bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of a Residue Affecting Fatty Alcohol Selectivity in Wax Ester Synthase
Authors: Barney Brett M. , Mann Rachel L. , Ohlert Janet M. 
Published: October 2012

ABSTRACT

The terminal enzyme in the bacterial wax ester biosynthetic pathway is the bifunctional wax ester synthase/acyl-coenzyme A:diacylglycerol acyltransferase (WS/DGAT), which utilizes a fatty alcohol and a fatty acyl-coenzyme A (CoA) to synthesize the corresponding wax ester. In this report, we identify a specific residue in WS/DGAT enzymes obtained from
Marinobacter aquaeolei
VT8 and
Acinetobacter baylyi
that alters fatty alcohol selectivity and kinetic parameters when modified to alternative residues.
Biochemical Properties and Crystal Structure of a β-Phenylalanine Aminotransferase from Variovorax paradoxus
Authors: Crismaru Ciprian G. , Wybenga Gjalt G. , Szymanski Wiktor , Wijma Hein J. , Wu Bian , Bartsch Sebastian , de Wildeman Stefaan , Poelarends Gerrit J. , Feringa Ben L. , Dijkstra Bauke W. , Janssen Dick B. 
Published: October 2012

ABSTRACTBy selective enrichment, we isolated a bacterium that can use β-phenylalanine as a sole nitrogen source. It was identified by 16S rRNA gene sequencing as a strain ofVariovorax paradoxus. Enzyme assays revealed an aminotransferase activity. Partial genome sequencing and screening of a cosmid DNA library resulted in the identification of a 1,302-bp aminotransferase gene, which encodes a 46,416-Da protein. The gene was cloned and overexpressed inEscherichia coli. The recombinant enzyme was purified and showed a specific activity of 17.5 U mg−1for (S)-β-phenylalanine at 30°C and 33 U mg−1at the optimum temperature of 55°C. The β-specific aminotransferase exhibits a broad substrate range, acceptingortho-,meta-, andpara-substituted β-phenylalanine derivatives as amino donors and 2-oxoglutarate and pyruvate as amino acceptors. The enzyme is highly enantioselective toward (S)-β-phenylalanine (enantioselectivity [E], >100) and derivatives thereof with different substituents on the phenyl ring, allowing the kinetic resolution of various racemic β-amino acids to yield (R)-β-amino acids with >95% enantiomeric excess (ee). The crystal structures of the holoenzyme and of the enzyme in complex with the inhibitor 2-aminooxyacetate revealed structural similarity to the β-phenylalanine aminotransferase fromMesorhizobiumsp. strain LUK. The crystal structure was used to rationalize the stereo- and regioselectivity ofV. paradoxusaminotransferase and to define a sequence motif with which new aromatic β-amino acid-converting aminotransferases may be identified.
Form and Function of Clostridium thermocellum Biofilms
Authors: Dumitrache Alexandru , Wolfaardt Gideon , Allen Grant , Liss Steven N. , Lynd Lee R. 
Published: October 2012

ABSTRACT

The importance of bacterial adherence has been acknowledged in microbial lignocellulose conversion studies; however, few reports have described the function and structure of biofilms supported by cellulosic substrates. We investigated the organization, dynamic formation, and carbon flow associated with biofilms of the obligately anaerobic cellulolytic bacterium
Clostridium thermocellum
27405. Using noninvasive,
in situ
fluorescence imaging, we showed biofilms capable of near complete substrate conversion with a characteristic monolayered cell structure without an extracellular polymeric matrix typically seen in biofilms. Cell division at the interface and terminal endospores appeared throughout all stages of biofilm growth. Using continuous-flow reactors with a rate of dilution (2 h
−1
) 12-fold higher than the bacterium's maximum growth rate, we compared biofilm activity under low (44 g/liter) and high (202 g/liter) initial cellulose loading. The average hydrolysis rate was over 3-fold higher in the latter case, while the proportions of oligomeric cellulose hydrolysis products lost from the biofilm were 13.7% and 29.1% of the total substrate carbon hydrolyzed, respectively. Fermentative catabolism was comparable between the two cellulose loadings, with ca. 4% of metabolized sugar carbon being utilized for cell production, while 75.4% and 66.7% of the two cellulose loadings, respectively, were converted to primary carbon metabolites (ethanol, acetic acid, lactic acid, carbon dioxide). However, there was a notable difference in the ethanol-to-acetic acid ratio (g/g), measured to be 0.91 for the low cellulose loading and 0.41 for the high cellulose loading. The results suggest that substrate availability for cell attachment rather than biofilm colonization rates govern the efficiency of cellulose conversion.
Development of Quantitative PCR Assays Targeting the 16S rRNA Genes of Enterococcus spp. and Their Application to the Identification of Enterococcus Species in Environmental Samples
Authors: Ryu Hodon , Henson Michael , Elk Michael , Toledo-Hernandez Carlos , Griffith John , Blackwood Denene , Noble Rachel , Gourmelon Michèle , Glassmeyer Susan , Santo Domingo Jorge W. 
Published: October 2012

ABSTRACT

The detection of environmental enterococci has been determined primarily by using culture-based techniques that might exclude some enterococcal species as well as those that are nonculturable. To address this, the relative abundances of enterococci were examined by challenging fecal and water samples against a currently available genus-specific assay (Entero1). To determine the diversity of enterococcal species, 16S rRNA gene-based group-specific quantitative PCR (qPCR) assays were developed and evaluated against eight of the most common environmental enterococcal species. Partial 16S rRNA gene sequences of 439 presumptive environmental enterococcal strains were analyzed to study further the diversity of enterococci and to confirm the specificities of group-specific assays. The group-specific qPCR assays showed relatively high amplification rates with targeted species (>98%), although some assays cross-amplified with nontargeted species (1.3 to 6.5%). The results with the group-specific assays also showed that different enterococcal species co-occurred in most fecal samples. The most abundant enterococci in water and fecal samples were
Enterococcus faecalis
and
Enterococcus faecium
, although we identified more water isolates as
Enterococcus casseliflavus
than as any of the other species. The prevalence of the Entero1 marker was in agreement with the combined number of positive signals determined by the group-specific assays in most fecal samples, except in gull feces. On the other hand, the number of group-specific assay signals was lower in all water samples tested, suggesting that other enterococcal species are present in these samples. While the results highlight the value of genus- and group-specific assays for detecting the major enterococcal groups in environmental water samples, additional studies are needed to determine further the diversity, distributions, and relative abundances of all enterococcal species found in water.
A Pericentrin-Related Protein Homolog in Aspergillus nidulans Plays Important Roles in Nucleus Positioning and Cell Polarity by Affecting Microtubule Organization
Authors: Chen Peiying , Gao Rongsui , Chen Shaochun , Pu Li , Li Pin , Huang Ying , Lu Ling 
Published: October 2012

ABSTRACT

Pericentrin is a large coiled-coil protein in mammalian centrosomes that serves as a multifunctional scaffold for anchoring numerous proteins. Recent studies have linked numerous human disorders with mutated or elevated levels of pericentrin, suggesting unrecognized contributions of pericentrin-related proteins to the development of these disorders. In this study, we characterized AnPcpA, a putative homolog of pericentrin-related protein in the model filamentous fungus
Aspergillus nidulans
, and found that it is essential for conidial germination and hyphal development. Compared to the hyphal apex localization pattern of calmodulin (CaM), which has been identified as an interactive partner of the pericentrin homolog, GFP-AnPcpA fluorescence dots are associated mainly with nuclei, while the accumulation of CaM at the hyphal apex depends on the function of AnPcpA. In addition, the depletion of AnPcpA by an inducible
alcA
promoter repression results in severe growth defects and abnormal nuclear segregation. Most interestingly, in mature hyphal cells, knockdown of pericentrin was able to significantly induce changes in cell shape and cytoskeletal remodeling; it resulted in some enlarged compartments with condensed nuclei and anucleate small compartments as well. Moreover, defects in AnPcpA significantly disrupted the microtubule organization and nucleation, suggesting that AnPcpA may affect nucleus positioning by influencing microtubule organization.
Microsporidian Infection in a Free-Living Marine Nematode
Authors: Ardila-Garcia A. M. , Fast N. M. 
Published: October 2012

ABSTRACTMicrosporidia are unicellular fungi that are obligate endoparasites. Although nematodes are one of the most abundant and diverse animal groups, the only confirmed report of microsporidian infection was that of the “nematode killer” (Nematocida parisii).N. parisiiwas isolated from a wildCaenorhabditissp. and causes an acute and lethal intestinal infection in a lab strain ofCaenorhabditis elegans. We set out to characterize a microsporidian infection in a wild nematode to determine whether the infection pattern ofN. parisiiin the lab is typical of microsporidian infections in nematodes. We describe a novel microsporidian species namedSporanauta perivermis(marine spore of roundworms) and characterize its infection in its natural host, the free-living marine nematodeOdontophora rectangula. S. perivermisis not closely related toN. parisiiand differs strikingly in all aspects of infection. Examination by transmission electron microscopy (TEM) revealed that the infection was localized in the hypodermal and muscle tissues only and did not involve the intestines. Fluorescentin situhybridization (FISH) confirmed infection in the muscle and hypodermis, and surprisingly, it also revealed that the parasite infectsO. rectangulaeggs, suggesting a vertical mode of transmission. Our observations highlight the importance of studying parasites in their natural hosts and indicate that not all nematode-infecting microsporidia are “nematode killers”; instead, microsporidiosis can be more versatile and chronic in the wild.
Inactivation of the Transcriptional Regulator-Encoding Gene
sdiA
Enhances Rice Root Colonization and Biofilm Formation in Enterobacter cloacae GS1
Authors: Shankar Manoharan , Ponraj Paramasivan , Illakkiam Devaraj , Rajendhran Jeyaprakash , Gunasekaran Paramasamy 
Published: October 2012

ABSTRACT

Enterobacter cloacae
GS1 is a plant growth-promoting bacterium which colonizes rice roots. In the rhizosphere environment,
N
-acyl homoserine lactone (NAHL)-like quorum-sensing signals are known to be produced by host plants and other microbial inhabitants.
E. cloacae
GS1 was unable to synthesize NAHL quorum-sensing signals but had the NAHL-dependent transcriptional regulator-encoding gene
sdiA
. This study was aimed at understanding the effects of SdiA and NAHL-dependent cross talk in rice root colonization by
E. cloacae
GS1. Pleiotropic effects of
sdiA
inactivation included substantial increases in root colonization and biofilm formation, suggesting a negative role for SdiA in bacterial adhesion. We provide evidence that
sdiA
inactivation leads to elevated levels of biosynthesis of curli, which is involved in cellular adhesion. Extraneous addition of NAHLs had a negative effect on root colonization and biofilm formation. However, the
sdiA
mutant of
E. cloacae
GS1 was insensitive to NAHLs, suggesting that this NAHL-induced inhibition of root colonization and biofilm formation is SdiA dependent. Therefore, it is proposed that NAHLs produced by both plant and microbes in the rice rhizosphere act as cross-kingdom and interspecies signals to negatively impact cellular adhesion and, thereby, root colonization in
E. cloacae
GS1.
The Sakaguchi Reaction Product Quenches Phycobilisome Fluorescence, Allowing Determination of the Arginine Concentration in Cells of Anabaena Strain PCC 7120
Authors: Ke Shan , Haselkorn Robert 
Published: October 2012

ABSTRACT

The filamentous cyanobacterium
Anabaena
fixes nitrogen in specialized cells called heterocysts. The immediate product of fixation, ammonia, is known to be assimilated by addition to glutamate to make glutamine. How fixed nitrogen is transported along the filament to the 10 to 20 vegetative cells that separate heterocysts is unknown. N-fixing heterocysts accumulate an insoluble polymer containing aspartate and arginine at the cell poles. Lockau's group has proposed that the polymer is degraded at the poles to provide a mobile carrier, arginine, to the vegetative cells (R. Richter, M. Hejazi, R. Kraft, K. Ziegler, and W. Lockau, Eur. J. Biochem.
263:
163–169, 1999). We wished to use the Sakaguchi reaction for arginine to determine the relative cellular concentration of arginine along the filament. At present, the methods for measuring absorption of the Sakaguchi reaction product at 520 nm are insufficiently sensitive for that purpose. However, that product quenches the fluorescence of phycobiliproteins, which we have adapted to a determination of arginine. Our results are consistent with the proposal that arginine is a principal nitrogen carrier from heterocysts to vegetative cells in
Anabaena
.
Identification and Characterization of a Highly Conserved Crenarchaeal Protein Lysine Methyltransferase with Broad Substrate Specificity
Authors: Chu Yindi , Zhang Zhenfeng , Wang Qian , Luo Yuanming , Huang Li 
Published: October 2012

ABSTRACTProtein lysine methylation occurs extensively in theCrenarchaeota, a major kingdom in theArchaea. However, the enzymes responsible for this type of posttranslational modification have not been found. Here we report the identification and characterization of the first crenarchaeal protein lysine methyltransferase, designated aKMT, from the hyperthermophilic crenarchaeonSulfolobus islandicus. The enzyme was capable of transferring methyl groups to selected lysine residues in a substrate protein usingS-adenosyl-l-methionine (SAM) as the methyl donor. aKMT, a non-SET domain protein, is highly conserved among crenarchaea, and distantly related homologs also exist inBacteriaandEukarya. aKMT was active over a wide range of temperatures, from ∼25 to 90°C, with an optimal temperature at ∼60 to 70°C. Amino acid residues Y9 and T12 at the N terminus appear to be the key residues in the putative active site of aKMT, as indicated by sequence conservation and site-directed mutagenesis. Although aKMT was identified based on its methylating activity on Cren7, the crenarchaeal chromatin protein, it exhibited broad substrate specificity and was capable of methylating a number of recombinantSulfolobusproteins overproduced inEscherichia coli. The finding of aKMT will help elucidate mechanisms underlining extensive protein lysine methylation and the functional significance of posttranslational protein methylation in crenarchaea.
AglS, a Novel Component of the Haloferax volcanii N-Glycosylation Pathway, Is a Dolichol Phosphate-Mannose Mannosyltransferase
Authors: Cohen-Rosenzweig Chen , Yurist-Doutsch Sophie , Eichler Jerry 
Published: October 2012

ABSTRACT

In
Haloferax volcanii
, a series of Agl proteins mediates protein N-glycosylation. The genes encoding all but one of the Agl proteins are sequestered into a single gene island. The same region of the genome includes sequences also suspected but not yet verified as serving N-glycosylation roles, such as
HVO
_
1526
. In the following, HVO_1526, renamed AglS, is shown to be necessary for the addition of the final mannose subunit of the pentasaccharide N-linked to the surface (S)-layer glycoprotein, a convenient reporter of N-glycosylation in
Hfx. volcanii
. Relying on bioinformatics, topological analysis, gene deletion, mass spectrometry, and biochemical assays, AglS was shown to act as a dolichol phosphate-mannose mannosyltransferase, mediating the transfer of mannose from dolichol phosphate to the tetrasaccharide corresponding to the first four subunits of the pentasaccharide N-linked to the S-layer glycoprotein.
Transcription Factor Family-Based Reconstruction of Singleton Regulons and Study of the Crp/Fnr, ArsR, and GntR Families in Desulfovibrionales Genomes
Authors: Kazakov Alexey E. , Rodionov Dmitry A. , Price Morgan N. , Arkin Adam P. , Dubchak Inna , Novichkov Pavel S. 
Published: October 2012

ABSTRACT

Accurate detection of transcriptional regulatory elements is essential for high-quality genome annotation, metabolic reconstruction, and modeling of regulatory networks. We developed a computational approach for reconstruction of regulons operated by transcription factors (TFs) from large protein families and applied this novel approach to three TF families in 10
Desulfovibrionales
genomes. Phylogenetic analyses of 125 regulators from the ArsR, Crp/Fnr, and GntR families revealed that 65% of these regulators (termed reference TFs) are well conserved in
Desulfovibrionales
, while the remaining 35% of regulators (termed singleton TFs) are species specific and show a mosaic distribution. For regulon reconstruction in the group of singleton TFs, the standard orthology-based approach was inefficient, and thus, we developed a novel approach based on the simultaneous study of all homologous TFs from the same family in a group of genomes. As a result, we identified binding for 21 singleton TFs and for all reference TFs in all three analyzed families. Within each TF family we observed structural similarities between DNA-binding motifs of different reference and singleton TFs. The collection of reconstructed regulons is available at the RegPrecise database (
http://regprecise.lbl.gov/RegPrecise/Desulfovibrionales.jsp
).
HIV among people who inject drugs in Central and Eastern Europe and Central Asia: a systematic review with implications for policy
Authors: Jolley Emma , Rhodes Tim , Platt Lucy , Hope Vivian , Latypov Alisher , Donoghoe Martin , Wilson David 
Published: October 2012

Background and objectivesHIV among people who inject drugs (PWID) is a major public health concern in Eastern and Central Europe and Central Asia. HIV transmission in this group is growing and over 27 000 HIV cases were diagnosed among PWID in 2010 alone. The objective of this systematic review was to examine risk factors associated with HIV prevalence among PWID in Central and Eastern Europe and Central Asia and to describe the response to HIV in this population and the policy environments in which they live.DesignA systematic review of peer-reviewed and grey literature addressing HIV prevalence and risk factors for HIV prevalence among PWID and a synthesis of key resources describing the response to HIV in this population. We used a comprehensive search strategy across multiple electronic databases to collect original research papers addressing HIV prevalence and risk factors among PWID since 2005. We summarised the extent of key harm reduction interventions, and using a simple index of ‘enabling’ environment described the policy environments in which they are implemented.Studies reviewedOf the 5644 research papers identified from electronic databases and 40 documents collected from our grey literature search, 70 documents provided unique estimates of HIV and 14 provided multivariate risk factors for HIV among PWID.ResultsHIV prevalence varies widely, with generally low or medium (<5%) prevalence in Central Europe and high (>10%) prevalence in Eastern Europe. We found evidence for a number of structural factors associated with HIV including gender, socio-economic position and contact with law enforcement agencies.ConclusionsThe HIV epidemic among PWID in the region is varied, with the greatest burden generally in Eastern Europe. Data suggest that the current response to HIV among PWID is insufficient, and hindered by multiple environmental barriers including restricted access to services and unsupportive policy or social environments.
Rheumatic heart disease: pilot study for a population-based evaluation of prevalence and cardiovascular outcomes among schoolchildren in Nepal
Authors: Shrestha Nikesh Raj , Kalesan Bindu , Karki Prahlad , Sherpa Kunjang , Basnet Anil , Urban Philip , Pilgrim Thomas 
Published: October 2012

ObjectivesTo evaluate a protocol for a population-based programme targeting the prevention of rheumatic heart disease (RHD) progression by early echocardiographic diagnosis of valvular lesions and timely implementation of secondary prevention.DesignObservational survey with a subsequent prospective cohort study.SettingPrivate boarding school in the urban area of the Sunsari district situated on the foothills of the Lower Himalayan Range in Eastern Nepal.ParticipantsFifty-four unselected school-going children 5–15 years of age, 24 girls and 30 boys.Primary outcome measureLogistic feasibility of a large-scale population-based screening study using the echocardiographic criteria formulated by the World Heart Federation, with longitudinal follow-up of children with definite or borderline RHD in a prospective cohort study.ResultsStandardised interview, physical examination and screening echocardiography were performed in a three-staged process and took approximately 6 min per child. Socio-economic status was assessed using surrogate markers such as the occupation of the primary caregiver, numbers of rooms at home, car, television, cell phone and internet connection. Physical examination was focused on cardiac auscultation and signs of acute rheumatic fever and targeted echocardiography was performed by an independent examiner without knowledge of the clinical findings. Two children with evidence of borderline RHD were re-examined at B.P. Koirala Institute of Health Sciences and the indication for secondary antibiotic prevention was discussed with the parents and the children. At 6 months of follow-up, echocardiographic findings were stable in both children. Implementation of secondary antibiotic prevention was challenged by impaired awareness of subclinical RHD among parents and inadequate cooperation with family physicians.ConclusionsThis pilot study shows that the methods outlined in the protocol can be translated into a large-scale population-based study. We learned that education and collaboration with teachers, parents and family physicians/paediatricians will be of key importance in order to establish a sustainable programme.