bluuebunny/crossref_metadata_2025_split huggingface dataset:
Bacteria Associated with Benthic Diatoms from Lake Constance: Phylogeny and Influences on Diatom Growth and Secretion of Extracellular Polymeric Substances
Authors: Bruckner Christian G. , Bahulikar Rahul , Rahalkar Monali , Schink Bernhard , Kroth Peter G. 
Published: October 2008

ABSTRACT

The composition of diatom-associated bacterial communities was studied with 14 different unialgal xenic diatom cultures isolated from freshwater epilithic biofilms of Lake Constance, Germany. A clear dominance of
Alphaproteobacteria
was observed, followed by
Betaproteobacteria, Gammaproteobacteria, Bacteroidetes
, and
Verrucomicrobia
. Pure cultures of the diatom
Cymbella microcephala
, which was found to be dominant in epilithic biofilms in Lake Constance, were cocultivated with six associated bacterial strains. All these bacterial strains were able to grow in
C. microcephala
cultures in the absence of organic cosubstrates. Diatom growth was generally enhanced in the presence of bacteria, and polysaccharide secretion was generally increased in the presence of
Proteobacteria
. The monomer composition of extracellular polysaccharides of
C. microcephala
changed in relation to the presence of different bacteria, but the dominant monomers were less affected. Our results indicate that these changes were caused by the diatom itself rather than by specific bacterial degradation. One
Bacteroidetes
strain strongly influenced carbohydrate secretion by the alga via extracellular soluble compounds. Biofilms were formed only in the presence of bacteria. Phylogenetic analysis and coculture studies indicate an adaptation of
Proteobacteria
and
Bacteroidetes
to the microenvironment created by the diatom biofilm.
The Twin-Arginine Signal Peptide of
Bacillus subtilis
YwbN Can Direct either Tat- or Sec-Dependent Secretion of Different Cargo Proteins: Secretion of Active Subtilisin via the
B. subtilis
Tat Pathway
Authors: Kolkman Marc A. B. , van der Ploeg René , Bertels Michael , van Dijk Maurits , van der Laan Joop , van Dijl Jan Maarten , Ferrari Eugenio 
Published: October 2008

ABSTRACT

Proteins that are produced for commercial purposes in
Bacillus subtilis
are commonly secreted via the Sec pathway. Despite its high secretion capacity, the secretion of heterologous proteins via the Sec pathway is often unsuccessful. Alternative secretion routes, like the Tat pathway, are therefore of interest. Two parallel Tat pathways with distinct specificities have previously been discovered in
B. subtilis
. To explore the application potential of these Tat pathways, several commercially relevant or heterologous model proteins were fused to the signal peptides of the known
B. subtilis
Tat substrates YwbN and PhoD. Remarkably, the YwbN signal peptide directed secretion of active subtilisin, a typical Sec substrate, via the
B. subtilis
TatAyCy route. In contrast, the same signal peptide directed Tat-independent secretion of the
Bacillus licheniformis
α-amylase (AmyL). Moreover, the YwbN signal peptide directed secretion of SufI, an
Escherichia coli
Tat substrate, in a Tat-independent manner, most likely via Sec. Our results suggest that cytoplasmic protein folding prior to translocation is probably a major determinant of Tat-dependent protein secretion in
B. subtilis
, as is the case with
E. coli
. We conclude that future applications for the Tat system of
B. subtilis
will most likely involve commercially interesting proteins that are Sec incompatible.
Identification of Components of Electron Transport Chains in the Extremely Thermoacidophilic Crenarchaeon
Metallosphaera sedula
through Iron and Sulfur Compound Oxidation Transcriptomes
Authors: Auernik Kathryne S. , Kelly Robert M. 
Published: October 2008

ABSTRACT

The crenarchaeal order
Sulfolobales
collectively contain at least five major terminal oxidase complexes. Based on genome sequence information, all five complexes are found only in
Metallosphaera sedula
and
Sulfolobus tokodaii
, the two sequenced
Sulfolobales
capable of iron oxidization. While specific respiratory complexes in certain
Sulfolobales
have been characterized previously as proton pumps for maintaining intracellular pH and generating proton motive force, their contribution to sulfur and iron biooxidation has not been considered. For
M. sedula
growing in the presence of ferrous iron and reduced inorganic sulfur compounds (RISCs), global transcriptional analysis was used to track the response of specific genes associated with these complexes, as well as other known and putative respiratory electron transport chain elements. Open reading frames from all five terminal oxidase or
bc
1
-like complexes were stimulated on one or more conditions tested. Components of the
fox
(Msed0467 to Msed0489) and
soxNL
-
cbsABA
(Msed0500 to Msed0505) terminal/quinol oxidase clusters were triggered by ferrous iron, while the
soxABCDD
′ terminal oxidase cluster (Msed0285 to Msed0291) were induced by tetrathionate and S
0
. Chemolithotrophic electron transport elements, including a putative tetrathionate hydrolase (Msed0804), a novel polysulfide/sulfur/dimethyl sulfoxide reductase-like complex (Msed0812 to Msed0818), and a novel heterodisulfide reductase-like complex (Msed1542 to Msed1550), were also stimulated by RISCs. Furthermore, several hypothetical proteins were found to have strong responses to ferrous iron or RISCs, suggesting additional candidates in iron or sulfur oxidation-related pathways. From this analysis, a comprehensive model for electron transport in
M. sedula
could be proposed as the basis for examining specific details of iron and sulfur oxidation in this bioleaching archaeon.
Development and Application of Flow-Cytometric Techniques for Analyzing and Sorting Endospore-Forming Clostridia
Authors: Tracy Bryan P. , Gaida Stefan M. , Papoutsakis Eleftherios T. 
Published: October 2008

ABSTRACT

The study of microbial heterogeneity at the single-cell level is a rapidly growing area of research in microbiology and biotechnology due to its significance in pathogenesis, environmental biology, and industrial biotechnologies. However, the tools available for efficiently and precisely probing such heterogeneity are limited for most bacteria. Here we describe the development and application of flow-cytometric (FC) and fluorescence-assisted cell-sorting techniques for the study of endospore-forming bacteria. We show that by combining FC light scattering (LS) with nucleic acid staining, we can discriminate, quantify, and enrich all sporulation-associated morphologies exhibited by the endospore-forming anaerobe
Clostridium acetobutylicum
. Using FC LS analysis, we quantitatively show that clostridial cultures commonly perform multiple rounds of sporulation and that sporulation is induced earlier by the overexpression of Spo0A, the master regulator of endospore formers. To further demonstrate the power of our approach, we employed FC LS analysis to generate compelling evidence to challenge the long-accepted view in the field that the clostridial cell form is the solvent-forming phenotype.
High-Temperature Ethanol Fermentation and Transformation with Linear DNA in the Thermotolerant Yeast
Kluyveromyces marxianus
DMKU3-1042
Authors: Nonklang Sanom , Abdel-Banat Babiker M. A. , Cha-aim Kamonchai , Moonjai Nareerat , Hoshida Hisashi , Limtong Savitree , Yamada Mamoru , Akada Rinji 
Published: October 2008

ABSTRACT

We demonstrate herein the ability of
Kluyveromyces marxianus
to be an efficient ethanol producer and host for expressing heterologous proteins as an alternative to
Saccharomyces cerevisiae
. Growth and ethanol production by strains of
K. marxianus
and
S. cerevisiae
were compared under the same conditions.
K. marxianus
DMKU3-1042 was found to be the most suitable strain for high-temperature growth and ethanol production at 45°C. This strain, but not
S. cerevisiae
, utilized cellobiose, xylose, xylitol, arabinose, glycerol, and lactose. To develop a
K. marxianus
DMKU3-1042 derivative strain suitable for genetic engineering, a uracil auxotroph was isolated and transformed with a linear DNA of the
S. cerevisiae
Sc
URA3
gene. Surprisingly, Ura
+
transformants were easily obtained. By Southern blot hybridization, the linear Sc
URA3
DNA was found to have inserted randomly into the
K. marxianus
genome. Sequencing of one Lys
−
transformant confirmed the disruption of the Km
LYS1
gene by the Sc
URA3
insertion. A PCR-amplified linear DNA lacking
K. marxianus
sequences but containing an
Aspergillus
α-amylase gene under the control of the Sc
TDH3
promoter together with an Sc
URA3
marker was subsequently used to transform
K. marxianus
DMKU3-1042 in order to obtain transformants expressing
Aspergillus
α-amylase. Our results demonstrate that
K. marxianus
DMKU3-1042 can be an alternative cost-effective bioethanol producer and a host for transformation with linear DNA by use of
S. cerevisiae
-based molecular genetic tools.
4-Dihydrotrisporin-Dehydrogenase, an Enzyme of the Sex Hormone Pathway of
Mucor mucedo
: Purification, Cloning of the Corresponding Gene, and Developmental Expression
Authors: Wetzel Jana , Scheibner Olaf , Burmester Anke , Schimek Christine , Wöstemeyer Johannes 
Published: October 2008

ABSTRACT

The NADP-dependent 4-dihydrotrisporin-dehydrogenase is a (−) mating-type-specific enzyme in the pathway from β-carotene to trisporic acid. This substance and its isomers and derivatives represent the general system of sexual communication in zygomycetes. The (−) mating type of
Mucor mucedo
was stimulated by trisporic acid and the enzyme was purified by ion exchange and affinity chromatography. Several peptides of the 26-kDa protein, digested with trypsin, were sequenced by mass spectrometry. Oligonucleotides based on protein sequence data were used for PCR amplification of genomic DNA. The primary PCR fragment was sequenced and the complete gene,
TSP2
, was isolated. A labeled
TSP2
hybridization probe detects a single-copy gene in the genome of
M. mucedo
. Northern blot analysis with RNAs from different growth stages reveals that the expression of the gene depends on the developmental stage of the mycelium in both mating types of
M. mucedo
. At the enzyme level, activity is found exclusively in the (−) mating type. However, renaturation of proteins in sodium dodecyl sulfate-containing gels revealed the
TSP2
gene product in both mating types. Analyzing the protein sequence places the enzyme in the short chain dehydrogenase superfamily. Thus, it has an evolutionary origin distinct from that of the previously isolated 4-dihydromethyltrisporate dehydrogenase, which belongs to the aldo/keto reductase superfamily. Apart from the
TSP2
genes in the three sequenced zygomycetous genomes (
Phycomyces blakesleeanus
,
Rhizopus oryzae
, and
Mucor circinelloides
), the closest relative is the
Myxococcus xanthus CsgA
gene product, which is also a short chain dehydrogenase, involved in C signaling and fruiting body formation.
Copy Number Suppressors of the
Aspergillus nidulans nimA1
Mitotic Kinase Display Distinctive and Highly Dynamic Cell Cycle-Regulated Locations
Authors: Ukil Leena , Varadaraj Archana , Govindaraghavan Meera , Liu Hui-Lin , Osmani Stephen A. 
Published: October 2008

ABSTRACT

The
Aspergillus nidulans
NIMA kinase is essential for mitosis and is the founding member of the conserved
N
IMA-r
e
lated
k
inase (Nek) family of protein kinases. To gain insight into NIMA function, a copy number suppression screen has been completed that defines three proteins termed MCNA, MCNB, and MCNC (multi-copy-number suppressor of
nimA1
A, B, and C). All display a distinctive and dynamic cell cycle-specific distribution. MCNC has weak similarity to
Saccharomyces cerevisiae
Def1 within a shared CUE-like domain. MCNC, like Def1, is a cytoplasmic protein with slow mobility during sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and its deletion causes polarization defects and a small colony phenotype. MCNC enters nuclei during mitosis. In contrast, MCNB is a nuclear protein displaying increased nuclear levels as cells progress through interphase but is lost from nuclei at mitosis. MCNB is highly related to the
Schizosaccharomyces pombe
forkhead transcription factor Sep1 and is likely a transcriptional activator of
nimA
. Most surprisingly, MCNA, a protein restricted to the aspergilli and pathogenic systemic dimorphic fungi (the
Eurotiomycetes
), defines a nuclear body located near nucleoli at the nuclear periphery of G
2
nuclei. During progression through mitosis, the MCNA body is excluded from nuclei. Cytoplasmic MCNA bodies then diminish during early stages of interphase, and single MCNA bodies are formed within nuclei as interphase progresses. Three sites of MCNA phosphorylation were mapped and mutated to implicate proline-directed phosphorylation in the equal segregation of MCNA during the cell cycle. The data indicate all three MCN proteins likely have cell cycle functions.
Cell Surface Enzyme Attachment Is Mediated by Family 37 Carbohydrate-Binding Modules, Unique to
Ruminococcus albus
Authors: Ezer Anat , Matalon Erez , Jindou Sadanari , Borovok Ilya , Atamna Nof , Yu Zhongtang , Morrison Mark , Bayer Edward A. , Lamed Raphael 
Published: October 2008

ABSTRACT

The rumen bacterium
Ruminococcus albus
binds to and degrades crystalline cellulosic substrates via a unique cellulose degradation system. A unique family of carbohydrate-binding modules (CBM37), located at the C terminus of different glycoside hydrolases, appears to be responsible both for anchoring these enzymes to the bacterial cell surface and for substrate binding.
Alanylclavam Biosynthetic Genes Are Clustered Together with One Group of Clavulanic Acid Biosynthetic Genes inStreptomyces clavuligerus
Authors: Zelyas Nathan J. , Cai Hui , Kwong Thomas , Jensen Susan E. 
Published: October 2008

ABSTRACTStreptomyces clavuligerusproduces at least five different clavam metabolites, including clavulanic acid and the methionine antimetabolite, alanylclavam. In vitro transposon mutagenesis was used to analyze a 13-kb region upstream of the known paralogue gene cluster. The paralogue cluster includes one group of clavulanic acid biosynthetic genes inS. clavuligerus. Twelve open reading frames (ORFs) were found in this area, and mutants were generated in each using either in vitro transposon or PCR-targeted mutagenesis. Mutants with defects in any of the genesorfA,orfB,orfC, ororfDwere unable to produce alanylclavam but could produce all of the other clavams, including clavulanic acid.orfAencodes a predicted hydroxymethyltransferase,orfBencodes a YjgF/YER057c/UK114-family regulatory protein,orfCencodes an aminotransferase, andorfDencodes a dehydratase. All of these types of proteins are normally involved in amino acid metabolism. Mutants inorfCororfDalso accumulated a novel clavam metabolite instead of alanylclavam, and a complementedorfCmutant was able to produce trace amounts of alanylclavam while still producing the novel clavam. Mass spectrometric analyses, together with consideration of the enzymes involved in its production, led to tentative identification of the novel clavam as 8-OH-alanylclavam, an intermediate in the proposed alanylclavam biosynthetic pathway.
Deletion of cgR_1596 and cgR_2070, Encoding NlpC/P60 Proteins, Causes a Defect in Cell Separation in
Corynebacterium glutamicum
R
Authors: Tsuge Yota , Ogino Hidetaka , Teramoto Haruhiko , Inui Masayuki , Yukawa Hideaki 
Published: October 2008

ABSTRACT

In previous work, random genome deletion mutants of
Corynebacterium glutamicum
R were generated using the insertion sequence (IS) element IS
31831
and the Cre/
loxP
excision system. One of these mutants,
C. glutamicum
strain RD41, resulting from the deletion of a 10.1-kb genomic region (ΔcgR_1595 through cgR_1604) from the WT strain, showed cell elongation, and several lines appeared on the cell surface (bamboo shape). The morphological changes were suppressed by overexpression of cgR_1596. Single disruption of cgR_1596 in WT
C. glutamicum
R resulted in morphological changes similar to those observed in the RD41 strain. CgR_1596 has a predicted secretion signal peptide in the amino-terminal region and a predicted NlpC/P60 domain, which is conserved in cell wall hydrolases, in the carboxyl-terminal region. In
C. glutamicum
R, CgR_0802, CgR_1596, CgR_2069, and CgR_2070 have the NlpC/P60 domain; however, only simultaneous disruption of cgR_1596 and cgR_2070, and not cgR_2070 single disruption, resulted in cell growth delay and more severe morphological changes than disruption of cgR_1596. Transmission electron microscopy revealed multiple septa within individual cells of cgR_1596 single and cgR_1596-cgR_2070 double disruptants. Taken together, these results suggest that cgR_1596 and cgR_2070 are involved in cell separation and cell growth in
C. glutamicum
.
A Multicopper Oxidase (Cj1516) and a CopA Homologue (Cj1161) Are Major Components of the Copper Homeostasis System of
Campylobacter jejuni
Authors: Hall Stephen J. , Hitchcock Andrew , Butler Clive S. , Kelly David J. 
Published: October 2008

ABSTRACT

Metal ion homeostasis mechanisms in the food-borne human pathogen
Campylobacter jejuni
are poorly understood. The Cj1516 gene product is homologous to the multicopper oxidase CueO, which is known to contribute to copper tolerance in
Escherichia coli
. Here we show, by optical absorbance and electron paramagnetic resonance spectroscopy, that purified recombinant Cj1516 contains both T1 and trinuclear copper centers, which are characteristic of multicopper oxidases. Inductively coupled plasma mass spectrometry revealed that the protein contained approximately six copper atoms per polypeptide. The presence of an N-terminal “twin arginine” signal sequence suggested a periplasmic location for Cj1516, which was confirmed by the presence of
p
-phenylenediamine (
p
-PD) oxidase activity in periplasmic fractions of wild-type but not Cj1516 mutant cells. Kinetic studies showed that the pure protein exhibited
p
-PD, ferroxidase, and cuprous oxidase activities and was able to oxidize an analogue of the bacterial siderophore anthrachelin (3,4-dihydroxybenzoate), although no iron uptake impairment was observed in a Cj1516 mutant. However, this mutant was very sensitive to increased copper levels in minimal media, suggesting a role in copper tolerance. This was supported by increased expression of the Cj1516 gene in copper-rich media. A mutation in a second gene, the Cj1161c gene, encoding a putative CopA homologue, was also found to result in copper hypersensitivity, and a Cj1516 Cj1161c double mutant was found to be more copper sensitive than either single mutant. These observations and the apparent lack of alternative copper tolerance systems suggest that Cj1516 (CueO) and Cj1161 (CopA) are major proteins involved in copper homeostasis in
C. jejuni
.
Regulation of Hemin Binding Proteins by a Novel Transcriptional Activator in
Porphyromonas gingivalis
Authors: Wu Jie , Lin Xinghua , Xie Hua 
Published: October 2008

ABSTRACT

One of the features of the periodontal pathogen
Porphyromonas gingivalis
is the presence of complex iron acquisition systems that include an
hmuYRSTUV
locus. HmuY and HmuR are hemin binding proteins required for
P. gingivalis
growth. Previous studies have demonstrated that expression of the
hmu
locus is regulated in response to environmental changes, such as growth phases. However, the mechanisms involved in
hmu
gene regulation are poorly understood. Here we report that a novel transcriptional activator, PG1237, is required for the expression of
humY
and
humR
, but not other iron acquisition-related genes, such as
fetB
and
tlr
, which also encode hemin binding proteins. Real-time reverse transcription-PCR analysis revealed that a mutation in the pg1237 gene decreased expression of
hmuY
and
hmuR
149- and 25-fold, respectively, compared to that observed in the wild-type strain. In addition, differential expression of
hmuY
,
hmuR
, and the pg1237 gene was found to be quorum-sensing dependent, such that higher expression levels of these genes were observed when
P. gingivalis
was grown at a lower cell density, such as that seen during the early exponential growth phase. This work demonstrates the involvement of a novel transcriptional activator, PG1237, in expression of the
hmu
operon in a cell density-dependent fashion.
Posttranscriptional Regulation of Flagellin Synthesis in
Helicobacter pylori
by the RpoN Chaperone HP0958
Authors: Douillard Francois P. , Ryan Kieran A. , Caly Delphine L. , Hinds Jason , Witney Adam A. , Husain Sarah E. , O'Toole Paul W. 
Published: October 2008

ABSTRACT

The
Helicobacter pylori
protein HP0958 is essential for flagellum biogenesis. It has been shown that HP0958 stabilizes the σ
54
factor RpoN. The aim of this study was to further investigate the role of HP0958 in flagellum production in
H. pylori
. Global transcript analysis identified a number of flagellar genes that were differentially expressed in an HP0958 mutant strain. Among these, the transcription of the major flagellin gene
flaA
was upregulated twofold, suggesting that HP0958 was a negative regulator of the
flaA
gene. However, the production of the FlaA protein was significantly reduced in the HP0958 mutant, and this was not due to the decreased stability of the FlaA protein. RNA stability analysis and binding assays indicated that HP0958 binds and destabilizes
flaA
mRNA. The HP0958 mutant was successfully complemented, confirming that the mutant phenotype described was due to the lack of HP0958. We conclude that HP0958 is a posttranscriptional regulator that modulates the amount of the
flaA
message available for translation in
H. pylori
.
Regulation of Fructose Transport and Its Effect on Fructose Toxicity in
Anabaena
spp
Authors: Ungerer Justin L. , Pratte Brenda S. , Thiel Teresa 
Published: October 2008

ABSTRACT

Anabaena variabilis
grows heterotrophically using fructose, while the close relative
Anabaena
sp. strain PCC 7120 does not. Introduction of a cluster of genes encoding a putative ABC transporter, herein named
frtRABC
, into
Anabaena
sp. strain PCC 7120 on a replicating plasmid allowed that strain to grow in the dark using fructose, indicating that these genes are necessary and sufficient for heterotrophic growth. FrtR, a putative LacI-like regulatory protein, was essential for heterotrophic growth of both cyanobacterial strains. Transcriptional analysis revealed that the transport system was induced by fructose and that in the absence of FrtR,
frtA
was very highly expressed, with or without fructose. In the
frtR
mutant, fructose uptake was immediate, in contrast to that in the wild-type strain, which required about 40 min for induction of transport. In the
frtR
mutant, high-level expression of the fructose transporter resulted in cells that were extremely sensitive to fructose. Even in the presence of the inducer, fructose, expression of
frtA
was low in the wild-type strain compared to that in the
frtR
mutant, indicating that FrtR repressed the transporter genes even in the presence of fructose. FrtR bound to the upstream region of
frtA
, but binding was not visibly altered by fructose, further supporting the hypothesis that fructose has only a modest effect in relieving repression of
frtA
by FrtR.
A. variabilis
grew better with increasing concentrations of fructose up to 50 mM, showing increased cell size and heterocyst frequency.
Anabaena
sp. strain PCC 7120 did not show any of these changes when it was grown with fructose. Thus, although
Anabaena
sp. strain PCC 7120 could take up fructose and use it in the dark, fructose did not improve growth in the light.