bluuebunny/crossref_metadata_2025_split huggingface dataset:
Role of the
gerI
Operon of
Bacillus cereus
569 in the Response of Spores to Germinants
Authors: Clements Mark O. , Moir Anne 
Published: December 2019

ABSTRACT

Bacillus cereus
569 (ATCC 10876) germinates in response to inosine or to
l
-alanine, but the most rapid germination response is elicited by a combination of these germinants. Mutants defective in their germination response to either inosine or to
l
-alanine were isolated after Tn 917 -LTV1 mutagenesis and enrichment procedures; one class of mutant could not germinate in response to inosine as a sole germinant but still germinated in response to
l
-alanine, although at a reduced rate; another mutant germinated normally in response to inosine but was slowed in its germination response to
l
-alanine. These mutants demonstrated that at least two signal response pathways are involved in the triggering of germination. Stimulation of germination in
l
-alanine by limiting concentrations of inosine and stimulation of germination in inosine by low concentrations of
l
-alanine were still detectable in these mutants, suggesting that such stimulation is not dependent on complete functionality of both these germination loci. Two transposon insertions that affected inosine germination were found to be located 2.2 kb apart on the chromosome. This region was cloned and sequenced, revealing an operon of three open reading frames homologous to those in the
gerA
and related operons of
Bacillus subtilis
. The individual genes of this
gerI
operon have been named
gerIA
,
gerIB
, and
gerIC
. The GerIA protein is predicted to possess an unusually long, charged, N-terminal domain containing nine tandem copies of a 13-amino-acid glutamine- and serine-rich sequence.
A Novel Serine/Threonine Protein Kinase Homologue of
Pseudomonas aeruginosa
Is Specifically Inducible within the Host Infection Site and Is Required for Full Virulence in Neutropenic Mice
Authors: Wang Jingyi , Li Caihe , Yang Hongjiang , Mushegian Arcady , Jin Shouguang 
Published: December 2019

ABSTRACT

A genetic locus of
Pseudomonas aeruginosa
was identified that is highly and specifically inducible during infection of neutropenic mice. This locus,
ppkA
, encodes a protein that is highly homologous to eukaryote-type serine/threonine protein kinases. A
ppkA
null mutant strain shows reduced virulence in neutropenic mice compared to the wild type. Overexpression of the PpkA protein greatly inhibited the growth of
Escherichia coli
or
P. aeruginosa
. However, a single amino acid change at the catalytic site of the kinase domain eliminated the toxic effect of PpkA on bacterial cells, suggesting that the kinase domain of PpkA is functional within bacterial cells.
Active Efflux of Organic Solvents by
Pseudomonas putida
S12 Is Induced by Solvents
Authors: Kieboom Jasper , Dennis Jonathan J. , Zylstra Gerben J. , de Bont Jan A. M. 
Published: December 2019

ABSTRACT

Induction of the membrane-associated organic solvent efflux system SrpABC of
Pseudomonas putida
S12 was examined by cloning a 312-bp DNA fragment, containing the
srp
promoter, in the broad-host-range reporter vector pKRZ-1. Compounds that are capable of inducing expression of the
srpABC
genes include aromatic and aliphatic solvents and alcohols. General stress conditions such as pH, temperature, NaCl, or the presence of organic acids did not induce
srp
transcription. Although the solvent efflux pump in
P. putida
S12 is a member of the resistance-nodulation-cell division family of transporters, the
srpABC
genes were not induced by antibiotics or heavy metals.
Identification of Two Binding Domains, One for Peptidoglycan and Another for a Secondary Cell Wall Polymer, on the N-Terminal Part of the S-Layer Protein SbsB from
Bacillus stearothermophilus
PV72/p2
Authors: Sára Margit , Egelseer Eva M. , Dekitsch Christine , Sleytr Uwe B. 
Published: December 2019

ABSTRACT

First studies on the structure-function relationship of the S-layer protein from
B. stearothermophilus
PV72/p2 revealed the coexistence of two binding domains on its N-terminal part, one for peptidoglycan and another for a secondary cell wall polymer (SCWP). The peptidoglycan binding domain is located between amino acids 1 to 138 of the mature S-layer protein comprising a typical S-layer homologous domain. The SCWP binding domain lies between amino acids 240 to 331 and possesses a high serine plus glycine content.
Two Roles for the DNA Recognition Site of the
Klebsiella aerogenes
Nitrogen Assimilation Control Protein
Authors: Pomposiello Pablo J. , Janes Brian K. , Bender Robert A. 
Published: December 2019

ABSTRACT

The nitrogen assimilation control protein (NAC) binds to a site within the promoter region of the histidine utilization operon (
hutUH
) of
Klebsiella aerogenes
, and NAC bound at this site activates transcription of
hutUH
. This NAC-binding site was characterized by a combination of random and directed DNA mutagenesis. Mutations that abolished or diminished in vivo transcriptional activation by NAC were found to lie within a 15-bp region contained within the 26-bp region protected by NAC from DNase I digestion. This 15-bp core has the palindromic ends ATA and TAT, and it matches the consensus for LysR family transcriptional regulators. Protein-binding experiments showed that transcriptional activation in vivo decreased with decreasing binding in vitro. In contrast to the NAC-binding site from
hutUH
, the NAC-binding site from the
gdhA
promoter failed to activate transcription from a semisynthetic promoter, and this failure was not due to weak binding or greatly distorted protein-DNA structure. Mutations in the promoter-proximal half-site of the NAC-binding site from
gdhA
allowed this site to activate transcription. Similar studies using the NAC-binding site from
hut
showed that two mutations in the promoter proximal half-site increased binding but abolished transcriptional activation. Interestingly, for symmetric mutations in the promoter-distal half-site, loss of transcriptional activation was always correlated with a decrease in binding. We conclude from these observations that if the binding in vitro reflects the binding in vivo, then binding of NAC to DNA is not sufficient for transcriptional activation and that the NAC-binding site can be functionally divided in two half-sites, with related but different functions.
Specific In Vivo Labeling of Cell Surface-Exposed Protein Loops: Reactive Cysteines in the Predicted Gating Loop Mark a Ferrichrome Binding Site and a Ligand-Induced Conformational Change of theEscherichia coliFhuA Protein
Authors: Bös Christoph , Lorenzen Dirk , Braun Volkmar 
Published: December 2019

ABSTRACTThe FhuA protein ofEscherichia coliK-12 transports ferrichrome, the antibiotic albomycin, colicin M, and microcin 25 across the outer membrane and serves as a receptor for the phages T1, T5, φ80, and UC-1. FhuA is activated by the electrochemical potential of the cytoplasmic membrane, which probably opens a channel in FhuA. It is thought that the proteins TonB, ExbB, and ExbD function as a coupling device between the cytoplasmic membrane and the outer membrane. Excision of 34 residues from FhuA, tentatively designated the gating loop, converts FhuA into a permanently open channel. FhuA contains two disulfide bridges, one in the gating loop and one close to the C-terminal end. Reduction of the disulfide bridges results in a low in vivo reaction of the cysteines in the gating loop and no reaction of the C-terminal cysteines with biotin-maleimide, as determined by streptavidin-β-galactosidase bound to biotin. In this study we show that a cysteine residue introduced into the gating loop by replacement of Asp-336 displayed a rather high reactivity and was used to monitor structural changes in FhuA upon binding of ferrichrome. Flow cytometric analysis revealed fluorescence quenching by ferrichrome and albomycin of fluorescein-maleimide bound to FhuA. Ferrichrome did not inhibit Cys-336 labeling. In contrast, labeling of Cys-347, obtained by replacing Val-347 in the gating loop, was inhibited by ferrichrome, but ferrichrome quenching was negligible. It is concluded that binding of ferrichrome causes a conformational change of the gating loop and that Cys-347 is part of or close to the ferrichrome binding site. Fluorescence quenching was independent of the TonB activity. The newly introduced cysteines and the replacement of the existing cysteines by serine did not alter sensitivity of cells to the FhuA ligands tested (T5, φ80, T1, colicin M, and albomycin) and fully supported growth on ferrichrome as the sole iron source. Since cells ofE. coliK-12 display no reactivity to thiol reagents, newly introduced cysteines can be used to determine surface-exposed regions of outer membrane proteins and to monitor conformational changes during their function.
Modulation of
Escherichia coli
Adenylyl Cyclase Activity by Catalytic-Site Mutants of Protein IIA
Glc
of the Phosphoenolpyruvate:Sugar Phosphotransferase System
Authors: Reddy Prasad , Kamireddi Madhavi 
Published: December 2019

ABSTRACT

It is demonstrated here that in
Escherichia coli
, the phosphorylated form of the glucose-specific phosphocarrier protein IIA
Glc
of the phosphoenolpyruvate:sugar phosphotransferase system is an activator of adenylyl cyclase and that unphosphorylated IIA
Glc
has no effect on the basal activity of adenylyl cyclase. To elucidate the specific role of IIA
Glc
phosphorylation in the regulation of adenylyl cyclase activity, both the phosphorylatable histidine (H90) and the interactive histidine (H75) of IIA
Glc
were mutated by site-directed mutagenesis to glutamine and glutamate. Wild-type IIA
Glc
and the H75Q mutant, in which the histidine in position 75 has been replaced by glutamine, were phosphorylated by the phosphohistidine-containing phosphocarrier protein (HPr∼P) and were equally potent activators of adenylyl cyclase. Neither the H90Q nor the H90E mutant of IIA
Glc
was phosphorylated by HPr∼P, and both failed to activate adenylyl cyclase. Furthermore, replacement of H75 by glutamate inhibited the appearance of a steady-state level of phosphorylation of H90 of this mutant protein by HPr∼P, yet the H75E mutant of IIA
Glc
was a partial activator of adenylyl cyclase. The H75E H90A double mutant, which cannot be phosphorylated, did not activate adenylyl cyclase. This suggests that the H75E mutant was transiently phosphorylated by HPr∼P but the steady-state level of the phosphorylated form of the mutant protein was decreased due to the repulsive forces of the negatively charged glutamate at position 75 in the catalytic pocket. These results are discussed in the context of the proximity of H75 and H90 in the IIA
Glc
structure and the disposition of the negative charge in the modeled glutamate mutants.
Analysis of
Bacillus subtilis tagAB
and
tagDEF
Expression during Phosphate Starvation Identifies a Repressor Role for PhoP∼P
Authors: Liu Wei , Eder Stephen , Hulett F. Marion 
Published: December 2019

ABSTRACT

The
tagAB
and
tagDEF
operons, which are adjacent and divergently transcribed, encode genes responsible for cell wall teichoic acid synthesis in
Bacillus subtilis
. The
Bacillus
data presented here suggest that PhoP and PhoR are required for direct repression of transcription of the two operons under phosphate starvation conditions but have no regulatory role under phosphate-replete conditions. These data identify for the first time that PhoP∼P has a negative role in Pho regulon gene regulation.
Chimeric Chemoreceptors in
Escherichia coli
: Signaling Properties of Tar-Tap and Tap-Tar Hybrids
Authors: Weerasuriya Siromi , Schneider Brian M. , Manson Michael D. 
Published: December 2019

ABSTRACT

The Tap (taxis toward peptides) receptor and the periplasmic dipeptide-binding protein (DBP) of
Escherichia coli
together mediate chemotactic responses to dipeptides. Tap is a low-abundance receptor. It is present in 5- to 10-fold-fewer copies than high-abundance receptors like Tar and Tsr. Cells expressing Tap as the sole receptor, even from a multicopy plasmid at 5- to 10-fold-overexpressed levels, do not generate sufficient clockwise (CW) signal to tumble and thus swim exclusively smoothly (run). To study the signaling properties of Tap in detail, we constructed reciprocal hybrids between Tap and Tar fused in the linker region between the periplasmic and cytoplasmic domains. The Tapr hybrid senses dipeptides and is a good CW-signal generator, whereas the Tarp hybrid senses aspartate but is a poor CW-signal generator. Thus, the poor CW signaling of Tap is a property of its cytoplasmic domain. Eighteen residues at the carboxyl terminus of high-abundance receptors, including the NWETF sequence that binds the CheR methylesterase, are missing in Tap. The Tart protein, created by removing these 18 residues from Tar, has diminished CW-signaling ability. The Tapl protein, made by adding the last 18 residues of Tar to the carboxyl terminus of Tap, also does not support CW flagellar rotation. However, Tart and Tapl cross-react well with antibody directed against the conserved cytoplasmic region of Tsr, whereas Tap does not cross-react with this antibody. Tap does cross-react, however, with antibody directed against the low-abundance chemoreceptor Trg. The hybrid, truncated, and extended receptors exhibit various levels of methylation. However, Tar and Tapl, which contain a consensus CheR-binding motif (NWETF) at their carboxyl termini, exhibit the highest basal levels of methylation, as expected. We conclude that no simple correlation exists between the abundance of a receptor, its methylation level, and its CW-signaling ability.
Attachment of
Vibrio alginolyticus
to Glass Surfaces Is Dependent on Swimming Speed
Authors: Kogure Kazuhiro , Ikemoto Eiko , Morisaki Hisao 
Published: December 2019

ABSTRACT

The attachment of
Vibrio alginolyticus
to glass surfaces was investigated with special reference to the swimming speed due to the polar flagellum. This bacterium has two types of flagella, i.e., one polar flagellum and numerous lateral flagella. The mutant YM4, which possesses only the polar flagellum, showed much faster attachment than the mutant YM18, which does not possess flagella, indicating that the polar flagellum plays an important role. The attachment of YM4 was dependent on Na
+
concentration and was specifically inhibited by amiloride, an inhibitor of polar flagellum rotation. These results are quite similar to those for swimming speed obtained under the same conditions. Observations with other mutants showed that chemotaxis is not critical and that the flagellum does not act as an appendage for attachment. From these results, it is concluded that the attachment of
V. alginolyticus
to glass surfaces is dependent on swimming speed.
Involvement of Outer Membrane Protein TolC, a Possible Member of the
mar-sox
Regulon, in Maintenance and Improvement of Organic Solvent Tolerance of
Escherichia coli
K-12
Authors: Aono Rikizo , Tsukagoshi Norihiko , Yamamoto Mami 
Published: December 2019

ABSTRACT

Escherichia coli
mutants with improved organic solvent tolerance levels showed high levels of outer membrane protein TolC and inner membrane protein AcrA. The TolC level was regulated positively by MarA, Rob, or SoxS. A possible
mar-rob-sox
box sequence was found upstream of the
tolC
gene. These findings suggest that
tolC
is a member of the
mar-sox
regulon responsive to stress conditions. When a defective
tolC
gene was transferred to
n
-hexane- or cyclohexane-tolerant strains by P1 transduction, the organic solvent tolerance level was lowered dramatically to the decane-tolerant and nonane-sensitive level. The tolerance level was restored by transformation of the transductants with a wild-type
tolC
gene. Therefore, it is evident that TolC is essential for
E. coli
to maintain organic solvent tolerance.
The StpA Protein Functions as a Molecular Adapter To Mediate Repression of the
bgl
Operon by Truncated H-NS in
Escherichia coli
Authors: Free Andrew , Williams Roy M. , Dorman Charles J. 
Published: December 2019

ABSTRACT

The mechanism of repression of the β-glucoside utilization (
bgl
) operon of
Escherichia coli
by a carboxy-terminally truncated derivative of the nucleoid-associated protein H-NS which is defective in DNA binding was investigated. The DNA-binding function of the H-NS-like protein StpA was found to be necessary for repression, which is consistent with a role for StpA as a DNA-binding adapter for mutant derivatives of H-NS.
An ABC Transporter System of
Yersinia pestis
Allows Utilization of Chelated Iron by
Escherichia coli
SAB11
Authors: Bearden Scott W. , Staggs Teanna M. , Perry Robert D. 
Published: December 2019

ABSTRACT

The acquisition of iron is an essential component in the pathogenesis of
Yersinia pestis
, the agent of bubonic and pneumonic plague. A cosmid library derived from the genomic DNA of
Y. pestis
KIM6+ was used for transduction of an
Escherichia coli
mutant (SAB11) defective in the biosynthesis of the siderophore enterobactin. Recombinant plasmids which had a common 13-kb
Bam
HI fragment were isolated from SAB11 transductants in which growth but not enterobactin synthesis was restored on media containing the iron chelator EDDA [ethylenediamine-di(
o
-hydroxyphenyl acetic acid)]. Subcloning and transposon mutagenesis revealed a 5.6-kb region, designated
yfe
, essential for SAB11 growth stimulation. In vitro transcription-translation analysis identified polypeptides of 18, 29.5, 32, and 33 kDa encoded by the
yfe
locus. Sequence analysis shows this locus to be comprised of five genes in two separate operons which have potential Fur-binding sequences in both promoters. A putative polycistronic operon,
yfeABCD
, is Fur regulated and responds to iron and manganese. A functional Fur protein is required for the observed manganese repression of this operon. This operon encodes polypeptides which have strong similarity to the ATP-binding cassette (ABC) family of transporters and include a periplasmic binding protein (YfeA), an ATP-binding protein (YfeB), and two integral membrane proteins (YfeC and -D), which likely function in the acquisition of inorganic iron and possibly other ions. The ∼21-kDa protein encoded by the separately transcribed
yfeE
gene may be located in the cell envelope, since a
yfeE
::Tn
phoA
fusion is PhoA
+
. Mutations in this gene abrogate growth of SAB11 on iron-chelated media.
Isolation of
Rhodospirillum centenum
Mutants Defective in Phototactic Colony Motility by Transposon Mutagenesis
Authors: Jiang Ze-Yu , Rushing Brenda G. , Bai Yong , Gest Howard , Bauer Carl E. 
Published: December 2019

ABSTRACT

The purple photosynthetic bacterium
Rhodospirillum centenum
is capable of forming swarm colonies that rapidly migrate toward or away from light, depending on the wavelength of excitation. To identify components specific for photoperception, we conducted mini-Tn
5
-mediated mutagenesis and screened approximately 23,000 transposition events for mutants that failed to respond to either continuous illumination or to a step down in light intensity. A majority of the ca. 250 mutants identified lost the ability to form motile swarm cells on an agar surface. These cells appeared to contain defects in the synthesis or assembly of surface-induced lateral flagella. Another large fraction of mutants that were unresponsive to light were shown to be defective in the formation of a functional photosynthetic apparatus. Several photosensory mutants also were obtained with defects in the perception and transmission of light signals. Twelve mutants in this class were shown to contain disruptions in a chemotaxis operon, and five mutants contained disruptions of components unique to photoperception. It was shown that screening for photosensory defective
R. centenum
swarm colonies is an effective method for genetic dissection of the mechanism of light sensing in eubacteria.
The
spoIIE
Locus Is Involved in the Spo0A-Dependent Switch in the Location of FtsZ Rings in
Bacillus subtilis
Authors: Khvorova Anastasia , Zhang Ling , Higgins Michael L. , Piggot Patrick J. 
Published: December 2019

ABSTRACT

A switch in the location of FtsZ ring structures from medial to polar is one of the earliest morphological indicators of sporulation in
Bacillus subtilis
. This switch can be artificially caused during vegetative growth by induction of an active form, Sad67, of the transcription regulator, Spo0A (P. A. Levin and R. Losick, Genes Dev. 10:478–488, 1996). We have used immunofluorescence microscopy to show that the switch in FtsZ ring location during vegetative growth caused by Sad67 induction is blocked by a
spoIIE
deletion mutation. The
spoIIE
mutation also impaired polar FtsZ ring formation during sporulation. These results suggest that SpoIIE mediates the Spo0A-directed formation of polar FtsZ rings.
Insertional Inactivation of Genes Encoding the Crystalline Inclusion Proteins of
Photorhabdus luminescens
Results in Mutants with Pleiotropic Phenotypes
Authors: Bintrim Scott B. , Ensign Jerald C. 
Published: December 2019

ABSTRACT

The entomopathogenic bacterium
Photorhabdus luminescens
exhibits phase variation when cultured in vitro. The variant forms of
P. luminescens
are pleiotropic and are designated phase I and phase II variants. One of the characteristic phenotypes of phase I cells is the production of two types of intracellular protein inclusions. The genes encoding the protein monomers that form these inclusions, designated
cipA
and
cipB
, were cloned and characterized.
cipA
and
cipB
encode hydrophobic proteins of 11,648 and 11,308 Da, respectively. The deduced amino acid sequences of CipA and CipB have no significant amino acid sequence similarity to any other known protein but have 25% identity and 49% similarity to each other. Insertional inactivation of
cipA
or
cipB
in phase I cells of
P. luminescens
produced mutants that differ from phase I cells in bioluminescence, the pattern and activities of extracellular products, biochemical traits, adsorption of dyes, and ability to support nematode growth and reproduction. In general, the
cip
mutants were phenotypically more similar to each other than to either phase I or phase II variants.
Sequence Diversity, Predicted Two-Dimensional Protein Structure, and Epitope Mapping of Neisserial Opa Proteins
Authors: Malorny Burkhard , Morelli Giovanna , Kusecek Barica , Kolberg Jan , Achtman Mark 
Published: December 2019

ABSTRACT

The sequence diversity of 45 Opa outer membrane proteins from
Neisseria meningitidis
,
Neisseria gonorrhoeae
,
Neisseria sicca
, and
Neisseria flava
indicates that horizontal genetic exchange of
opa
alleles has been rare between these species. A two-dimensional structural model containing four surface-exposed loops was constructed based on rules derived from porin crystal structure and on conservation of sequence homology within transmembrane β-strands. The minimal continuous epitopes recognized by 23 monoclonal antibodies were mapped to loops 2 and 3. Some of these epitopes are localized on the bacterial cell surface, in support of the model.
Purification of an Extracellular Signaling Molecule Involved in Production of Aerial Mycelium by
Streptomyces coelicolor
Authors: Nodwell Justin R. , Losick Richard 
Published: December 2019

ABSTRACT

We have extensively purified a factor from conditioned medium that restores aerial mycelium formation to a mutant of
Streptomyces coelicolor
that is defective in morphological differentiation. Response to this factor is shown to depend on the presence of the BldK oligopeptide import system. We suggest that this substance acts at the first step in a putative cascade of developmental regulatory signals.