bluuebunny/crossref_metadata_2025_split huggingface dataset:
Supplementation of Intracellular XylR Leads to Coutilization of Hemicellulose Sugars
Authors: Groff Dan , Benke Peter I. , Batth Tanveer S. , Bokinsky Gregory , Petzold Christopher J. , Adams Paul D. , Keasling Jay D. 
Published: January 2012

ABSTRACT

Escherichia coli
has the potential to be a powerful biocatalyst for the conversion of lignocellulosic biomass into useful materials such as biofuels and polymers. One important challenge in using
E. coli
for the transformation of biomass sugars is diauxie, or sequential utilization of different types of sugars. We demonstrate that, by increasing the intracellular levels of the transcription factor XylR, the preferential consumption of arabinose before xylose can be eliminated. In addition, XylR augmentation must be finely tuned for robust coutilization of these two hemicellulosic sugars. Using a novel technique for scarless gene insertion, an additional copy of
xylR
was inserted into the
araBAD
operon. The resulting strain was superior at cometabolizing mixtures of arabinose and xylose and was able to produce at least 36% more ethanol than wild-type strains. This strain is a useful starting point for the development of an
E. coli
biocatalyst that can simultaneously convert all biomass sugars.
Engineered Escherichia coli Silver-Binding Periplasmic Protein That Promotes Silver Tolerance
Authors: Hall Sedlak Ruth , Hnilova Marketa , Grosh Carolynn , Fong Hanson , Baneyx Francois , Schwartz Dan , Sarikaya Mehmet , Tamerler Candan , Traxler Beth 
Published: January 2012

ABSTRACT

Silver toxicity is a problem that microorganisms face in medical and environmental settings. Through exposure to silver compounds, some bacteria have adapted to growth in high concentrations of silver ions. Such adapted microbes may be dangerous as pathogens but, alternatively, could be potentially useful in nanomaterial-manufacturing applications. While naturally adapted isolates typically utilize efflux pumps to achieve metal resistance, we have engineered a silver-tolerant
Escherichia coli
strain by the use of a simple silver-binding peptide motif. A silver-binding peptide, AgBP2, was identified from a combinatorial display library and fused to the C terminus of the
E. coli
maltose-binding protein (MBP) to yield a silver-binding protein exhibiting nanomolar affinity for the metal. Growth experiments performed in the presence of silver nitrate showed that cells secreting MBP-AgBP2 into the periplasm exhibited silver tolerance in a batch culture, while those expressing a cytoplasmic version of the fusion protein or MBP alone did not. Transmission electron microscopy analysis of silver-tolerant cells revealed the presence of electron-dense silver nanoparticles. This is the first report of a specifically engineered metal-binding peptide exhibiting a strong
in vivo
phenotype, pointing toward a novel ability to manipulate bacterial interactions with heavy metals by the use of short and simple peptide motifs. Engineered metal-ion-tolerant microorganisms such as this
E. coli
strain could potentially be used in applications ranging from remediation to interrogation of biomolecule-metal interactions
in vivo
.
Significant Role for Microbial Autotrophy in the Sequestration of Soil Carbon
Authors: Yuan Hongzhao , Ge Tida , Chen Caiyan , O'Donnell Anthony G. , Wu Jinshui 
Published: January 2012

ABSTRACT

Soils were incubated for 80 days in a continuously labeled
14
CO
2
atmosphere to measure the amount of labeled C incorporated into the microbial biomass. Microbial assimilation of
14
C differed between soils and accounted for 0.12% to 0.59% of soil organic carbon (SOC). Assuming a terrestrial area of 1.4 × 10
8
km
2
, this represents a potential global sequestration of 0.6 to 4.9 Pg C year
−1
. Estimated global C sequestration rates suggest a “missing sink” for carbon of between 2 and 3 Pg C year
−1
. To determine whether
14
CO
2
incorporation was mediated by autotrophic microorganisms, the diversity and abundance of CO
2
-fixing bacteria and algae were investigated using clone library sequencing, terminal restriction fragment length polymorphism (T-RFLP), and quantitative PCR (qPCR) of the ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) gene (
cbbL
). Phylogenetic analysis showed that the dominant
cbbL
-containing bacteria were
Azospirillum lipoferum
,
Rhodopseudomonas palustris
,
Bradyrhizobium japonicum
,
Ralstonia eutropha
, and
cbbL
-containing chromophytic algae of the genera
Xanthophyta
and
Bacillariophyta
. Multivariate analyses of T-RFLP profiles revealed significant differences in
cbbL
-containing microbial communities between soils. Differences in
cbbL
gene diversity were shown to be correlated with differences in SOC content. Bacterial and algal
cbbL
gene abundances were between 10
6
and 10
8
and 10
3
to 10
5
copies g
−1
soil, respectively. Bacterial
cbbL
abundance was shown to be positively correlated with RubisCO activity (
r
= 0.853;
P
< 0.05), and both
cbbL
abundance and RubisCO activity were significantly related to the synthesis rates of [
14
C]SOC (
r
= 0.967 and 0.946, respectively;
P
< 0.01). These data offer new insights into the importance of microbial autotrophy in terrestrial C cycling.
Functional Cloning and Characterization of Antibiotic Resistance Genes from the Chicken Gut Microbiome
Authors: Zhou Wei , Wang Ying , Lin Jun 
Published: January 2012

ABSTRACT

Culture-independent sampling in conjunction with a functional cloning approach identified diverse antibiotic resistance genes for different classes of antibiotics in gut microbiomes from both conventionally raised and free-range chickens. Many of the genes are phylogenetically distant from known resistance genes. Two unique genes that conferred ampicillin and spectinomycin resistance were also functional in
Campylobacter
, a distant relative of the
Escherichia coli
host used to generate the genomic libraries.
Disulfide Bond Formation and Activation of Escherichia coli β-Galactosidase under Oxidizing Conditions
Authors: Seras-Franzoso Joaquin , Affentranger Roman , Ferrer-Navarro Mario , Daura Xavier , Villaverde Antonio , García-Fruitós Elena 
Published: January 2012

ABSTRACT

Escherichia coli
β-galactosidase is probably the most widely used reporter enzyme in molecular biology, cell biology, and biotechnology because of the easy detection of its activity. Its large size and tetrameric structure make this bacterial protein an interesting model for crystallographic studies and atomic mapping. In the present study, we investigate a version of
Escherichia coli
β-galactosidase produced under oxidizing conditions, in the cytoplasm of an Origami strain. Our data prove the activation of this microbial enzyme under oxidizing conditions and clearly show the occurrence of a disulfide bond in the β-galactosidase structure. Additionally, the formation of this disulfide bond is supported by the analysis of a homology model of the protein that indicates that two cysteines located in the vicinity of the catalytic center are sufficiently close for disulfide bond formation.
Influence of Cations on Growth of Thermophilic Geobacillus spp. and Anoxybacillus flavithermus in Planktonic Culture
Authors: Somerton Ben , Palmer Jon , Brooks John , Smolinski Edward , Lindsay Denise , Flint Steve 
Published: January 2012

ABSTRACT

Free ions of Na
+
, K
+
, Ca
2+
, and Mg
2+
influenced the optical density of planktonic cultures of thermophilic bacilli.
Anoxybacillus flavithermus
E16 and
Geobacillus
sp. strain F75 (milk powder manufacturing plant isolates) and
A. flavithermus
DSM 2641 and
G. thermoleovorans
DSM 5366 were studied. Ca
2+
and Mg
2+
were associated with increases in optical density more so than Na
+
and K
+
. Overall, it appeared that Ca
2+
and/or Mg
2+
was required for the production of protein in thermophilic bacilli, as shown by results obtained with
A. flavithermus
E16, which was selected for further study.
Roles of Protein Kinase A and Adenylate Cyclase in Light-Modulated Cellulase Regulation in Trichoderma reesei
Authors: Schuster André , Tisch Doris , Seidl-Seiboth Verena , Kubicek Christian P. , Schmoll Monika 
Published: January 2012

ABSTRACT

The cyclic AMP (cAMP) pathway represents a central signaling cascade with crucial functions in all organisms. Previous studies of
Trichoderma reesei
(anamorph of
Hypocrea jecorina
) suggested a function of cAMP signaling in regulation of cellulase gene expression. We were therefore interested in how the crucial components of this pathway, adenylate cyclase (ACY1) and cAMP-dependent protein kinase A (PKA), would affect cellulase gene expression. We found that both ACY1 and PKA catalytic subunit 1 (PKAC1) are involved in regulation of vegetative growth but are not essential for sexual development. Interestingly, our results showed considerably increased transcript abundance of cellulase genes in darkness compared to light (light responsiveness) upon growth on lactose. This effect is strongly enhanced in mutant strains lacking PKAC1 or ACY1. Comparison to the wild type showed that ACY1 has a consistently positive effect on cellulase gene expression in light and darkness, while PKAC1 influences transcript levels of cellulase genes positively in light but negatively in darkness. A function of PKAC1 in light-modulated cellulase gene regulation is also reflected by altered complex formation within the
cel6a/cbh2
promoter in light and darkness and in the absence of
pkac1
. Analysis of transcript levels of cellulase regulator genes indicates that the regulatory output of the cAMP pathway may be established via adjustment of XYR1 abundance. Consequently, both adenylate cyclase and protein kinase A are involved in light-modulated cellulase gene expression in
T. reesei
and have a dampening effect on the light responsiveness of this process.
Population Structure of and Mycotoxin Production by Fusarium graminearum from Maize in South Korea
Authors: Lee Jungkwan , Kim Hun , Jeon Jae-Jin , Kim Hye-Seon , Zeller Kurt A. , Carter Laurel L. A. , Leslie John F. , Lee Yin-Won 
Published: January 2012

ABSTRACT

Fusarium graminearum
(
Gibberella zeae
) is an important pathogen of wheat, maize, barley, and rice in South Korea, and harvested grain often is contaminated with trichothecenes such as deoxynivalenol and nivalenol. In this study, we examined 568 isolates of
F. graminearum
collected from maize at eight locations in South Korea. We used amplified fragment length polymorphisms (AFLPs) to identify four lineages (2, 3, 6, and 7); lineage 7 was the most common (75%), followed by lineage 6 (12%), lineage 3 (12%), and lineage 2 (1%). The genetic identity among populations was high (>0.98), and the effective migration rate between locations was higher than that between lineages. Female fertility varied by lineage: all lineage 7 isolates were fertile, while 70%, 26%, and 14% of the isolates in lineages 6, 3, and 2, respectively, were fertile. All lineage 3 and lineage 7 isolates produced deoxynivalenol, whereas most lineage 2 and 6 isolates produced nivalenol. Genotypic diversity in lineage 3 and lineage 6 populations is similar to that found in previously described Korean rice populations, but genotypic diversity in lineage 7 is much lower, even though similar levels of gene flow occur between lineage 7 populations. We conclude that lineage 7 was relatively recently introduced into South Korea, perhaps accompanying imported maize seeds.
Chimeric Phage Lysins Act Synergistically with Lysostaphin To Kill Mastitis-Causing Staphylococcus aureus in Murine Mammary Glands
Authors: Schmelcher Mathias , Powell Anne M. , Becker Stephen C. , Camp Mary J. , Donovan David M. 
Published: January 2012

ABSTRACT

Staphylococci cause bovine mastitis, with
Staphylococcus aureus
being responsible for the majority of the mastitis-based losses to the dairy industry (up to $2 billion/annum). Treatment is primarily with antibiotics, which are often ineffective and potentially contribute to resistance development. Bacteriophage endolysins (peptidoglycan hydrolases) present a promising source of alternative antimicrobials. Here we evaluated two fusion proteins consisting of the streptococcal λSA2 endolysin endopeptidase domain fused to staphylococcal cell wall binding domains from either lysostaphin (λSA2-E-Lyso-SH3b) or the staphylococcal phage K endolysin, LysK (λSA2-E-LysK-SH3b). We demonstrate killing of 16 different
S. aureus
mastitis isolates, including penicillin-resistant strains, by both constructs. At 100 μg/ml in processed cow milk, λSA2-E-Lyso-SH3b and λSA2-E-LysK-SH3b reduced the
S. aureus
bacterial load by 3 and 1 log units within 3 h, respectively, compared to a buffer control. In contrast to λSA2-E-Lyso-SH3b, however, λSA2-E-LysK-SH3b permitted regrowth of the pathogen after 1 h. In a mouse model of mastitis, infusion of 25 μg of λSA2-E-Lyso-SH3b or λSA2-E-LysK-SH3b into mammary glands reduced
S. aureus
CFU by 0.63 or 0.81 log units, compared to >2 log for lysostaphin. Both chimeras were synergistic with lysostaphin against
S. aureus
in plate lysis checkerboard assays. When tested in combination in mice, λSA2-E-LysK-SH3b and lysostaphin (12.5 μg each/gland) caused a 3.36-log decrease in CFU. Furthermore, most protein treatments reduced gland wet weights and intramammary tumor necrosis factor alpha (TNF-α) concentrations, which serve as indicators of inflammation. Overall, our animal model results demonstrate the potential of fusion peptidoglycan hydrolases as antimicrobials for the treatment of
S. aureus
-induced mastitis.
Abundance and Single-Cell Activity of Heterotrophic Bacterial Groups in the Western Arctic Ocean in Summer and Winter
Authors: Nikrad Mrinalini P. , Cottrell M. T. , Kirchman D. L. 
Published: January 2012

ABSTRACTEnvironmental conditions in the western Arctic Ocean range from constant light and nutrient depletion in summer to complete darkness and sea ice cover in winter. This seasonal environmental variation is likely to have an effect on the use of dissolved organic matter (DOM) by heterotrophic bacteria in surface water. However, this effect is not well studied and we know little about the activity of specific bacterial clades in the surface oceans. The use of DOM by three bacterial subgroups in both winter and summer was examined by microautoradiography combined with fluorescencein situhybridization. We found selective use of substrates by these groups, although the abundances of Ant4D3 (AntarcticGammaproteobacteria),Polaribacter(Bacteroidetes), and SAR11 (Alphaproteobacteria) were not different between summer and winter in the Beaufort and Chukchi Seas. The number of cells taking up glucose within all three bacterial groups decreased significantly from summer to winter, while the percentage of cells using leucine did not show a clear pattern between seasons. The uptake of the amino acid mix increased substantially from summer to winter by the Ant4D3 group, although such a large increase in uptake was not seen for the other two groups. Use of glucose by bacteria, but not use of leucine or the amino acid mix, related strongly to inorganic nutrients, chlorophylla, and other environmental factors. Our results suggest a switch in use of dissolved organic substrates from summer to winter and that the three phylogenetic subgroups examined fill different niches in DOM use in the two seasons.
Construction and Characterization of a Gradually Inducible Expression Vector for Halobacterium salinarum, Based on the
kdp
Promoter
Authors: Kixmüller Dorthe , Greie Jörg-Christian 
Published: January 2012

ABSTRACT

Gradually inducible expression vectors which are governed by variations of growth conditions are powerful tools for gene expression of conditionally lethal mutants. Furthermore, controlled expression allows monitoring of overproduction of proteins at various stages in their expressing hosts. For
Halobacterium salinarum
, which is often used as a paradigm for halophilic archaea, such an inducible expression system is not available to date. Here we show that the
kdp
promoter (P
kdp
), which facilitates gene expression upon K
+
limitation, can be used to establish such a system for molecular applications. P
kdp
features a rather high expression rate, with an approximately 50-fold increase that can be easily varied by K
+
concentrations in the growth medium. Besides the construction of an expression vector, our work describes the characterization of expression patterns and, thus, offers a gradually inducible expression system to the scientific community.
Three Types of Taxis Used in the Response of
Acidovorax
sp. Strain JS42 to 2-Nitrotoluene
Authors: Rabinovitch-Deere Christine A. , Parales Rebecca E. 
Published: January 2012

ABSTRACT

Acidovorax
sp. strain JS42 is able to utilize 2-nitrotoluene (2NT) as its sole carbon, nitrogen, and energy source. We report here that strain JS42 is chemotactic to 2NT and that the response is increased when cells are grown on compounds such as 2NT that are known to induce the first step of 2NT degradation. Assays with JS42 mutants unable to oxidize 2NT showed that the first step of 2NT metabolism was required for the induced response, but not for a portion of the constitutive response, indicating that 2NT itself is an attractant. The 2NT metabolite nitrite was shown to be a strong attractant for strain JS42, and sufficient nitrite was produced during the taxis assay to account for a large part of the induced response. A mutant with an inactivated
ntdY
gene, which is located adjacent to the 2NT degradation genes and codes for a putative methyl-accepting chemotaxis protein, showed a defect in taxis toward 2NT that may involve a reduced response to nitrite. Responses of a mutant defective for the energy-taxis receptor, Aer, indicated that a functional
aer
gene is required for a substantial part of the wild-type induced response to 2NT. In summary, strain JS42 utilizes three types of taxis to sense and respond to 2NT: constitutive 2NT-specific chemotaxis to directly sense 2NT, metabolism-dependent nitrite-specific chemotaxis that may be mediated by NtdY, and energy taxis mediated by Aer.
Differential Isotopic Fractionation during Cr(VI) Reduction by an Aquifer-Derived Bacterium under Aerobic versus Denitrifying Conditions
Authors: Han Ruyang , Qin Liping , Brown Shaun T. , Christensen John N. , Beller Harry R. 
Published: January 2012

ABSTRACT

We studied Cr isotopic fractionation during Cr(VI) reduction by
Pseudomonas stutzeri
strain RCH2. Despite the fact that strain RCH2 reduces Cr(VI) cometabolically under both aerobic and denitrifying conditions and at similar specific rates, fractionation was markedly different under these two conditions (ε was ∼2‰ aerobically and ∼0.4‰ under denitrifying conditions).
Geraniol and Geranial Dehydrogenases Induced in Anaerobic Monoterpene Degradation by Castellaniella defragrans
Authors: Lüddeke Frauke , Wülfing Annika , Timke Markus , Germer Frauke , Weber Johanna , Dikfidan Aytac , Rahnfeld Tobias , Linder Dietmar , Meyerdierks Anke , Harder Jens 
Published: January 2012

ABSTRACT

Castellaniella defragrans
is a
Betaproteobacterium
capable of coupling the oxidation of monoterpenes with denitrification. Geraniol dehydrogenase (GeDH) activity was induced during growth with limonene in comparison to growth with acetate. The N-terminal sequence of the purified enzyme directed the cloning of the corresponding open reading frame (ORF), the first bacterial gene for a GeDH (
geoA
, for
ge
raniol
o
xidation pathway). The
C. defragrans
geraniol dehydrogenase is a homodimeric enzyme that affiliates with the zinc-containing benzyl alcohol dehydrogenases in the superfamily of medium-chain-length dehydrogenases/reductases (MDR). The purified enzyme most efficiently catalyzes the oxidation of perillyl alcohol (
k
cat
/
K
m
= 2.02 × 10
6
M
−1
s
−1
), followed by geraniol (
k
cat
/
K
m
= 1.57 × 10
6
M
−1
s
−1
). Apparent
K
m
values of <10 μM are consistent with an
in vivo
toxicity of geraniol above 5 μM. In the genetic vicinity of
geoA
is a putative aldehyde dehydrogenase that was named
geoB
and identified as a highly abundant protein during growth with phellandrene. Extracts of
Escherichia coli
expressing
geoB
demonstrated
in vitro
a geranial dehydrogenase (GaDH) activity. GaDH activity was independent of coenzyme A. The irreversible formation of geranic acid allows for a metabolic flux from β-myrcene via linalool, geraniol, and geranial to geranic acid.
Molecular Basis and Phylogenetic Implications of Deoxycylindrospermopsin Biosynthesis in the Cyanobacterium Raphidiopsis curvata
Authors: Jiang Yongguang , Xiao Peng , Yu Gongliang , Sano Tomoharu , Pan Qianqian , Li Renhui 
Published: January 2012

ABSTRACT

New insights into the distribution and biochemistry of the cyanotoxin cylindrospermopsin (CYN) have been provided by the recent determination of its biosynthesis gene cluster (
cyr
) in several cyanobacterial species.
Raphidiopsis curvata
CHAB1150 isolated from China was analyzed for CYN analogues. Only 7-deoxy-CYN was detected in the cell extracts. The
cyr
gene cluster of
R. curvata
CHAB1150 was sequenced, and the
cyr
genes of this strain were found to have extremely high similarities (96% to 100%) to those from other nostocalean species. These species include
Cylindrospermopsis raciborskii
AWT205,
Aphanizomenon
sp. strain 10E6, and
Aphanizomenon ovalisporum
ILC-146. Insertion mutation was identified within the
cyrI
gene, and transcripts of
cyrI
and another functional gene
cyrJ
were detected in
R. curvata
CHAB1150. General congruence between the phylogenetic trees based on both
cyr
and 16S
rrn
was displayed. Neutral evolution was found on the whole sequences of the
cyr
genes, and 0 to 89 negative selected codons were detected in each gene. Therefore, the function of CyrI is to catalyze the oxygenation of 7-deoxy-CYN in CYN biosynthesis. The transcripts of the mutated
cyrI
gene may result from polycistronic transcription. The high conservation of the
cyr
genes may be ascribed to purifying selection and horizontal gene transfer.
Toward Biosynthetic Design and Implementation of Escherichia coli-Derived Paclitaxel and Other Heterologous Polyisoprene Compounds
Authors: Jiang Ming , Stephanopoulos Gregory , Pfeifer Blaine A. 
Published: January 2012

ABSTRACT

Escherichia coli
offers unparalleled engineering capacity in the context of heterologous natural product biosynthesis. However, as with other heterologous hosts, cellular metabolism must be designed or redesigned to support final compound formation. This task is at once complicated and aided by the fact that the cell does not natively produce an abundance of natural products. As a result, the metabolic engineer avoids complicated interactions with native pathways closely associated with the outcome of interest, but this convenience is tempered by the need to implement the required metabolism to allow functional biosynthesis. This review focuses on engineering
E. coli
for the purpose of polyisoprene formation, as it is related to isoprenoid compounds currently being pursued through a heterologous approach. In particular, the review features the compound paclitaxel and early efforts to design and overproduce intermediates through
E. coli
.
Cloning and Sequencing of Inulinase and β-Fructofuranosidase Genes of a Deep-Sea Microbulbifer Species and Properties of Recombinant Enzymes
Authors: Kobayashi Tohru , Uchimura Kohsuke , Deguchi Shigeru , Horikoshi Koki 
Published: January 2012

ABSTRACT

An inulinase-producing
Microbulbifer
sp. strain, JAM-3301, was isolated from a deep-sea sediment. An inulin operon that contained three open reading frames was cloned and sequenced. Two of the three genes were expressed. One product was an endo-inulinase, and the other was a β-fructofuranosidase. Both enzymes worked together to effectively degrade inulin.