
sbc

+
,
recBC sbcA
,
recBC sbcBC
,
umuDC
,
recF
, and
recD
backgrounds. However,
hupAB
mutations do not enhance the UV sensitivity of resolvase-deficient
recG ruvA
strains.
hupAB uvrA
and
hupAB recG
strains are supersensitive to UV.
hup
mutations enhance the UV sensitivity of
ruvA
strains to a much lesser extent but enhance that of
rus-1 ruvA
strains to the same extent as for

rus
+
ruv

+
strains. Our results suggest that HU plays a role in recombinational DNA repair that is not specifically limited to double-strand break repair or daughter strand gap repair; the lack of HU affects the RecG RusA and RuvABC pathways for Holliday junction processing equally if the two pathways are equally active in recombinational repair; the function of HU is not in the substrate processing step or in the RecFOR-directed synapsis action during recombinational repair. Furthermore, the UV sensitivity of
hup
mutants cannot be suppressed by overexpression of wild-type or mutant
gyrB
, which confers novobiocin resistance, or by different concentrations of a gyrase inhibitor that can increase or decrease the supercoiling of chromosomal DNA.
Lack of Consistent Short Sequence Repeat Polymorphisms in Genetically Homologous Colonizing and Invasive
Candida albicans
Strains
Authors: Lunel Frans Verduyn , Licciardello Lidia , Stefani Stefania , Verbrugh Henri A. , Melchers Willem J. G. , Meis Jacques F. G. M. , Scherer Stewart , van Belkum Alex 
Published: December 2019

ABSTRACT

Short sequence repeats (SSRs), potentially representing variable numbers of tandem repeat (VNTR) loci, were identified for the human-pathogenic yeast species
Candida albicans
by computerized DNA sequence scanning. The individual SSR regions were investigated in different clinical isolates of
C. albicans
. Most of the
C. albicans
SSRs were identified as genuine VNTRs. They appeared to be present in multiple allelic variants and were demonstrated to be diverse in length among nonrelated strains. As such, these loci provide adequate targets for the molecular typing of
C. albicans
strains. VNTRs encountered in other microbial species sometimes participate in regulation of gene expression and function as molecular switches at the transcriptional or translational level. Interestingly, the VNTRs identified here often encode polyglutamine stretches and are frequently located within genes potentially involved in the regulation of transcription. DNA sequencing of these VNTRs demonstrated that the length variability was restricted to the CAA/CAG repeats encoding the polyglutamine stretches. For these reasons, paired
C. albicans
isolates of similar genotype, either found as noninvasive colonizers or encountered in an invasive state in the same individual, were studied with respect to potentially invasion-related alterations in the VNTR profiles. However, none of the VNTRs analyzed thus far varied systematically with the transition from colonization to invasion. In contrast to the situation described for some prokaryotic species, this finding suggests that VNTRs of
C. albicans
may not simply function as contingency loci related to straightforward on/off regulation of invasion-related gene expression.
Disruption of the
Candida albicans TPS1
Gene Encoding Trehalose-6-Phosphate Synthase Impairs Formation of Hyphae and Decreases Infectivity
Authors: Zaragoza Oscar , Blazquez Miguel A. , Gancedo Carlos 
Published: December 2019

ABSTRACT

The
TPS1
gene from
Candida albicans
, which encodes trehalose-6-phosphate synthase, has been cloned by functional complementation of a
tps1
mutant from
Saccharomyces cerevisiae
. In contrast with the wild-type strain, the double
tps1/tps1
disruptant did not accumulate trehalose at stationary phase or after heat shock. Growth of the
tps1/tps1
disruptant at 30°C was indistinguishable from that of the wild type. However, at 42°C it did not grow on glucose or fructose but grew normally on galactose or glycerol. At 37°C, the yeast-hypha transition in the mutant in glucose-calf serum medium did not occur. During growth at 42°C, the mutant did not form hyphae in galactose or in glycerol. Some of the growth defects observed may be traced to an unbalanced sugar metabolism that reduces the cellular content of ATP. Mice inoculated with 10
6
CFU of the
tps1/tps1
mutant did not show visible symptoms of infection 16 days after inoculation, while those similarly inoculated with wild-type cells were dead 12 days after inoculation.
Physical and Genetic Map of the Obligate Intracellular Bacterium
Coxiella burnetii
Authors: Willems H. , Jäger Cornelie , Baljer Georg 
Published: December 2019

ABSTRACT

Pulsed-field gel electrophoresis and PCR techniques have been used to construct a
Not
I macrorestriction map of the obligate intracellular bacterium
Coxiella burnetii
Nine Mile. The size of the chromosome has been determined to be 2,103 kb comprising 29
Not
I restriction fragments. The average resolution is 72.5 kb, or about 3.5% of the genome. Experimental data support the presence of a linear chromosome. Published genes were localized on the physical map by Southern hybridization. One gene, recognized as transposable element, was found to be present in at least nine sites evenly distributed over the whole chromosome. There is only one copy of a 16S rRNA gene. The putative
oriC
has been located on a 27.5-kb
Not
I fragment. Gene organization upstream the
oriC
is almost identical to that of
Pseudomonas putida
and
Bacillus subtilis
, whereas gene organization downstream the
oriC
seems to be unique among bacteria. The physical map will be helpful in investigations of the great heterogeneity in restriction fragment length polymorphism patterns of different isolates and the great variation in genome size. The genetic map will help to determine whether gene order in different isolates is conserved.
Purification and Characterization of EDTA Monooxygenase from the EDTA-Degrading Bacterium BNC1
Authors: Payne Jason W. , Bolton Harvey , Campbell James A. , Xun Luying 
Published: December 2019

ABSTRACT

The synthetic chelating agent EDTA can mobilize radionuclides and heavy metals in the environment. Biodegradation of EDTA should reduce this mobilization. Although several bacteria have been reported to mineralize EDTA, little is known about the biochemistry of EDTA degradation. Understanding the biochemistry will facilitate the removal of EDTA from the environment. EDTA-degrading activities were detected in cell extracts of bacterium BNC1 when flavin mononucleotide (FMN), NADH, and O
2
were present. The degradative enzyme system was separated into two different enzymes, EDTA monooxygenase and an FMN reductase. EDTA monooxygenase oxidized EDTA to glyoxylate and ethylenediaminetriacetate (ED3A), with the coconsumption of FMNH
2
and O
2
. The FMN reductase provided EDTA monooxygenase with FMNH
2
by reducing FMN with NADH. The FMN reductase was successfully substituted in the assay mixture by other FMN reductases. EDTA monooxygenase was purified to greater than 95% homogeneity and had a single polypeptide with a molecular weight of 45,000. The enzyme oxidized both EDTA complexed with various metal ions and uncomplexed EDTA. The optimal conditions for activity were pH 7.8 and 35°C.

K
m

s were 34.1 μM for uncomplexed EDTA and 8.5 μM for MgEDTA
2−
; this difference in

K
m

indicates that the enzyme has greater affinity for MgEDTA
2−
. The enzyme also catalyzed the release of glyoxylate from nitrilotriacetate and diethylenetriaminepentaacetate. EDTA monooxygenase belongs to a small group of FMNH
2
-utilizing monooxygenases that attack carbon-nitrogen, carbon-sulfur, and carbon-carbon double bonds.
Regulation of Porin-Mediated Outer Membrane Permeability by Nutrient Limitation in
Escherichia coli
Authors: Liu XueQiao , Ferenci Thomas 
Published: December 2019

ABSTRACT

OmpF and OmpC porins were differentially regulated by nutrient limitation and growth rate in glucose- or nitrogen-limited chemostat cultures of
Escherichia coli
. Transcriptional and translational
ompF
fusions showed a sharp peak of expression under glucose limitation at
D
= 0.3 h
−1
, with lower amounts at lower and higher growth rates. The peak of OmpR-dependent transcriptional stimulation of
ompF
under glucose limitation in minimal salts media was about 20-fold above nutrient excess levels and 3-fold higher than that achieved with low osmolarity. Analysis of outer membrane protein levels and results of growth competition experiments with porin mutants were consistent with the enhanced role of OmpF under glucose limitation, but not N limitation. In contrast, OmpC was the major porin under N limitation but was increasingly expressed under glucose limitation at very low growth rates approaching starvation, when OmpF was downregulated. In summary, outer membrane permeability under N-limited, sugar-rich conditions is largely based on OmpC, whereas porin activity is a complex, highly sensitive function of OmpF, OmpC, and LamB glycoporin expression under different levels of glucose limitation. Indeed, the OmpF level was more responsive to nutrient limitation than to medium osmolarity and suggested a significant additional layer of control over the porin-regulatory network.
Effect of Oxygen on Translation and Posttranslational Steps in Expression of Photosynthesis Genes in
Rhodobacter capsulatus
Authors: Hebermehl Markus , Klug Gabriele 
Published: December 2019

ABSTRACT

The formation of the photosynthetic apparatus in
Rhodobacter capsulatus
is regulated by oxygen tension. Previous studies have shown a regulatory effect of oxygen on the transcription of photosynthesis genes and on the stability of certain mRNA segments. Here we show that oxygen affects
puf
and
puc
gene expression posttranslationally and that this regulation depends on the presence of bacteriochlorophyll. Our data suggest that this posttranslational effect of oxygen on
puf
and
puc
expression is due to the primary effect of oxygen on bacteriochlorophyll synthesis or assembly of pigment protein complexes. Oxygen does not affect the rates of translation of
puf
-encoded proteins.
Binding Interaction between Tet(M) and the Ribosome: Requirements for Binding
Authors: Dantley Kathi A. , Dannelly H. Kathleen , Burdett Vickers 
Published: December 2019

ABSTRACT
Tet(M) protein interacts with the protein biosynthesis machinery to render this process resistant to tetracycline by a mechanism which involves release of the antibiotic from the ribosome in a reaction dependent on GTP hydrolysis. To clarify this resistance mechanism further, the interaction of Tet(M) with the ribosome has been examined by using a gel filtration assay with radioactively labelled Tet(M) protein. The presence of GTP and 5′-guanylyl imido diphosphate, but not GDP, promoted Tet(M)-ribosome complex formation. Furthermore, thiostrepton, which inhibits the activities of elongation factor G (EF-G) and EF-Tu by binding to the ribosome, blocks stable Tet(M)-ribosome complex formation. Direct competition experiments show that Tet(M) and EF-G bind to overlapping sites on the ribosome.
Rhs
Elements Comprise Three Subfamilies Which Diverged Prior to Acquisition by
Escherichia coli
Authors: Wang Yong-Dong , Zhao Sheng , Hill Charles W. 
Published: December 2019

ABSTRACT

The
Rhs
elements are complex genetic composites widely spread among
Escherichia coli
isolates. One of their components, a 3.7-kb, GC-rich core, maintains a single open reading frame that extends the full length of the core and then 400 to 600 bp beyond into an AT-rich region. Whereas
Rhs
cores are homologous, core extensions from different elements are dissimilar. Two new
Rhs
elements from strains of the ECOR reference collection have been characterized.
RhsG
(from strain ECOR-11) maps to min 5.3, and
RhsH
(from strain ECOR-45) maps to min 32.8, where it lies in tandem with
RhsE
. Comparison of strain K-12 to ECOR-11 indicates that
RhsG
was once present in but has been largely deleted from an ancestor of K-12. Phylogenetic analysis shows that the cores from eight known elements fall into three subfamilies,
RhsA-B-C-F
,
RhsD-E
, and
RhsG-H
. Cores from different subfamilies diverge 22 to 29%. Analysis of substitutions that distinguish between subfamilies shows that the origin of the ancestral core as well as the process of subfamily separation occurred in a GC-rich background. Furthermore, each subfamily independently passed from the GC-rich background to a less GC-rich background such as
E. coli
. A new example of core-extension shuffling provides the first example of exchange between cores of different subfamilies. A novel component of
RhsE
and
RhsG
,
vgr
, encodes a large protein distinguished by 18 to 19 repetitions of a Val-Gly dipeptide occurring with a eight-residue periodicity.
TherelA/spoT-Homologous Gene inStreptomyces coelicolorEncodes Both Ribosome-Dependent (p)ppGppSynthesizing and -Degrading Activities
Authors: Martínez-Costa Oscar H. , Fernández-Moreno Miguel A. , Malpartida Francisco 
Published: December 2019

ABSTRACTStreptomyces coelicolor(p)ppGpp synthetase (Rel protein) belongs to the RelA and SpoT (RelA/SpoT) family, which is involved in (p)ppGpp metabolism and the stringent response. The potential functions of therelgene have been examined.S. coelicolorRel has been shown to be ribosome associated, and its activity in vitro is ribosome dependent. Analysis in vivo of the active recombinant protein in well-definedEscherichia coli relAandrelA/spoTmutants provides evidence thatS. coelicolorRel, like nativeE. coliRelA, is functionally ribosome associated, resulting in ribosome-dependent (p)ppGpp accumulation upon amino acid deprivation. Expression of anS. coelicolorC-terminally deleted Rel, comprised of only the first 489 amino acids, catalyzes a ribosome-independent (p)ppGpp formation, in the same manner as theE. colitruncated RelA protein (1 to 455 amino acids). AnE. coli relA spoTdouble deletion mutant transformed withS. coelicolor relgene suppresses the phenotype associated with (p)ppGpp deficiency. However, in such a strain, arel-mediated (p)ppGpp response apparently occurs after glucose depletion, but only in the absence of amino acids. Analysis of ppGpp decay inE. coliexpressing theS. coelicolor relgene suggests that it also encodes a (p)ppGpp-degrading activity. By deletion analysis, the catalytic domains ofS. coelicolorRel for (p)ppGpp synthesis and degradation have been located within its N terminus (amino acids 267 to 453 and 93 to 397, respectively). In addition,E. coli relAin anS. coelicolor reldeletion mutant restores actinorhodine production and shows a nearly normal morphological differentiation, as does the wild-typerelgene, which is in agreement with the proposed role of (p)ppGpp nucleotides in antibiotic biosynthesis.
Influence of the Secondary Cell Wall Polymer on the Reassembly, Recrystallization, and Stability Properties of the S-Layer Protein from
Bacillus stearothermophilus
PV72/p2
Authors: Sára Margit , Dekitsch Christine , Mayer Harald F. , Egelseer Eva M. , Sleytr Uwe B. 
Published: December 2019

ABSTRACT

The high-molecular-weight secondary cell wall polymer (SCWP) from
Bacillus stearothermophilus
PV72/p2 is mainly composed of
N
-acetylglucosamine (GlcNAc) and
N
-acetylmannosamine (ManNAc) and is involved in anchoring the S-layer protein via its N-terminal region to the rigid cell wall layer. In addition to this binding function, the SCWP was found to inhibit the formation of self-assembly products during dialysis of the guanidine hydrochloride (GHCl)-extracted S-layer protein. The degree of assembly (DA; percent assembled from total S-layer protein) that could be achieved strongly depended on the amount of SCWP added to the GHCl-extracted S-layer protein and decreased from 90 to 10% when the concentration of the SCWP was increased from 10 to 120 μg/mg of S-layer protein. The SCWP kept the S-layer protein in the water-soluble state and favored its recrystallization on solid supports such as poly-
l
-lysine-coated electron microscopy grids. Derived from the orientation of the base vectors of the oblique S-layer lattice, the subunits had bound with their charge-neutral outer face, leaving the N-terminal region with the polymer binding domain exposed to the ambient environment. From cell wall fragments about half of the S-layer protein could be extracted with 1 M GlcNAc, indicating that the linkage type between the S-layer protein and the SCWP could be related to that of the lectin-polysaccharide type. Interestingly, GlcNAc had an effect on the in vitro self-assembly and recrystallization properties of the S-layer protein that was similar to that of the isolated SCWP. The SCWP generally enhanced the stability of the S-layer protein against endoproteinase Glu-C attack and specifically protected a potential cleavage site in position 138 of the mature S-layer protein.
The
Escherichia coli
Citrate Carrier CitT: a Member of a Novel Eubacterial Transporter Family Related to the 2-Oxoglutarate/Malate Translocator from Spinach Chloroplasts
Authors: Pos Klaas Martinus , Dimroth Peter , Bott Michael 
Published: December 2019

ABSTRACT

Under anoxic conditions in the presence of an oxidizable cosubstrate such as glucose or glycerol,
Escherichia coli
converts citrate to acetate and succinate. Two enzymes are specifically required for the fermentation of the tricarboxylic acid, i.e., a citrate uptake system and citrate lyase. Here we report that the open reading frame (designated
citT
) located at 13.90 min on the
E. coli
chromosome between
rna
and the citrate lyase genes encodes a citrate carrier.
E. coli
transformed with a plasmid expressing
citT
was capable of aerobic growth on citrate, which provides convincing evidence for a function of CitT as a citrate carrier. Transport studies with cell suspensions of the transformed strain indicated that CitT catalyzes a homologous exchange of citrate or a heterologous exchange against succinate, fumarate, or tartrate. Since succinate is the end product of citrate fermentation in
E. coli
, it is likely that CitT functions in vivo as a citrate/succinate antiporter. Analysis of the primary sequence showed that CitT (487 amino acids, 53.1 kDa) is a highly hydrophobic protein with 12 putative transmembrane helices. Sequence comparisons revealed that CitT is related to the 2-oxoglutarate/malate translocator (SODiT1 gene product) from spinach chloroplasts and five bacterial gene products, none of which has yet been functionally characterized. It is suggested that the
E. coli
CitT protein is a member of a novel family of eubacterial transporters involved in the transport of di- and tricarboxylic acids.