bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification and Properties of the Genes Encoding Microcin E492 and Its Immunity Protein
Authors: Lagos Rosalba , Villanueva Jorge E. , Monasterio Octavio 
Published: December 2019

ABSTRACT

The gene coding for the immunity protein (
mceB
) and the structural gene of microcin E492 (
mceA
), a low-molecular-weight channel-forming bacteriocin produced by a strain of
Klebsiella pneumoniae
, have been characterized. The microcin gene codes for a precursor protein of either 99 or 103 amino acids. Protein sequencing of the N-terminal region of microcin E492 unequivocally identified this gene as the microcin structural gene and indicated that this microcin is synthesized as a precursor protein that is cleaved at either amino acid 15 or 19, at a site resembling the double-glycine motif. The gene encoding the 95-amino-acid immunity protein (
mceB
) was identified by cloning the DNA segment that encodes only this polypeptide into an expression vector and demonstrating the acquisition of immunity to microcin E492. As expected, the immunity protein was found to be associated with the inner membrane. Analysis of the DNA sequence indicates that these genes belong to the same family as microcin 24, and they do not share structural motifs with any other known channel-forming bacteriocin. The organization of the microcin- and immunity protein-encoding genes suggests that they are coordinately expressed.
Isolation of a Putative
Candida albicans
Transcriptional Regulator Involved in Pleiotropic Drug Resistance by Functional Complementation of a
pdr1 pdr3
Mutation in
Saccharomyces cerevisiae
Authors: Talibi Driss , Raymond Martine 
Published: December 2019

ABSTRACT

Three
Candida albicans
genes, designated
FCR
(for fluconazole resistance), have been isolated by their ability to complement the fluconazole (FCZ) hypersensitivity of a
Saccharomyces cerevisiae
mutant lacking the transcription factors Pdr1p and Pdr3p. Overexpression of any of the three
FCR
genes in the
pdr1 pdr3
mutant resulted in increased resistance of the cells to FCZ and cycloheximide and in increased expression of
PDR5
, a gene coding for a drug efflux transporter of the ATP-binding cassette superfamily and whose transcription is under the control of Pdr1p and Pdr3p. Deletion of
PDR5
in the
pdr1 pdr3
strain completely abrogated the ability of the three
FCR
genes to confer FCZ resistance, demonstrating that
PDR5
is required for
FCR
-mediated FCZ resistance in
S. cerevisiae
. The
FCR1
gene encodes a putative 517-amino-acid protein with an N-terminal Zn
2
C
6
-type zinc finger motif homologous to that found in fungal zinc cluster proteins, including
S. cerevisiae
Pdr1p and Pdr3p. We have constructed a
C. albicans
CAI4-derived mutant strain carrying a homozygous deletion of the
FCR1
gene and analyzed its ability to grow in the presence of FCZ. We found that the
fcr1Δ/fcr1
Δ mutant displays hyperresistance to FCZ and other antifungal drugs compared to the parental CAI4 strain. This hyperresistance could be reversed to wild-type levels by reintroduction of a plasmid-borne copy of
FCR1
into the
fcr1Δ/fcr1
Δ mutant. Taken together, our results indicate that the
FCR1
gene behaves as a negative regulator of drug resistance in
C. albicans
and constitute the first evidence that FCZ resistance can result from the inactivation of a regulatory factor such as Fcr1p.
The
Myxococcus xanthus pilQ
(
sglA
) Gene Encodes a Secretin Homolog Required for Type IV Pilus Biogenesis, Social Motility, and Development
Authors: Wall Daniel , Kolenbrander Paul E. , Kaiser Dale 
Published: December 2019

ABSTRACT

The
Myxococcus xanthus sglA1
spontaneous mutation was originally isolated because it allowed dispersed cell growth in liquid yet retained the ability to form fruiting bodies. Consequently, most of today’s laboratory strains either contain the
sglA1
mutation or were derived from strains that carry it. Subsequent work showed that
sglA
was a gene for social gliding motility, a process which is mediated by type IV pili. Here
sglA
is shown to map to the major
pil
cluster and to encode a 901-amino-acid open reading frame (ORF) that is homologous to the secretin superfamily of proteins. Secretins form a channel in the outer membrane for the transport of macromolecules. The closest homologs found were PilQ proteins from
Pseudomonas aeruginosa
and
Neisseria gonorrhoeae
, which are required for type IV pili biogenesis and twitching motility. To signify these molecular and functional similarities, we have changed the name of
sglA
to
pilQ
. The hypomorphic
pilQ1
(
sglA1
) allele was sequenced and found to contain two missense mutations at residues 741 (G→S) and 762 (N→G). In addition, 19 independent social (S)-motility mutations are shown to map to the
pilQ
locus. In-frame deletions of
pilQ
and its downstream gene,
orfL
, were constructed.
pilQ
is shown to be essential for pilus biogenesis, S-motility, rippling, and fruiting body formation, while
orfL
is dispensable for these processes. The
pilQ1
allele, but not the Δ
pilQ
allele, was found to render cells hypersensitive to vancomycin, suggesting that PilQ1 alters the permeability properties of the outer membrane. Many differences between
pilQ1
and
pilQ
+
strains have been noted in the literature. We discuss some of these observations and how they may be rationalized in the context of our molecular and functional findings.
Activities of the
Porphyromonas gingivalis
PrtP Proteinase Determined by Construction of
prtP
-Deficient Mutants and Expression of the Gene in
Bacteroides
Species
Authors: Barkocy-Gallagher Genevieve A. , Foley Joseph W. , Lantz Marilyn S. 
Published: December 2019

ABSTRACT

PrtP is a major cysteine proteinase of
Porphyromonas gingivalis
. The gene encoding this proteinase,
prtP
, was cloned into the
Escherichia coli-Bacteroides
shuttle vectors pFD288 and pFD340 and was expressed in
Bacteroides
cells, apparently under the control of its own promoter, when in pFD288, or a
Bacteroides
promoter present on pFD340. Proteolytically active PrtP was detected by fibrinogen zymography in cells or spent growth medium of several
Bacteroides
species harboring the recombinant plasmids. The proteinase was recovered from
Bacteroides fragilis
ATCC 25285(pFD340-
prtP
) cells by 3-[(3-cholamidopropyl)-dimethyl-ammonio]-1-propanesulfonate (CHAPS) extraction and characterized with regard to exopeptidase specificity and sensitivity to proteinase inhibitors. Lys-amidolytic activity, but not Arg-amidolytic activity, was detected. PrtP was activated by cysteine and, to a lesser extent, dithiothreitol, and it was stimulated by glycine-containing compounds. It also was inhibited by
Nα-p
-tosyl-
l
-lysine chloromethyl ketone (TLCK) and, to a lesser extent, H-
d
-Tyr-
l
-Pro-
l
-arginyl chloromethyl ketone (YPRCK) and was relatively insensitive to EDTA and leupeptin. Neither
B. fragilis
ATCC 25285(pFD340-
prtP
) cells nor the CHAPS extract effected hemagglutination of sheep red blood cells or collagen cleavage, but the cells did cleave gelatin. Furthermore,
P. gingivalis
W12, ATCC 33277, KDP110, and HG66 with knockout mutations in
prtP
were constructed by allelic replacement. Unlike the parent strains, the mutant strains produced beige colonies on plates containing sheep blood. These strains also were affected in their ability to effect hemagglutination, cleave collagen, and cleave a Lys-specific peptide substrate. This report presents the results of the first characterization of the PrtP proteinase clearly in the absence of any influence by other
P. gingivalis
proteins and describes the properties of
P. gingivalis
cells defective in the production of PrtP.
Adaptation of
Mycobacterium smegmatis
to Stationary Phase
Authors: Smeulders Marjan J. , Keer Jacquie , Speight Richard A. , Williams Huw D. 
Published: December 2019

ABSTRACT

Mycobacterium tuberculosis
can persist for many years within host lung tissue without causing clinical disease. Little is known about the state in which the bacilli survive, although it is frequently referred to as dormancy. Some evidence suggests that cells survive in nutrient-deprived stationary phase. Therefore, we are studying stationary-phase survival of
Mycobacterium smegmatis
as a model for mycobacterial persistence.
M. smegmatis
cultures could survive 650 days of either carbon, nitrogen, or phosphorus starvation. In carbon-limited medium, cells entered stationary phase before the carbon source (glycerol) had been completely depleted and glycerol uptake from the medium continued during the early stages of stationary phase. These results suggest that the cells are able to sense when the glycerol is approaching limiting concentrations and initiate a shutdown into stationary phase, which involves the uptake of the remaining glycerol from the medium. During early stationary phase, cells underwent reductive cell division and became more resistant to osmotic and acid stress and pool mRNA stabilized. Stationary-phase cells were also more resistant to oxidative stress, but this resistance was induced during late exponential phase in a cell-density-dependent manner. Upon recovery in fresh medium, stationary-phase cultures showed an immediate increase in protein synthesis irrespective of culture age. Colony morphology variants accumulated in stationary-phase cultures. A flat colony variant was seen in 75% of all long-term-stationary-phase cultures and frequently took over the whole population. Cryo scanning electron microscopy showed that the colony organization was different in flat colony strains, flat colonies appearing less well organized than wild-type colonies. Competition experiments with an exponential-phase-adapted wild-type strain showed that the flat strain had a competitive advantage in stationary phase, as well a providing evidence that growth and cell division occur in stationary-phase cultures of
M. smegmatis
. These results argue against stationary-phase
M. smegmatis
cultures entering a quiescent state akin to dormancy but support the idea that they are a dynamic population of cells.
Doxorubicin Overproduction in
Streptomyces peucetius
: Cloning and Characterization of the
dnrU
Ketoreductase and
dnrV
Genes and the
doxA
Cytochrome P-450 Hydroxylase Gene
Authors: Lomovskaya Natalie , Otten Sharee L. , Doi-Katayama Yukiko , Fonstein Leonid , Liu Xiao-Chun , Takatsu Toshio , Inventi-Solari Augusto , Filippini Silvia , Torti Francesca , Colombo Anna Luisa , Hutchinson C. Richard 
Published: December 2019

ABSTRACT

Doxorubicin-overproducing strains of
Streptomyces peucetius
ATCC 29050 can be obtained through manipulation of the genes in the region of the doxorubicin (DXR) gene cluster that contains
dpsH
, the
dpsG
polyketide synthase gene, the putative
dnrU
ketoreductase gene,
dnrV
, and the
doxA
cytochrome P-450 gene. These five genes were characterized by sequence analysis, and the effects of replacing
dnrU
,
dnrV
,
doxA
, or
dpsH
with mutant alleles and of
doxA
overexpression on the production of the principal anthracycline metabolites of
S. peucetius
were studied. The exact roles of
dpsH
and
dnrV
could not be established, although
dnrV
is implicated in the enzymatic reactions catalyzed by DoxA, but
dnrU
appears to encode a ketoreductase specific for the C-13 carbonyl of daunorubicin (DNR) and DXR or their biosynthetic precursors. The highest DXR titers were obtained in a
dnrX dnrU
(N. Lomovskaya, Y. Doi-Katayama, S. Filippini, C. Nastro, L. Fonstein, M. Gallo, A. L. Colombo, and C. R. Hutchinson, J. Bacteriol. 180:2379–2386, 1998) double mutant and a
dnrX dnrU dnrH
(C. Scotti and C. R. Hutchinson, J. Bacteriol. 178:7316–7321, 1996) triple mutant. Overexpression of
doxA
in a
doxA::aphII
mutant resulted in the accumulation of DXR precursors instead of in a notable increase in DXR production. In contrast, overexpression of
dnrV
and
doxA
jointly in the
dnrX dnrU
double mutant or the
dnrX dnrU dnrH
triple mutant increased the DXR titer 36 to 86%.
Convergent Pathways for Utilization of the Amino Sugars
N
-Acetylglucosamine,
N
-Acetylmannosamine, and
N
-Acetylneuraminic Acid by
Escherichia coli
Authors: Plumbridge Jacqueline , Vimr Eric 
Published: December 2019

ABSTRACT

N
-Acetylglucosamine (GlcNAc) and
N
-acetylneuraminic acid (NANA) are good carbon sources for
Escherichia coli
K-12, whereas
N
-acetylmannosamine (ManNAc) is metabolized very slowly. The isolation of regulatory mutations which enhanced utilization of ManNAc allowed us to elucidate the pathway of its degradation. ManNAc is transported by the
manXYZ
-encoded phosphoenolpyruvate-dependent phosphotransferase system (PTS) transporter producing intracellular ManNAc-6-P. This phosphorylated hexosamine is subsequently converted to GlcNAc-6-P, which is further metabolized by the
nagBA
-encoded deacetylase and deaminase of the GlcNAc-6-P degradation pathway. Two independent mutations are necessary for good growth on ManNAc. One mutation maps to
mlc
, and mutations in this gene are known to enhance the expression of
manXYZ
. The second regulatory mutation was mapped to the
nanAT
operon, which encodes the NANA transporter and NANA lyase. The combined action of the
nanAT
gene products converts extracellular NANA to intracellular ManNAc. The second regulatory mutation defines an open reading frame (ORF), called
yhcK
, as the gene for the repressor of the
nan
operon (
nanR
). Mutations in the repressor enhance expression of the
nanAT
genes and, presumably, three distal, previously unidentified genes,
yhcJIH
. Expression of just one of these downstream ORFs,
yhcJ
, is necessary for growth on ManNAc in the presence of an
mlc
mutation. The
yhcJ
gene appears to encode a ManNAc-6-P-to-GlcNAc-6-P epimerase (
nanE
). Another putative gene in the
nan
operon,
yhcI
, likely encodes ManNAc kinase (
nanK
), which should phosphorylate the ManNAc liberated from NANA by the NanA protein. Use of NANA as carbon source by
E. coli
also requires the
nagBA
gene products. The existence of a ManNAc kinase and epimerase within the
nan
operon allows us to propose that the pathways for dissimilation of the three amino sugars GlcNAc, ManNAc, and NANA, all converge at the step of GlcNAc-6-P.
Multiple Small Heat Shock Proteins in Rhizobia
Authors: Münchbach Martin , Nocker Andreas , Narberhaus Franz 
Published: December 2019

ABSTRACTSeven genes coding for small heat shock proteins (sHsps) inBradyrhizobium japonicumhave been identified. They are organized in five operons that are coordinately regulated by ROSE, a negativelycis-acting DNA element. The deduced sHsps can be divided into two separate classes: class A, consisting of proteins that show similarity toEscherichia coliIbpA and IbpB, and class B, whose members display significant similarity to other sHsps from prokaryotes and eukaryotes. Two-dimensional gel electrophoresis and Edman sequencing revealed the presence of at least 12 sHsps inB. japonicum, indicating a remarkable abundance of sHsps in this organism. Three additional members of class A and two potentially novel heat shock proteins were identified on the basis of their amino termini. The presence of multiple sHsps was also demonstrated for a variety ofRhizobiumandBradyrhizobiumspecies by immunoblot analysis and two-dimensional gel electrophoresis. An extensive database survey revealed that, in contrast to the rhizobia, other bacteria contain maximally two sHsps whereas many plants have been reported to possess a sHsp superfamily.
Role of Quinolinate Phosphoribosyl Transferase in Degradation of Phthalate by
Burkholderia cepacia
DBO1
Authors: Chang Hung-Kuang , Zylstra Gerben J. 
Published: December 2019

ABSTRACT

Two distinct regions of DNA encode the enzymes needed for phthalate degradation by
Burkholderia cepacia
DBO1. A gene coding for an enzyme (quinolinate phosphoribosyl transferase) involved in the biosynthesis of NAD
+
was identified between these two regions by sequence analysis and functional assays. Southern hybridization experiments indicate that DBO1 and other phthalate-degrading
B. cepacia
strains have two dissimilar genes for this enzyme, while non-phthalate-degrading
B. cepacia
strains have only a single gene. The sequenced gene was labeled
ophE
, due to the fact that it is specifically induced by phthalate as shown by
lacZ
gene fusions. Insertional knockout mutants lacking
ophE
grow noticeably slower on phthalate while exhibiting normal rates of growth on other substrates. The fact that elevated levels of quinolinate phosphoribosyl transferase enhance growth on phthalate stems from the structural similarities between phthalate and quinolinate: phthalate is a competitive inhibitor of this enzyme and the phthalate catabolic pathway cometabolizes quinolinate. The recruitment of this gene for growth on phthalate thus gives
B. cepacia
an advantage over other phthalate-degrading bacteria in the environment.
Diverse Oxygenations Catalyzed by Carbazole 1,9a-Dioxygenase from
Pseudomonas
sp. Strain CA10
Authors: Nojiri Hideaki , Nam Jeong-Won , Kosaka Mikiko , Morii Ken-Ichi , Takemura Tetsuo , Furihata Kazuo , Yamane Hisakazu , Omori Toshio 
Published: December 2019

ABSTRACT

Carbazole 1,9a-dioxygenase (CARDO) from
Pseudomonas
sp. strain CA10 is a multicomponent enzyme that catalyzes the angular dioxygenation of carbazole, dibenzofuran, and dibenzo-
p
-dioxin. It was revealed by gas chromatography-mass spectrometry and
1
H and
13
C nuclear magnetic resonance analyses that xanthene and phenoxathiin were converted to 2,2′,3-trihydroxydiphenylmethane and 2,2′,3-trihydroxydiphenyl sulfide, respectively. Thus, for xanthene and phenoxathiin, angular dioxygenation by CARDO occurred at the angular position adjacent to the oxygen atom to yield hetero ring-cleaved compounds. In addition to the angular dioxygenation, CARDO catalyzed the
cis
dihydroxylation of polycyclic aromatic hydrocarbons and biphenyl. Naphthalene and biphenyl were converted by CARDO to
cis
-1,2-dihydroxy-1,2-dihydronaphthalene and
cis
-2,3-dihydroxy-2,3-dihydrobiphenyl, respectively. On the other hand, CARDO also catalyzed the monooxygenation of sulfur heteroatoms in dibenzothiophene and of the benzylic methylenic group in fluorene to yield dibenzothiophene-5-oxide and 9-hydroxyfluorene, respectively. These results indicate that CARDO has a broad substrate range and can catalyze diverse oxygenation: angular dioxygenation,
cis
dihydroxylation, and monooxygenation. The diverse oxygenation catalyzed by CARDO for several aromatic compounds might reflect the differences in the binding of the substrates to the reaction center of CARDO.
Identification of Genetic Determinants for the Hemolytic Activity of
Streptococcus agalactiae
by IS
S1
Transposition
Authors: Spellerberg Barbara , Pohl Barbara , Haase Gerhard , Martin Simone , Weber-Heynemann Josephine , Lütticken Rudolf 
Published: December 2019

ABSTRACT

Streptococcus agalactiae
is a poorly transformable bacterium and studies of molecular mechanisms are difficult due to the limitations of genetic tools. Employing the novel pGh9:IS
S1
transposition vector we generated plasmid-based mutant libraries of
S. agalactiae
strains O90R and AC475 by random chromosomal integration. A screen for mutants with a nonhemolytic phenotype on sheep blood agar led to the identification of a genetic locus harboring several genes that are essential for the hemolytic function and pigment production of
S. agalactiae
. Nucleotide sequence analysis of nonhemolytic mutants revealed that four mutants had distinct insertion sites in a single genetic locus of 7 kb that was subsequently designated
cyl
. Eight different open reading frames were identified:
cylX
,
cylD
,
cylG
,
acpC
,
cylZ
,
cylA
,
cylB
, and
cylE
, coding for predicted proteins with molecular masses of 11, 33, 26, 11, 15, 35, 32, and 78 kDa, respectively. The deduced amino acid sequence of the protein encoded by
cylA
harbors a conserved ATP-binding cassette (ABC) motif, and the predicted proteins encoded by
cylA
and
cylB
have significant similarities to the nucleotide binding and transmembrane proteins of typical ABC transporter systems. Transcription analysis by reverse transcription-PCR suggests that
cylX
to
cylE
are part of an operon. The requirement of
acpC
and
cylZABE
for hemolysin production of
S. agalactiae
was confirmed either by targeted mutagenesis with the vector pGh5, complementation studies with pAT28, or analysis of insertion elements in naturally occurring nonhemolytic mutants.
Histidinol Phosphate Phosphatase, Catalyzing the Penultimate Step of the Histidine Biosynthesis Pathway, Is Encoded by
ytvP
(
hisJ
) in
Bacillus subtilis
Authors: le Coq Dominique , Fillinger Sabine , Aymerich Stéphane 
Published: December 2019

ABSTRACT

The deduced product of the
Bacillus subtilis ytvP
gene is similar to that of
ORF13
, a gene of unknown function in the
Lactococcus lactis
histidine biosynthesis operon. A
B. subtilis ytvP
mutant was auxotrophic for histidine. The only enzyme of the histidine biosynthesis pathway that remained uncharacterized in
B. subtilis
was histidinol phosphate phosphatase (HolPase), catalyzing the penultimate step of this pathway. HolPase activity could not be detected in crude extracts of the
ytvP
mutant, while purified glutathione
S
-transferase-YtvP fusion protein exhibited strong HolPase activity. These observations demonstrated that HolPase is encoded by
ytvP
in
B. subtilis
and led us to rename this gene
hisJ
. Together with the HolPase of
Saccharomyces cerevisiae
and the presumed HolPases of
L. lactis
and
Schizosaccharomyces pombe
, HisJ constitutes a family of related enzymes that are not homologous to the HolPases of
Escherichia coli
,
Salmonella typhimurium
, and
Haemophilus influenzae
.
A Novel
Campylobacter jejuni
Two-Component Regulatory System Important for Temperature-Dependent Growth and Colonization
Authors: Brás Ana M. , Chatterjee Shimonti , Wren Brendan W. , Newell Diane G. , Ketley Julian M. 
Published: December 2019

ABSTRACT

Campylobacter jejuni
colonizes the intestines of domestic and wild animals and is a common cause of human diarrheal disease. We identified a two-component regulatory system, designated the RacR-RacS (reduced ability to colonize) system, that is involved in a temperature-dependent signalling pathway. A mutation of the response regulator gene
racR
reduced the organism’s ability to colonize the chicken intestinal tract and resulted in temperature-dependent changes in its protein profile and growth characteristics.
NahY, a Catabolic Plasmid-Encoded Receptor Required for Chemotaxis of
Pseudomonas putida
to the Aromatic Hydrocarbon Naphthalene
Authors: Grimm Ann C. , Harwood Caroline S. 
Published: December 2019

ABSTRACT

Pseudomonas putida
G7 exhibits chemotaxis to naphthalene, but the molecular basis for this was not known. A new gene,
nahY
, was found to be cotranscribed with
meta
cleavage pathway genes on the NAH7 catabolic plasmid for naphthalene degradation. The
nahY
gene encodes a 538-amino-acid protein with a membrane topology and a C-terminal region that resemble those of chemotaxis transducer proteins. A
P. putida
G7
nahY
mutant grew on naphthalene but was not chemotactic to this aromatic hydrocarbon. The protein NahY thus appears to function as a chemoreceptor for naphthalene or a related compound. The presence of
nahY
on a catabolic plasmid implies that chemotaxis may facilitate biodegradation.