bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Clinical Candidate VT-1161 Is a Highly Potent Inhibitor of Candida albicans CYP51 but Fails To Bind the Human Enzyme
Authors: Warrilow A. G. S. , Hull C. M. , Parker J. E. , Garvey E. P. , Hoekstra W. J. , Moore W. R. , Schotzinger R. J. , Kelly D. E. , Kelly S. L. 
Published: September 2014

ABSTRACT

The binding and cytochrome P45051 (CYP51) inhibition properties of a novel antifungal compound, VT-1161, against purified recombinant
Candida albicans
CYP51 (
ERG11
) and
Homo sapiens
CYP51 were compared with those of clotrimazole, fluconazole, itraconazole, and voriconazole. VT-1161 produced a type II binding spectrum with
Candida albicans
CYP51, characteristic of heme iron coordination. The binding affinity of VT-1161 for
Candida albicans
CYP51 was high (dissociation constant [

K
d

], ≤39 nM) and similar to that of the pharmaceutical azole antifungals (

K
d

, ≤50 nM). In stark contrast, VT-1161 at concentrations up to 86 μM did not perturb the spectrum of recombinant human CYP51, whereas all the pharmaceutical azoles bound to human CYP51. In reconstitution assays, VT-1161 inhibited
Candida albicans
CYP51 activity in a tight-binding fashion with a potency similar to that of the pharmaceutical azoles but failed to inhibit the human enzyme at the highest concentration tested (50 μM). In addition, VT-1161 (MIC = 0.002 μg ml
−1
) had a more pronounced fungal sterol disruption profile (increased levels of methylated sterols and decreased levels of ergosterol) than the known CYP51 inhibitor voriconazole (MIC = 0.004 μg ml
−1
). Furthermore, VT-1161 weakly inhibited human CYP2C9, CYP2C19, and CYP3A4, suggesting a low drug-drug interaction potential. In summary, VT-1161 potently inhibited
Candida albicans
CYP51 and culture growth but did not inhibit human CYP51, demonstrating a >2,000-fold selectivity. This degree of potency and selectivity strongly supports the potential utility of VT-1161 in the treatment of
Candida
infections.
Evaluation of a Loop-Mediated Isothermal Amplification-Based Methodology To Detect Carbapenemase Carriage in Acinetobacter Clinical Isolates
Authors: Vergara Andrea , Zboromyrska Yuliya , Mosqueda Noraida , Morosini María Isabel , García-Fernández Sergio , Roca Ignasi , Cantón Rafael , Marco Francesc , Vila Jordi 
Published: September 2014

ABSTRACT

Carbapenem-resistant
Acinetobacter baumannii
is a major source of nosocomial infections worldwide and is mainly associated with the acquisition of OXA-type carbapenemases and, to a lesser extent, metallo-β-lactamases (MBLs). In this study, 82 nonepidemiologically related
Acinetobacter
strains carrying different types of OXA or MBL enzymes were tested using the Eazyplex system, a loop-mediated isothermal amplification (LAMP)-based method to rapidly detect carbapenemase carriage. The presence/absence of carbapenem-hydrolyzing enzymes was correctly determined for all isolates in <30 min.
Role of Toll-Like Receptor 13 in Innate Immune Recognition of Group B Streptococci
Authors: Signorino Giacomo , Mohammadi Nastaran , Patanè Francesco , Buscetta Marco , Venza Mario , Venza Isabella , Mancuso Giuseppe , Midiri Angelina , Alexopoulou Lena , Teti Giuseppe , Biondo Carmelo , Beninati Concetta 
Published: September 2014

ABSTRACTMurine Toll-like receptor 13 (TLR13), an endosomal receptor that is not present in humans, is activated by an unmethylated motif present in the large ribosomal subunit of bacterial RNA (23S rRNA). Little is known, however, of the impact of TLR13 on antibacterial host defenses. Here we examined the role of this receptor in the context of infection induced by the model pathogen group B streptococcus (GBS). To this end, we used bacterial strains masked from TLR13 recognition by virtue of constitutive expression of the ErmC methyltransferase, which results in dimethylation of the 23S rRNA motif at a critical adenine residue. We found that TLR13-mediated rRNA recognition was required for optimal induction of tumor necrosis factor alpha and nitrous oxide in dendritic cell and macrophage cultures stimulated with heat-killed bacteria or purified bacterial RNA. However, TLR13-dependent recognition was redundant when live bacteria were used as a stimulus. Moreover, masking bacterial rRNA from TLR13 recognition did not increase the ability of GBS to avoid host defenses and replicatein vivo. In contrast, increased susceptibility to infection was observed under conditions in which signaling by all endosomal TLRs was abolished, i.e., in mice with a loss-of-function mutation in the chaperone protein UNC93B1. Our data lend support to the conclusion that TLR13 participates in GBS recognition, although blockade of the function of this receptor can be compensated for by other endosomal TLRs. Lack of selective pressure by bacterial infections might explain the evolutionary loss of TLR13 in humans. However, further studies using different bacterial species are needed to prove this hypothesis.
Mycobacterium tuberculosis MtrB Sensor Kinase Interactions with FtsI and Wag31 Proteins Reveal a Role for MtrB Distinct from That Regulating MtrA Activities
Authors: Plocinska Renata , Martinez Luis , Gorla Purushotham , Pandeeti Emmanuel , Sarva Krishna , Blaszczyk Ewelina , Dziadek Jaroslaw , Madiraju Murty V. , Rajagopalan Malini 
Published: September 2014

ABSTRACT

The septal association of
Mycobacterium tuberculosis
MtrB, the kinase partner of the MtrAB two-component signal transduction system, is necessary for the optimal expression of the MtrA regulon targets, including
ripA
,
fbpB
, and
ftsI
, which are involved in cell division and cell wall synthesis. Here, we show that MtrB, irrespective of its phosphorylation status, interacts with Wag31, whereas only phosphorylation-competent MtrB interacts with FtsI. We provide evidence that FtsI depletion compromises the MtrB septal assembly and MtrA regulon expression; likewise, the absence of MtrB compromises FtsI localization and, possibly, FtsI activity. We conclude from these results that FtsI and MtrB are codependent for their activities and that FtsI functions as a positive modulator of MtrB activation and MtrA regulon expression. In contrast to FtsI, Wag31 depletion does not affect MtrB septal assembly and MtrA regulon expression, whereas the loss of MtrB increased Wag31 localization and the levels of PknA/PknB (PknA/B) serine-threonine protein kinase-mediated Wag31 phosphorylation. Interestingly, we found that FtsI decreased levels of phosphorylated Wag31 (Wag31∼P) and that MtrB interacted with PknA/B. Overall, our results indicate that MtrB interactions with FtsI, Wag31, and PknA/B are required for its optimal localization, MtrA regulon expression, and phosphorylation of Wag31. Our results emphasize a new role for MtrB in cell division and cell wall synthesis distinct from that regulating the MtrA phosphorylation activities.
A Novel, Broad-Range, CTXΦ-Derived Stable Integrative Expression Vector for Functional Studies
Authors: Das Bhabatosh , Kumari Reena , Pant Archana , Sen Gupta Sourav , Saxena Shruti , Mehta Ojasvi , Nair Gopinath Balakrish 
Published: September 2014

ABSTRACT

CTXΦ, a filamentous vibriophage encoding cholera toxin, uses a unique strategy for its lysogeny. The single-stranded phage genome forms intramolecular base-pairing interactions between two inversely oriented XerC and XerD binding sites (XBS) and generates a functional phage attachment site,
attP
(
+
), for integration. The
attP
(
+
) structure is recognized by the host-encoded tyrosine recombinases XerC and XerD (XerCD), which enables irreversible integration of CTXΦ into the chromosome dimer resolution site (
dif
) of
Vibrio cholerae
. The
dif
site and the XerCD recombinases are widely conserved in bacteria. We took advantage of these conserved attributes to develop a broad-host-range integrative expression vector that could irreversibly integrate into the host chromosome using XerCD recombinases without altering the function of any known open reading frame (ORF). In this study, we engineered two different arabinose-inducible expression vectors, pBD62 and pBD66, using XBS of CTXΦ. pBD62 replicates conditionally and integrates efficiently into the
dif
of the bacterial chromosome by site-specific recombination using host-encoded XerCD recombinases. The expression level of the gene of interest could be controlled through the P
BAD
promoter by modulating the functions of the vector-encoded transcriptional factor AraC. We validated the irreversible integration of pBD62 into a wide range of pathogenic and nonpathogenic bacteria, such as
V. cholerae
,
Vibrio fluvialis
,
Vibrio parahaemolyticus
,
Escherichia coli
,
Salmonella enterica
, and
Klebsiella pneumoniae
. Gene expression from the P
BAD
promoter of integrated vectors was confirmed in
V. cholerae
using the well-studied reporter genes
mCherry
,
eGFP
, and
lacZ
.
Methylamine Utilization via the
N
-Methylglutamate Pathway in Methylobacterium extorquens PA1 Involves a Novel Flow of Carbon through C
1
Assimilation and Dissimilation Pathways
Authors: Nayak Dipti D. , Marx Christopher J. 
Published: September 2014

ABSTRACT

Methylotrophs grow on reduced single-carbon compounds like methylamine as the sole source of carbon and energy. In
Methylobacterium extorquens
AM1, the best-studied aerobic methylotroph, a periplasmic methylamine dehydrogenase that catalyzes the primary oxidation of methylamine to formaldehyde has been examined in great detail. However, recent metagenomic data from natural ecosystems are revealing the abundance and importance of lesser-known routes, such as the
N
-methylglutamate pathway, for methylamine oxidation. In this study, we used
M. extorquens
PA1, a strain that is closely related to
M. extorquens
AM1 but is lacking methylamine dehydrogenase, to dissect the genetics and physiology of the ecologically relevant
N
-methylglutamate pathway for methylamine oxidation. Phenotypic analyses of mutants with null mutations in genes encoding enzymes of the
N
-methylglutamate pathway suggested that γ-glutamylmethylamide synthetase is essential for growth on methylamine as a carbon source but not as a nitrogen source. Furthermore, analysis of
M. extorquens
PA1 mutants with defects in methylotrophy-specific dissimilatory and assimilatory modules suggested that methylamine use via the
N
-methylglutamate pathway requires the tetrahydromethanopterin (H
4
MPT)-dependent formaldehyde oxidation pathway but not a complete tetrahydrofolate (H
4
F)-dependent formate assimilation pathway. Additionally, we present genetic evidence that formaldehyde-activating enzyme (FAE) homologs might be involved in methylotrophy. Null mutants of FAE and homologs revealed that FAE and FAE2 influence the growth rate and FAE3 influences the yield during the growth of
M. extorquens
PA1 on methylamine.
Unique Genomic Arrangements in an Invasive Serotype M23 Strain of Streptococcus pyogenes Identify Genes That Induce Hypervirulence
Authors: Bao Yunjuan , Liang Zhong , Booyjzsen Claire , Mayfield Jeffrey A. , Li Yang , Lee Shaun W. , Ploplis Victoria A. , Song Hui , Castellino Francis J. 
Published: September 2014

ABSTRACT

The first genome sequence of a group A
Streptococcus pyogenes
serotype M23 (
emm23
) strain (M23ND), isolated from an invasive human infection, has been completed. The genome of this opacity factor-negative (SOF
−
) strain is composed of a circular chromosome of 1,846,477 bp. Gene profiling showed that this strain contained six phage-encoded and 24 chromosomally inherited well-known virulence factors, as well as 11 pseudogenes. The bacterium has acquired four large prophage elements, ΦM23ND.1 to ΦM23ND.4, harboring genes encoding streptococcal superantigen (
ssa
), streptococcal pyrogenic exotoxins (
speC
,
speH
, and
speI
), and DNases (
spd1
and
spd3
), with phage integrase genes being present at one flank of each phage insertion, suggesting that the phages were integrated by horizontal gene transfer. Comparative analyses revealed unique large-scale genomic rearrangements that result in genomic rearrangements that differ from those of previously sequenced GAS strains. These rearrangements resulted in an imbalanced genomic architecture and translocations of chromosomal virulence genes. The
covS
sensor in M23ND was identified as a pseudogene, resulting in the attenuation of
speB
function and increased expression of the genes for the chromosomal virulence factors multiple-gene activator (
mga
), M protein (
emm23
), C5a peptidase (
scpA
), fibronectin-binding proteins (
sfbI
and
fbp54
), streptolysin O (
slo
), hyaluronic acid capsule (
hasA
), streptokinase (
ska
), and DNases (
spd
and
spd3
), which were verified by PCR. These genes are responsible for facilitating host epithelial cell binding and and/or immune evasion, thus further contributing to the virulence of M23ND. In conclusion, strain M23ND has become highly pathogenic as the result of a combination of multiple genetic factors, particularly gene composition and mutations, prophage integrations, unique genomic rearrangements, and regulated expression of critical virulence factors.
Charge Requirements of Lipid II Flippase Activity in Escherichia coli
Authors: Butler Emily K. , Tan Wee Boon , Joseph Hildy , Ruiz Natividad 
Published: September 2014

ABSTRACT

Peptidoglycan (PG) is an extracytoplasmic glycopeptide matrix essential for the integrity of the envelope of most bacteria. The PG building block is a disaccharide-pentapeptide that is synthesized as a lipid-linked precursor called lipid II. The translocation of the amphipathic lipid II across the cytoplasmic membrane is required for subsequent incorporation of the disaccharide-pentapeptide into PG. In
Escherichia coli
, the essential inner membrane protein MurJ is the lipid II flippase. Previous studies showed that 8 charged residues in the central cavity region of MurJ are crucial for function. Here, we completed the functional analysis of all 57 charged residues in MurJ and demonstrated that the respective positive or negative charge of the 8 aforementioned residues is required for proper MurJ function. Loss of the negative charge in one of these residues, D39, causes a severe defect in MurJ biogenesis; by engineering an intragenic suppressor mutation that restores MurJ biogenesis, we found that this charge is also essential for MurJ function. Because of the low level of homology between MurJ and putative orthologs from Gram-positive bacteria, we explored the conservation of these 8 charged residues in YtgP, a homolog from
Streptococcus pyogenes
. We found that only 3 positive charges are similarly positioned and essential in YtgP; YtgP possesses additional charged residues within its predicted cavity that are essential for function and conserved among Gram-positive bacteria. From these data, we hypothesize that some charged residues in the cavity region of MurJ homologs are required for interaction with lipid II and/or energy coupling during transport.
Characterization of Five β-Glycoside Hydrolases from Cellulomonas fimi ATCC 484
Authors: Gao Juan , Wakarchuk Warren 
Published: September 2014

ABSTRACT

The Gram-positive bacterium
Cellulomonas fimi
produces a large array of carbohydrate-active enzymes. Analysis of the collection of carbohydrate-active enzymes from the recent genome sequence of
C. fimi
ATCC 484 shows a large number of uncharacterized genes for glycoside hydrolase (GH) enzymes potentially involved in biomass utilization. To investigate the enzymatic activity of potential β-glucosidases in
C. fimi
, genes encoding several GH3 enzymes and one GH1 enzyme were cloned and recombinant proteins were expressed in
Escherichia coli
. Biochemical analysis of these proteins revealed that the enzymes exhibited different substrate specificities for
para
-nitrophenol-linked substrates (
p
NP), disaccharides, and oligosaccharides. Celf_2726 encoded a bifunctional enzyme with β-
d
-xylopyranosidase and α-
l
-arabinofuranosidase activities, based on
p
NP-linked substrates (CfXyl3A). Celf_0140 encoded a β-
d
-glucosidase with activity on β-1,3- and β-1,6-linked glucosyl disaccharides as well as
p
NP-β-Glc (CfBgl3A). Celf_0468 encoded a β-
d
-glucosidase with hydrolysis of
p
NP-β-Glc and hydrolysis/transglycosylation activities only on β-1,6-linked glucosyl disaccharide (CfBgl3B). Celf_3372 encoded a GH3 family member with broad aryl-β-
d
-glycosidase substrate specificity. Celf_2783 encoded the GH1 family member (CfBgl1), which was found to hydrolyze
p
NP-β-Glc/Fuc/Gal, as well as cellotetraose and cellopentaose. CfBgl1 also had good activity on β-1,2- and β-1,3-linked disaccharides but had only very weak activity on β-1,4/6-linked glucose.
Role of Adaptor TrfA and ClpPC in Controlling Levels of SsrA-Tagged Proteins and Antitoxins in Staphylococcus aureus
Authors: Donegan Niles P. , Marvin Jonathan S. , Cheung Ambrose L. 
Published: September 2014

ABSTRACTStaphylococcus aureusresponds to changing extracellular environments in part by adjusting its proteome through alterations of transcriptional priorities and selective degradation of the preexisting pool of proteins. InBacillus subtilis, the proteolytic adaptor protein MecA has been shown to play a role in assisting with the proteolytic degradation of proteins involved in competence and the oxidative stress response. However, the targets of TrfA, the MecA homolog inS. aureus, have not been well characterized. In this work, we investigated how TrfA assists chaperones and proteases to regulate the proteolysis of several classes of proteins inS. aureus. By fusing the last 3 amino acids of the SsrA degradation tag to Venus, a rapidly folding yellow fluorescent protein, we obtained both fluorescence-based and Western blot assay-based evidence that TrfA and ClpCP are the adaptor and protease, respectively, responsible for the degradation of the SsrA-tagged protein inS. aureus. Notably, the impact of TrfA on degradation was most prominent during late log phase and early stationary phase, due in part to a combination of transcriptional regulation and proteolytic degradation of TrfA by ClpCP. We also characterized the temporal transcriptional regulation governing TrfA activity, wherein Spx, a redox-sensitive transcriptional regulator degraded by ClpXP, activatestrfAtranscription while repressing its own promoter. Finally, the scope of TrfA-mediated proteolysis was expanded by identifying TrfA as the adaptor that works with ClpCP to degrade antitoxins inS. aureus. Together, these results indicate that the adaptor TrfA adds temporal nuance to protein degradation by ClpCP inS. aureus.
Serum Interleukin-8 Reflects Tumor Burden and Treatment Response across Malignancies of Multiple Tissue Origins
Authors: Sanmamed Miguel F. , Carranza-Rua Omar , Alfaro Carlos , Oñate Carmen , Martín-Algarra Salvador , Perez Guiomar , Landazuri Sara F. , Gonzalez Álvaro , Gross Stefanie , Rodriguez Inmaculada , Muñoz-Calleja Cecilia , Rodríguez-Ruiz María , Sangro Bruno , López-Picazo José M. , Rizzo Manglio , Mazzolini Guillermo , Pascual Juan I. , Andueza Maria Pilar , Perez-Gracia Jose Luis , Melero Ignacio 
Published: September 2014

AbstractPurpose: Interleukin-8 (IL8) is a chemokine produced by malignant cells of multiple cancer types. It exerts various functions in shaping protumoral vascularization and inflammation/immunity. We evaluated sequential levels of serum IL8 in preclinical tumor models and in patients to assess its ability to estimate tumor burden.Experimental Design: IL8 levels were monitored by sandwich ELISAs in cultured tumor cells supernatants, tumor-xenografted mice serum, and in samples from 126 patients with cancer. We correlated IL8 serum levels with baseline tumor burden and with treatment-induced changes in tumor burden, as well as with prognosis.Results: IL8 concentrations correlated with the number of IL8-producing tumor cells in culture. In xenografted neoplasms, IL8 serum levels rapidly dropped after surgical excision, indicating an accurate correlation with tumor burden. In patients with melanoma (n = 16), renal cell carcinoma (RCC; n = 23), non–small cell lung cancer (NSCLC; n = 21), or hepatocellular carcinoma (HCC; n = 30), serum IL8 concentrations correlated with tumor burden and stage, survival (melanoma, n = 16; RCC, n = 23; HCC, n = 33), and objective responses to therapy, including those to BRAF inhibitors (melanoma, n = 16) and immunomodulatory monoclonal antibodies (melanoma, n = 8). IL8 concentrations in urine (n = 18) were mainly elevated in tumors with direct contact with the urinary tract.Conclusions: IL8 levels correlate with tumor burden in preclinical models and in patients with cancer. IL8 is a potentially useful biomarker to monitor changes in tumor burden following anticancer therapy, and has prognostic significance. Clin Cancer Res; 20(22); 5697–707. ©2014 AACR.
Long-Lived Response of the Midlatitude Circulation and Storm Tracks to Pulses of Tropical Heating
Authors: Branstator Grant 
Published: September 2014

Abstract
The midlatitude response to localized equatorial heating events that last 2 days is examined through experimentation with an atmospheric general circulation model. Such responses are argued to be important because many tropical rainfall events only last a short time and because the responses to such pulses serve as building blocks with which to study the impacts of more general heating fluctuations.
The experiments indicate that short-lived heating produces responses in midlatitudes at locations far removed from the source and these responses persist much longer than the pulses themselves. Indeed pulse forcing, which is essentially white in time, produces upper-tropospheric responses that have an e-damping time of almost a week and that are detectable for more than two weeks in the experiments. Moreover the upper-tropospheric structure of the reaction to short pulses is remarkably similar to the reaction to steady tropical heating, including having a preference for occurring at special geographical locations and being composed of recurring patterns that resemble the leading patterns of responses to steady heating. This similarity is argued to be a consequence of the responses to pulses having little or no phase propagation in the extratropics. The impact of short-lived tropical heating also produces a persistent response in midlatitude surface fields and in the statistics of synoptic eddies.
The implications these results have for subseasonal variability are discussed. These include 1) the potential for improving subseasonal prediction through improved assimilation and short-range forecasts of tropical precipitation and 2) the difficulties involved in attributing subseasonal midlatitude events to tropical heating.
High-Water Alerts from Coinciding High Astronomical Tide and High Mean Sea Level Anomaly in the Pacific Islands Region
Authors: Stephens Scott A. , Bell Robert G. , Ramsay Douglas , Goodhue Nigel 
Published: September 2014

AbstractA technique to produce high-water alerts from coinciding high astronomical tide and high mean sea level anomaly is demonstrated for the Pacific Islands region. Low-lying coastal margins are vulnerable to episodic inundation that often coincides with times of higher-than-normal high tides. Prior knowledge of the dates of the highest tides can assist with efforts to minimize the impacts of increased exposure to inundation. It is shown that the climate-driven mean sea level anomaly is an important component of total sea level elevation in the Pacific Islands region, which should be accounted for in medium-term (1–7 months) sea level forecasts. An empirical technique is applied to develop a mean sea level–adjusted high-water alert calendar that accounts for both sea level components and provides a practical tool to assist with coastal inundation hazard planning and management.