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purification, crystallization and preliminary crystallographic analysis of katb, a manganese catalase from anabaena pcc 7120.catalases are enzymes that play an important role in the detoxification of hydrogen peroxide (h2o2) in aerobic organisms. among catalases, haem-containing catalases are ubiquitously distributed and their enzymatic mechanism is very well understood. on the other hand, manganese catalases that contain a bimanganese core in the active site have been less well characterized and their mode of action is not fully understood. the genome of anabaena pcc 7120 does not show the presence of a haem catalase ...201324192374
cloning, expression, purification, crystallization and preliminary crystallographic analysis of the putative nlpc/p60 endopeptidase, ttha0266, from thermus thermophilus hb8.autolysins belong to a protein family involved in peptidoglycan degradation and remodelling. within this family, nlpc/p60 endopeptidases are involved in the hydrolysis of the peptide arm of peptidoglycan. in this work, the putative nlpc/p60 endopeptidase ttha0266 from thermus thermophilus hb8 was overexpressed, purified and crystallized. the crystals diffracted to 2.4 å resolution and belonged to the hexagonal space group p6(1), with unit-cell parameters a = b = 71.19, c = 198.68 å, γ = 120°. se ...201324192372
cloning, overexpression, purification and crystallization of malate dehydrogenase from thermus thermophilus.malate dehydrogenase (mdh) has been used as a conjugate for enzyme immunoassay of a wide variety of compounds, such as drugs of abuse, drugs used in repetitive therapeutic application and hormones. in consideration of the various biotechnological applications of mdh, investigations of mdh from thermus thermophilus were carried out to further understand the properties of this enzyme. the dna fragment containing the open reading frame of mdh was amplified from the genomic dna of t. thermophilus an ...201324192361
biophysical characterization of a recombinant aminopeptidase ii from the thermophilic bacterium bacillus stearothermophilus.in the present study, the biophysical properties of his6-tagged bacillus stearothermophilus aminopeptidase ii (his6-tagged bsampii) are characterized in detail by gel-filtration, analytical ultracentrifugation, and various spectroscopic techniques. using size-exclusion chromatography and analytical ultracentrifugation, we demonstrate that his6-tagged bsampii exists predominantly as a dimer in solution. the enzyme is active and stable at phs ranging from 6.5 to 8.5. far-uv circular dichroism anal ...201324165863
biophysical characterization of a recombinant aminopeptidase ii from the thermophilic bacterium bacillus stearothermophilus.in the present study, the biophysical properties of his6-tagged bacillus stearothermophilus aminopeptidase ii (his6-tagged bsampii) are characterized in detail by gel-filtration, analytical ultracentrifugation, and various spectroscopic techniques. using size-exclusion chromatography and analytical ultracentrifugation, we demonstrate that his6-tagged bsampii exists predominantly as a dimer in solution. the enzyme is active and stable at phs ranging from 6.5 to 8.5. far-uv circular dichroism anal ...201324165863
metagenomic approach for the isolation of a thermostable β-galactosidase with high tolerance of galactose and glucose from soil samples of turpan basin.β-galactosidases can be used to produce low-lactose milk and dairy products for lactose intolerant people. although commercial β-galactosidases have outstanding lactose hydrolysis ability, their thermostability is low, and reaction products have strong inhibition to these enzymes. in addition, the β-galactosidases possessing simultaneously high thermostability and tolerance of galactose and glucose are still seldom reported until now. therefore, identification of novel β-galactosidases with high ...201324156692
translational regulation of yersinia enterocolitica mrna encoding a type iii secretion substrate.yersinia enterocolitica type iii secretion machines transport yopq and other yop effectors into host immune cells. yopd and its chaperone lcrh are essential components of the yersinia type iii pathway, enabling effector translocation into host cells. yopd, lcrh, and yscm1 also regulate yop expression post-transcriptionally in response to environmental signals; however, the molecular mechanisms for this regulation and yop secretion are unknown. we show here that yopd associates with 30 s ribosoma ...201324158443
a comparative study of ribosomal proteins: linkage between amino acid distribution and ribosomal assembly.assembly of the ribosome from its protein and rna constituents must occur quickly and efficiently in order to synthesize the proteins necessary for all cellular activity. since the early 1960's, certain characteristics of possible assembly pathways have been elucidated, yet the mechanisms that govern the precise recognition events remain unclear.we utilize a comparative analysis to investigate the amino acid composition of ribosomal proteins (r-proteins) with respect to their role in the assembl ...201324152303
structure of the secy channel during initiation of protein translocation.many secretory proteins are targeted by signal sequences to a protein-conducting channel, formed by prokaryotic secy or eukaryotic sec61 complexes, and are translocated across the membrane during their synthesis. crystal structures of the inactive channel show that the secy subunit of the heterotrimeric complex consists of two halves that form an hourglass-shaped pore with a constriction in the middle of the membrane and a lateral gate that faces the lipid phase. the closed channel has an empty ...201324153188
structure of the secy channel during initiation of protein translocation.many secretory proteins are targeted by signal sequences to a protein-conducting channel, formed by prokaryotic secy or eukaryotic sec61 complexes, and are translocated across the membrane during their synthesis. crystal structures of the inactive channel show that the secy subunit of the heterotrimeric complex consists of two halves that form an hourglass-shaped pore with a constriction in the middle of the membrane and a lateral gate that faces the lipid phase. the closed channel has an empty ...201324153188
crystal structure of a bioactive pactamycin analog bound to the 30s ribosomal subunit.biosynthetically and chemically derived analogs of the antibiotic pactamycin and de-6-methylsalicylyl (msa)-pactamycin have attracted recent interest as potential antiprotozoal and antitumor drugs. here, we report a 3.1-å crystal structure of de-6-msa-pactamycin bound to its target site on the thermus thermophilus 30s ribosomal subunit. although de-6-msa-pactamycin lacks the msa moiety, it shares the same binding site as pactamycin and induces a displacement of nucleic acid template bound at the ...201323702293
evolution of crispr rna recognition and processing by cas6 endonucleases.in many bacteria and archaea, small rnas derived from clustered regularly interspaced short palindromic repeats (crisprs) associate with crispr-associated (cas) proteins to target foreign dna for destruction. in type i and iii crispr/cas systems, the cas6 family of endoribonucleases generates functional crispr-derived rnas by site-specific cleavage of repeat sequences in precursor transcripts. crispr repeats differ widely in both sequence and structure, with varying propensity to form hairpin fo ...201324150936
evolution of crispr rna recognition and processing by cas6 endonucleases.in many bacteria and archaea, small rnas derived from clustered regularly interspaced short palindromic repeats (crisprs) associate with crispr-associated (cas) proteins to target foreign dna for destruction. in type i and iii crispr/cas systems, the cas6 family of endoribonucleases generates functional crispr-derived rnas by site-specific cleavage of repeat sequences in precursor transcripts. crispr repeats differ widely in both sequence and structure, with varying propensity to form hairpin fo ...201324150936
12-fold symmetry of the putative portal protein from the thermus thermophilus bacteriophage g20c determined by x-ray analysis.in tailed bacteriophages and several animal viruses, the portal protein forms the gateway through which viral dna is translocated into the head structure during viral particle assembly. in the mature virion the portal protein exists as a dodecamer, while recombinant portal proteins from several phages, including spp1 and cnph82, have been shown to form 13-subunit assemblies. a putative portal protein from the thermostable bacteriophage g20c has been cloned, overexpressed and purified. crystals o ...201324192358
crystallization and preliminary x-ray study of alanine dehydrogenase from bacillus pseudofirmus of4.alanine dehydrogenase (of4ald) from the alkaliphilic bacillus pseudofirmus of4 was expressed and purified with a his6 tag in a form suitable for x-ray crystallographic analysis. crystals were grown by the hanging-drop vapour-diffusion method at 289 k using a solution consisting of 0.1 m tris-hcl ph 8.0, 0.2 m liso4, 22%(w/v) peg 3350. x-ray diffraction data were collected to 2.8 å resolution. the crystal belonged to the triclinic space group p1, with unit-cell parameters a = 88.04, b = 105.59, c ...201324192355
crystallization and preliminary x-ray diffraction studies of staphylococcus aureus homoserine dehydrogenase.staphylococcus aureus is a gram-positive nosocomial pathogen. the prevalence of multidrug-resistant s. aureus strains in both hospital and community settings makes it imperative to characterize new drug targets to combat s. aureus infections. in this context, enzymes involved in cell-wall maintenance and essential amino-acid biosynthesis are significant drug targets. homoserine dehydrogenase (hsd) is an oxidoreductase that is involved in the reversible conversion of l-aspartate semialdehyde to l ...201324192352
temperature dependence of the dna double helix at the nanoscale: structure, elasticity, and fluctuations.biological organisms exist over a broad temperature range of -15°c to +120°c, where many molecular processes involving dna depend on the nanoscale properties of the double helix. here, we present results of extensive molecular dynamics simulations of dna oligomers at different temperatures. we show that internal basepair conformations are strongly temperature-dependent, particularly in the stretch and opening degrees of freedom whose harmonic fluctuations can be considered the initial steps of t ...201324138866
structure of the homodimeric glycine decarboxylase p-protein from synechocystis sp. pcc 6803 suggests a mechanism for redox regulation.glycine decarboxylase, or p-protein, is a pyridoxal 5'-phosphate (plp)-dependent enzyme in one-carbon metabolism of all organisms, in the glycine and serine catabolism of vertebrates, and in the photorespiratory pathway of oxygenic phototrophs. p-protein from the cyanobacterium synechocystis sp. pcc 6803 is an α2 homodimer with high homology to eukaryotic p-proteins. the crystal structure of the apoenzyme shows the c terminus locked in a closed conformation by a disulfide bond between cys(972) i ...201324121504
structure and activity of the rna-targeting type iii-b crispr-cas complex of thermus thermophilus.the crispr-cas system is a prokaryotic host defense system against genetic elements. the type iii-b crispr-cas system of the bacterium thermus thermophilus, the ttcmr complex, is composed of six different protein subunits (cmr1-6) and one crrna with a stoichiometry of cmr112131445361:crrna1. the ttcmr complex copurifies with crrna species of 40 and 46 nt, originating from a distinct subset of crispr loci and spacers. the ttcmr complex cleaves the target rna at multiple sites with 6 nt intervals ...201324119403
cloning and expression of dnak gene from bacillus pumilus of hot water spring origin.a set of thermotolerant strains isolated from hot springs of manikaran and bakreshwar (india) were selected with an aim to isolate dnak gene which encodes dnak protein. the gene dnak along with its flanking region was successfully amplified from 5 different strains (4 from bakreshwar and one from manikaran). restriction fragment length polymorphism (rflp) revealed that amplicons were almost identical in sequence. the dnak gene from one representative, bacillus pumilus strain b3 isolated from bak ...201327275408
cloning and expression of dnak gene from bacillus pumilus of hot water spring origin.a set of thermotolerant strains isolated from hot springs of manikaran and bakreshwar (india) were selected with an aim to isolate dnak gene which encodes dnak protein. the gene dnak along with its flanking region was successfully amplified from 5 different strains (4 from bakreshwar and one from manikaran). restriction fragment length polymorphism (rflp) revealed that amplicons were almost identical in sequence. the dnak gene from one representative, bacillus pumilus strain b3 isolated from bak ...201327275408
improvement of biocatalysts for industrial and environmental purposes by saturation mutagenesis.laboratory evolution techniques are becoming increasingly widespread among protein engineers for the development of novel and designed biocatalysts. the palette of different approaches ranges from complete randomized strategies to rational and structure-guided mutagenesis, with a wide variety of costs, impacts, drawbacks and relevance to biotechnology. a technique that convincingly compromises the extremes of fully randomized vs. rational mutagenesis, with a high benefit/cost ratio, is saturatio ...201324970191
staphylococcus epidermidis csm1 is a 3'-5' exonuclease.clustered regularly interspaced short palindromic repeats (crispr) offer an adaptive immune system that protects bacteria and archaea from nucleic acid invaders through an rna-mediated nucleic acid cleavage mechanism. our knowledge of nucleic acid cleavage mechanisms is limited to three examples of widely different ribonucleoprotein particles that target either dna or rna. staphylococcus epidermidis belongs to the type iii-a crispr system and has been shown to interfere with invading dna in vivo ...201324121684
staphylococcus epidermidis csm1 is a 3'-5' exonuclease.clustered regularly interspaced short palindromic repeats (crispr) offer an adaptive immune system that protects bacteria and archaea from nucleic acid invaders through an rna-mediated nucleic acid cleavage mechanism. our knowledge of nucleic acid cleavage mechanisms is limited to three examples of widely different ribonucleoprotein particles that target either dna or rna. staphylococcus epidermidis belongs to the type iii-a crispr system and has been shown to interfere with invading dna in vivo ...201324121684
conformational and thermodynamic hallmarks of dna operator site specificity in the copper sensitive operon repressor from streptomyces lividans.metal ion homeostasis in bacteria relies on metalloregulatory proteins to upregulate metal resistance genes and enable the organism to preclude metal toxicity. the copper sensitive operon repressor (csor) family is widely distributed in bacteria and controls the expression of copper efflux systems. csor operator sites consist of g-tract containing pseudopalindromes of which the mechanism of operator binding is poorly understood. here, we use a structurally characterized csor from streptomyces li ...201324121681
conformational and thermodynamic hallmarks of dna operator site specificity in the copper sensitive operon repressor from streptomyces lividans.metal ion homeostasis in bacteria relies on metalloregulatory proteins to upregulate metal resistance genes and enable the organism to preclude metal toxicity. the copper sensitive operon repressor (csor) family is widely distributed in bacteria and controls the expression of copper efflux systems. csor operator sites consist of g-tract containing pseudopalindromes of which the mechanism of operator binding is poorly understood. here, we use a structurally characterized csor from streptomyces li ...201324121681
menaquinone biosynthesis: formation of aminofutalosine requires a unique radical sam enzyme.menaquinone (mk, vitamin k2) is a lipid-soluble molecule that participates in the bacterial electron transport chain. in mammalian cells, mk functions as an essential vitamin for the activation of various proteins involved in blood clotting and bone metabolism. recently, a new pathway for the biosynthesis of this cofactor was discovered in streptomyces coelicolor a3(2) in which chorismate is converted to aminofutalosine in a reaction catalyzed by mqna and an unidentified enzyme. here, we reconst ...201324083939
construction of an in vitro bypassed pyruvate decarboxylation pathway using thermostable enzyme modules and its application to n-acetylglutamate production.metabolic engineering has emerged as a practical alternative to conventional chemical conversion particularly in biocommodity production processes. however, this approach is often hampered by as yet unidentified inherent mechanisms of natural metabolism. one of the possible solutions for the elimination of the negative effects of natural regulatory mechanisms on artificially engineered metabolic pathway is to construct an in vitro pathway using a limited number of enzymes. employment of thermost ...201324099461
minimal mechanistic model of sirna-dependent target rna slicing by recombinant human argonaute 2 protein.argonaute (ago) proteins are the key component of the rna-induced silencing complex and mediate rna interference (rnai) in association with small rnas. although overall the mechanism of rnai is well understood, many molecular details of this complex process are not. here we report about in-depth steady-state and, in particular, pre-steady-state characterization of sirna binding, target rna recognition, sequence-specific cleavage and product release by recombinant human ago 2 (hago2). in combinin ...201324101500
archaeal ubiquitin-like samp3 is isopeptide-linked to proteins via a ubaa-dependent mechanism.samp1 and samp2 are ubiquitin-like proteins that function as protein modifiers and are required for the production of sulfur-containing biomolecules in the archaeon haloferax volcanii. here we report a novel small archaeal modifier protein (named samp3) with a β-grasp fold and c-terminal diglycine motif characteristic of ubiquitin that is functional in protein conjugation in hfx. volcanii. samp3 conjugates were dependent on the ubiquitin-activating e1 enzyme homolog of archaea (ubaa) for synthes ...201324097257
archaeal ubiquitin-like samp3 is isopeptide-linked to proteins via a ubaa-dependent mechanism.samp1 and samp2 are ubiquitin-like proteins that function as protein modifiers and are required for the production of sulfur-containing biomolecules in the archaeon haloferax volcanii. here we report a novel small archaeal modifier protein (named samp3) with a β-grasp fold and c-terminal diglycine motif characteristic of ubiquitin that is functional in protein conjugation in hfx. volcanii. samp3 conjugates were dependent on the ubiquitin-activating e1 enzyme homolog of archaea (ubaa) for synthes ...201324097257
crystal structure of human seryl-trna synthetase and ser-sa complex reveals a molecular lever specific to higher eukaryotes.seryl-trna synthetase (serrs), an essential enzyme for translation, also regulates vascular development. this "gain-of-function" has been linked to the une-s domain added to vertebrate serrs during evolution. however, the significance of two insertions also specific to higher eukaryotic serrs remains elusive. here, we determined the crystal structure of human serrs in complex with ser-sa, an aminoacylation reaction intermediate analog, at 2.9 å resolution. despite a 70 å distance, binding of ser ...201324095058
thermal adaptation of conformational dynamics in ribonuclease h.the relationship between inherent internal conformational processes and enzymatic activity or thermodynamic stability of proteins has proven difficult to characterize. the study of homologous proteins with differing thermostabilities offers an especially useful approach for understanding the functional aspects of conformational dynamics. in particular, ribonuclease hi (rnase h), an 18 kd globular protein that hydrolyzes the rna strand of rna:dna hybrid substrates, has been extensively studied by ...201324098095
the role of the mitochondrial ribosome in human disease: searching for mutations in 12s mitochondrial rrna with high disruptive potential.mutations of mitochondrial dna are linked to many human diseases. despite the identification of a large number of variants in the mitochondrially encoded rrna (mt-rrna) genes, the evidence supporting their pathogenicity is, at best, circumstantial. establishing the pathogenicity of these variations is of major diagnostic importance. here, we aim to estimate the disruptive effect of mt-rrna variations on the function of the mitochondrial ribosome. in the absence of direct biochemical methods to s ...201324092330
the role of the mitochondrial ribosome in human disease: searching for mutations in 12s mitochondrial rrna with high disruptive potential.mutations of mitochondrial dna are linked to many human diseases. despite the identification of a large number of variants in the mitochondrially encoded rrna (mt-rrna) genes, the evidence supporting their pathogenicity is, at best, circumstantial. establishing the pathogenicity of these variations is of major diagnostic importance. here, we aim to estimate the disruptive effect of mt-rrna variations on the function of the mitochondrial ribosome. in the absence of direct biochemical methods to s ...201324092330
ndufaf7 methylates arginine 85 in the ndufs2 subunit of human complex i.complex i (nadh ubiquinone oxidoreductase) in mammalian mitochondria is an l-shaped assembly of 44 subunits. one arm is embedded in the inner membrane with the other protruding ∼100 å into the matrix of the organelle. the extrinsic arm contains binding sites for nadh and the primary electron acceptor fmn, and it provides a scaffold for seven iron-sulfur clusters that form an electron pathway linking fmn to the terminal electron acceptor, ubiquinone, which is bound in the region of the junction b ...201324089531
basic properties of rotary dynamics of the molecular motor enterococcus hirae v1-atpase.v-atpases are rotary molecular motors that generally function as proton pumps. we recently solved the crystal structures of the v1 moiety of enterococcus hirae v-atpase (ehv1) and proposed a model for its rotation mechanism. here, we characterized the rotary dynamics of ehv1 using single-molecule analysis employing a load-free probe. ehv1 rotated in a counterclockwise direction, exhibiting two distinct rotational states, namely clear and unclear, suggesting unstable interactions between the roto ...201324089518
protein folding on the ribosome studied using nmr spectroscopy.nmr spectroscopy is a powerful tool for the investigation of protein folding and misfolding, providing a characterization of molecular structure, dynamics and exchange processes, across a very wide range of timescales and with near atomic resolution. in recent years nmr methods have also been developed to study protein folding as it might occur within the cell, in a de novo manner, by observing the folding of nascent polypeptides in the process of emerging from the ribosome during synthesis. des ...201324083462
nanometer scale pores similar in size to the entrance of the ribosomal exit cavity are a common feature of large rnas.the highly conserved peptidyl transferase center (ptc) of the ribosome contains an rna pore that serves as the entrance to the exit tunnel. analysis of available ribosome crystal structures has revealed the presence of multiple additional well-defined pores of comparable size in the ribosomal (rrna) rnas. these typically have dimensions of 1-2 nm, with a total area of ∼100 å(2) or more, and most are associated with one or more ribosomal proteins. the ptc example and the other rrna pores result f ...201323940386
s6:s18 ribosomal protein complex interacts with a structural motif present in its own mrna.prokaryotic ribosomal protein genes are typically grouped within highly conserved operons. in many cases, one or more of the encoded proteins not only bind to a specific site in the ribosomal rna, but also to a motif localized within their own mrna, and thereby regulate expression of the operon. in this study, we computationally predicted an rna motif present in many bacterial phyla within the 5' untranslated region of operons encoding ribosomal proteins s6 and s18. we demonstrated that the s6:s ...201323980204
automated classification of rna 3d motifs and the rna 3d motif atlas.the analysis of atomic-resolution rna three-dimensional (3d) structures reveals that many internal and hairpin loops are modular, recurrent, and structured by conserved non-watson-crick base pairs. structurally similar loops define rna 3d motifs that are conserved in homologous rna molecules, but can also occur at nonhomologous sites in diverse rnas, and which often vary in sequence. to further our understanding of rna motif structure and sequence variability and to provide a useful resource for ...201323970545
mechanisms of eukaryotic transcription.a report on the cold spring harbor laboratory mechanisms of eukaryotic transcription meeting, cold spring harbor, new york, usa, august 27–31, 2013.201324079829
fast fitting to low resolution density maps: elucidating large-scale motions of the ribosome.determining the conformational rearrangements of large macromolecules is challenging experimentally and computationally. case in point is the ribosome; it has been observed by high-resolution crystallography in several states, but many others are known only from low-resolution methods including cryo-electron microscopy. combining these data into dynamical trajectories that may aid understanding of its largest-scale conformational changes has so far remained out of reach of computational methods. ...201324081579
fast fitting to low resolution density maps: elucidating large-scale motions of the ribosome.determining the conformational rearrangements of large macromolecules is challenging experimentally and computationally. case in point is the ribosome; it has been observed by high-resolution crystallography in several states, but many others are known only from low-resolution methods including cryo-electron microscopy. combining these data into dynamical trajectories that may aid understanding of its largest-scale conformational changes has so far remained out of reach of computational methods. ...201324081579
crystallization and preliminary crystallographic analysis of ganb, a gh42 intracellular β-galactosidase from geobacillus stearothermophilus.geobacillus stearothermophilus t-6 is a gram-positive thermophilic soil bacterium that contains a multi-enzyme system for the utilization of plant cell-wall polysaccharides, including xylan, arabinan and galactan. the bacterium uses a number of endo-acting extracellular enzymes that break down the high-molecular-weight polysaccharides into decorated oligosaccharides. these oligosaccharides enter the cell and are further hydrolyzed into sugar monomers by a set of intracellular glycoside hydrolase ...201324100561
a nondiscriminating glutamyl-trna synthetase in the plasmodium apicoplast: the first enzyme in an indirect aminoacylation pathway.the malaria parasite plasmodium falciparum and related organisms possess a relict plastid known as the apicoplast. apicoplast protein synthesis is a validated drug target in malaria because antibiotics that inhibit translation in prokaryotes also inhibit apicoplast protein synthesis and are sometimes used for malaria prophylaxis or treatment. we identified components of an indirect aminoacylation pathway for gln-trna(gln) biosynthesis in plasmodium that we hypothesized would be essential for api ...201324072705
comparative analysis of barophily-related amino acid content in protein domains of pyrococcus abyssi and pyrococcus furiosus.amino acid substitution patterns between the nonbarophilic pyrococcus furiosus and its barophilic relative p. abyssi confirm that hydrostatic pressure asymmetry indices reflect the extent to which amino acids are preferred by barophilic archaeal organisms. substitution patterns in entire protein sequences, shared protein domains defined at fold superfamily level, domains in homologous sequence pairs, and domains of very ancient and very recent origin now provide further clues about the environme ...201324187517
single mutations that redirect internal proton transfer in the ba3 oxidase from thermus thermophilus.the ba3-type cytochrome c oxidase from thermus thermophilus is a membrane-bound proton pump. results from earlier studies have shown that with the aa3-type oxidases proton uptake to the catalytic site and "pump site" occurs simultaneously. however, with ba3 oxidase the pump site is loaded before proton transfer to the catalytic site because the proton transfer to the latter is slower than that with the aa3 oxidases. in addition, the timing of formation and decay of catalytic intermediates is dif ...201324004023
structural, biochemical and genetic characterization of dissimilatory atp sulfurylase from allochromatium vinosum.atp sulfurylase (atps) catalyzes a key reaction in the global sulfur cycle by reversibly converting inorganic sulfate (so4 (2-)) with atp to adenosine 5'-phosphosulfate (aps) and pyrophosphate (ppi). in this work we report on the sat encoded dissimilatory atp sulfurylase from the sulfur-oxidizing purple sulfur bacterium allochromatium vinosum. in this organism, the sat gene is located in one operon and co-transcribed with the aprmba genes for membrane-bound aps reductase. like aps reductase, sat ...201324073218
mutations in gtp binding protein obg of mycoplasma synoviae vaccine strain ms-h: implications in temperature-sensitivity phenotype.mycoplasma synoviae strain ms-h, developed by chemical mutagenesis of the australian field strain 86079/7ns, is a live temperature-sensitive (ts (+)) vaccine used for control of m. synoviae infection in poultry worldwide. genetic basis of temperature sensitivity and attenuation of ms-h has not been revealed thus far. comparison of the complete genome sequence of ms-h, its parent strain 86079/7ns and two non-temperature sensitive (ts (-)) reisolates of ms-h revealed a mutation in a highly conserv ...201324069254
loose binding of the df axis with the a3b3 complex stimulates the initial activity of enterococcus hirae v1-atpase.vacuolar atpases (v-atpases) function as proton pumps in various cellular membrane systems. the hydrophilic v1 portion of the v-atpase is a rotary motor, in which a central-axis df complex rotates inside a hexagonally arranged catalytic a3b3 complex by using atp hydrolysis energy. we have previously reported crystal structures of enterococcushirae v-atpase a3b3 and a3b3df (v1) complexes; the result suggested that the df axis induces structural changes in the a3b3 complex through extensive protei ...201324058539
an ftsh protease is recruited to the mitochondrion of plasmodium falciparum.the two organelles, apicoplast and mitochondrion, of the malaria parasite plasmodium falciparum have unique morphology in liver and blood stages; they undergo complex branching and looping prior to division and segregation into daughter merozoites. little is known about the molecular processes and proteins involved in organelle biogenesis in the parasite. we report the identification of an aaa+/ftsh protease homolog (pfftsh1) that exhibits atp- and zn(2+)-dependent protease activity. pfftsh1 und ...201324058559
chaperone machines for protein folding, unfolding and disaggregation.molecular chaperones are diverse families of multidomain proteins that have evolved to assist nascent proteins to reach their native fold, protect subunits from heat shock during the assembly of complexes, prevent protein aggregation or mediate targeted unfolding and disassembly. their increased expression in response to stress is a key factor in the health of the cell and longevity of an organism. unlike enzymes with their precise and finely tuned active sites, chaperones are heavy-duty molecul ...201324026055
bacterial argonaute samples the transcriptome to identify foreign dna.eukaryotic argonautes bind small rnas and use them as guides to find complementary rna targets and induce gene silencing. though homologs of eukaryotic argonautes are present in many bacteria and archaea, their small rna partners and functions are unknown. we found that the argonaute of rhodobacter sphaeroides (rsago) associates with 15-19 nt rnas that correspond to the majority of transcripts. rsago also binds single-stranded 22-24 nt dna molecules that are complementary to the small rnas and e ...201324034694
involvement of protein if2 n domain in ribosomal subunit joining revealed from architecture and function of the full-length initiation factor.translation initiation factor 2 (if2) promotes 30s initiation complex (ic) formation and 50s subunit joining, which produces the 70s ic. the architecture of full-length if2, determined by small angle x-ray diffraction and cryo electron microscopy, reveals a more extended conformation of if2 in solution and on the ribosome than in the crystal. the n-terminal domain is only partially visible in the 30s ic, but in the 70s ic, it stabilizes interactions between if2 and the l7/l12 stalk of the 50s, a ...201324029017
initiation factor 2 crystal structure reveals a different domain organization from eukaryotic initiation factor 5b and mechanism among translational gtpases.the initiation of protein synthesis uses initiation factor 2 (if2) in prokaryotes and a related protein named eukaryotic initiation factor 5b (eif5b) in eukaryotes. if2 is a gtpase that positions the initiator trna on the 30s ribosomal initiation complex and stimulates its assembly to the 50s ribosomal subunit to make the 70s ribosome. the 3.1-å resolution x-ray crystal structures of the full-length thermus thermophilus apo if2 and its complex with gdp presented here exhibit two different confor ...201324029018
systematic mutational analysis of the putative hydrolase pqse: toward a deeper molecular understanding of virulence acquisition in pseudomonas aeruginosa.pseudomonas aeruginosa is an important opportunistic human pathogen that can establish bacterial communication by synchronizing the behavior of individual cells in a molecular phenomenon known as "quorum sensing". through an elusive mechanism involving gene products of the pqs operon, the pqse enzyme is absolutely required for the synthesis of extracellular phenazines, including the toxic blue pigment pyocyanin, effectively allowing cells to achieve full-fledged virulence. despite several functi ...201324040042
complete genomic dna sequence of the east asian spotted fever disease agent rickettsia japonica.rickettsia japonica is an obligate intracellular alphaproteobacteria that causes tick-borne japanese spotted fever, which has spread throughout east asia. we determined the complete genomic dna sequence of r. japonica type strain yh (vr-1363), which consists of 1,283,087 base pairs (bp) and 971 protein-coding genes. comparison of the genomic dna sequence of r. japonica with other rickettsiae in the public databases showed that 2 regions (4,323 and 216 bp) were conserved in a very narrow range of ...201324039725
aminoacylation of trna 2'- or 3'-hydroxyl by phosphoseryl- and pyrrolysyl-trna synthetases.class i and ii aminoacyl-trna synthetases (aarss) attach amino acids to the 2'- and 3'-oh of the trna terminal adenosine, respectively. one exception is phenylalanyl-trna synthetase (phers), which belongs to class ii but attaches phenylalanine to the 2'-oh. here we show that two class ii aarss, o-phosphoseryl- (seprs) and pyrrolysyl-trna (pylrs) synthetases, aminoacylate the 2'- and 3'-oh, respectively. structure-based-phylogenetic analysis reveals that seprs is more closely related to phers tha ...201324021645
significance of neuropilin-1 expression in acute myeloid leukemia.neuropilin-1 is a vascular endothelial growth factor receptor that acts as a mediator of angiogenesis. its importance in hematological malignancies such as acute myeloid leukemia (aml) remains to be elucidated. the aim of this study was to evaluate the significance of neuropilin-1 expression in aml patients by both flow cytometry and real-time polymerase chain reaction (pcr) in regard to its diagnostic and prognostic values.201324385810
the uve1 endonuclease is regulated by the white collar complex to protect cryptococcus neoformans from uv damage.the pathogenic fungus cryptococcus neoformans uses the bwc1-bwc2 photoreceptor complex to regulate mating in response to light, virulence and ultraviolet radiation tolerance. how the complex controls these functions is unclear. here, we identify and characterize a gene in cryptococcus, uve1, whose mutation leads to a uv hypersensitive phenotype. the homologous gene in fission yeast schizosaccharomyces pombe encodes an apurinic/apyrimidinic endonuclease acting in the uvde-dependent excision repai ...201324039606
splicefinder - a fast and easy screening method for active protein trans-splicing positions.split intein enabled protein trans-splicing (pts) is a powerful method for the ligation of two protein fragments, thereby paving the way for various protein modification or protein function control applications. pts activity is strongly influenced by the amino acids directly flanking the splice junctions. however, to date no reliable prediction can be made whether or not a split intein is active in a particular foreign extein context. here we describe splicefinder, a pcr-based method, allowing f ...201324023792
a two-domain elevator mechanism for sodium/proton antiport.sodium/proton (na(+)/h(+)) antiporters, located at the plasma membrane in every cell, are vital for cell homeostasis. in humans, their dysfunction has been linked to diseases, such as hypertension, heart failure and epilepsy, and they are well-established drug targets. the best understood model system for na(+)/h(+) antiport is nhaa from escherichia coli, for which both electron microscopy and crystal structures are available. nhaa is made up of two distinct domains: a core domain and a dimeriza ...201323995679
physiological role of acyl coenzyme a synthetase homologs in lipid metabolism in neurospora crassa.acyl coenzyme a (coa) synthetase (acs) enzymes catalyze the activation of free fatty acids (fas) to coa esters by a two-step thioesterification reaction. activated fas participate in a variety of anabolic and catabolic lipid metabolic pathways, including de novo complex lipid biosynthesis, fa β-oxidation, and lipid membrane remodeling. analysis of the genome sequence of the filamentous fungus neurospora crassa identified seven putative fatty acss (acs-1 through acs-7). acs-3 was found to be the ...201323873861
genome-wide transcriptional profiling of the purple sulfur bacterium allochromatium vinosum dsm 180t during growth on different reduced sulfur compounds.the purple sulfur bacterium allochromatium vinosum dsm 180(t) is one of the best-studied sulfur-oxidizing anoxygenic phototrophic bacteria, and it has been developed into a model organism for laboratory-based studies of oxidative sulfur metabolism. here, we took advantage of the organism's high metabolic versatility and performed whole-genome transcriptional profiling to investigate the response of a. vinosum cells upon exposure to sulfide, thiosulfate, elemental sulfur, or sulfite compared to p ...201323873913
sensitive and specific quantitative detection of rotavirus a by one-step real-time reverse transcription-pcr assay without antecedent double-stranded-rna denaturation.a real-time quantitative reverse transcription-pcr (qrt-pcr) assay using the recombinant thermostable thermus thermophilus (rtth) enzyme was developed to detect and quantify rotavirus a (rva). by using rtth polymerase, significant improvement was achieved over the existing real-time rt-pcr assays, which require denaturation of the rva double-stranded rna (dsrna) prior to assay setup. using a dsrna transcript for segment 7, which encodes the assay target nsp3 gene, the limit of detection for the ...201323850952
two atp-binding cassette transporters involved in (s)-2-aminoethyl-cysteine uptake in thermus thermophilus.thermus thermophilus exhibits hypersensitivity to a lysine analog, (s)-2-aminoethyl-cysteine (aec). cosmid libraries were constructed using genomes from two aec-resistant mutants, at10 and at14, and the cosmids that conferred aec resistance on the wild-type strain were isolated. when the cosmid library for mutant at14 was screened, two independent cosmids, conferring partial aec resistance to the wild type, were obtained. two cosmids carried a common genomic region from ttc0795 to ttc0810. this ...201323794618
polysaccharide-degrading thermophiles generated by heterologous gene expression in geobacillus kaustophilus hta426.thermophiles have important advantages over mesophiles as host organisms for high-temperature bioprocesses, functional production of thermostable enzymes, and efficient expression of enzymatic activities in vivo. to capitalize on these advantages of thermophiles, we describe here a new inducible gene expression system in the thermophile geobacillus kaustophilus hta426. six promoter regions in the hta426 genome were identified and analyzed for expression profiles using β-galactosidase reporter as ...201323793634
mechanism of activation of elongation factor tu by ribosome: catalytic histidine activates gtp by protonation.elongation factor tu (ef-tu) is central to prokaryotic protein synthesis as it has the role of delivering amino-acylated trnas to the ribosome. release of ef-tu, after correct binding of the ef-tu:aa-trna complex to the ribosome, is initiated by gtp hydrolysis. this reaction, whose mechanism is uncertain, is catalyzed by ef-tu, but requires activation by the ribosome. there have been a number of mechanistic proposals, including those spurred by a recent x-ray crystallographic analysis of a ribos ...201323864225
translation of cga codon repeats in yeast involves quality control components and ribosomal protein l1.translation of cga codon repeats in the yeast saccharomyces cerevisiae is inefficient, resulting in dose-dependent reduction in expression and in production of an mrna cleavage product, indicative of a stalled ribosome. here, we use genetics and translation inhibitors to understand how ribosomes respond to cga repeats. we find that cga codon repeats result in a truncated polypeptide that is targeted for degradation by ltn1, an e3 ubiquitin ligase involved in nonstop decay, although deletion of l ...201323825054
an insertion peptide in yeast glycyl-trna synthetase facilitates both productive docking and catalysis of cognate trnas.the yeast saccharomyces cerevisiae possesses two distinct glycyl-trna synthetase (glyrs) genes: grs1 and grs2. grs1 is dually functional, encoding both cytoplasmic and mitochondrial activities, while grs2 is dysfunctional and not required for growth. the protein products of these two genes, glyrs1 and glyrs2, are much alike but are distinguished by an insertion peptide of glyrs1, which is absent from glyrs2 and other eukaryotic homologues. we show that deletion or mutation of the insertion pepti ...201323816885
divergent contributions of conserved active site residues to transcription by eukaryotic rna polymerases i and ii.multisubunit rna polymerases (msrnaps) exhibit high sequence and structural homology, especially within their active sites, which is generally thought to result in msrnap functional conservation. however, we show that mutations in the trigger loop (tl) in the largest subunit of rna polymerase i (pol i) yield phenotypes unexpected from studies of pol ii. for example, a well-characterized gain-of-function mutation in pol ii results in loss of function in pol i (pol ii: rpb1- e1103g; pol i: rpa190- ...201323994471
ppr-smrs: ancient proteins with enigmatic functions.a small subset of the large pentatricopeptide repeat (ppr) protein family in higher plants contain a c-terminal small muts-related (smr) domain. although few in number, they figure prominently in the chloroplast biogenesis and retrograde signaling literature due to their striking mutant phenotypes. in this review, we summarize current knowledge of ppr-smr proteins focusing on arabidopsis and maize proteomic and mutant studies. we also examine their occurrence in other organisms and have determin ...201324004908
the temperature sensitivity of a mutation in the essential trna modification enzyme trna methyltransferase d (trmd).conditional temperature-sensitive (ts) mutations are important reagents to study essential genes. although it is commonly assumed that the ts phenotype of a specific mutation arises from thermal denaturation of the mutant enzyme, the possibility also exists that the mutation decreases the enzyme activity to a certain level at the permissive temperature and aggravates the negative effect further upon temperature upshifts. resolving these possibilities is important for exploiting the ts mutation f ...201323986443
exploration of freely available web-interfaces for comparative homology modelling of microbial proteins.this study was conducted to find the best suited freely available software for modelling of proteins by taking a few sample proteins. the proteins used were small to big in size with available crystal structures for the purpose of benchmarking. key players like phyre2, swiss-model, cphmodels-3.0, homer, (ps)2, (ps)(2)-v(2), modweb were used for the comparison and model generation.201324023424
generation of a mouse model with down-regulated u50 snorna (snord50) expression and its organ-specific phenotypic modulation.box c/d-type small nucleolar rnas (snornas) are functional rnas responsible for mediating 2'-o-ribose methylation of ribosomal rnas (rrnas) within the nucleolus. in the past years, evidence for the involvement of human u50 snorna in tumorigenesis has been accumulating. we previously identified u50hg, a non-protein-coding gene that hosted a box c/d-type u50 snorna, in a chromosomal breakpoint in a human b-cell lymphoma. mouse genome analysis revealed four mouse u50 (mu50) host-genes: three mu50hg ...201323991050
reca protein recruits structural maintenance of chromosomes (smc)-like recn protein to dna double-strand breaks.escherichia coli recn is an smc (structural maintenance of chromosomes) family protein that is required for dna double-strand break (dsb) repair. previous studies show that gfp-recn forms nucleoid-associated foci in response to dna damage, but the mechanism by which recn is recruited to the nucleoid is unknown. here, we show that the assembly of gfp-recn foci on the nucleoid in response to dna damage involves a functional interaction between recn and reca. a novel reca allele identified in this ...201323974212
crystal structure of ruvc resolvase in complex with holliday junction substrate.the key intermediate in genetic recombination is the holliday junction (hj), a four-way dna structure. at the end of recombination, hjs are cleaved by specific nucleases called resolvases. in gram-negative bacteria, this cleavage is performed by ruvc, a dimeric endonuclease that belongs to the retroviral integrase superfamily. here, we report the first crystal structure of ruvc in complex with a synthetic hj solved at 3.75 å resolution. the junction in the complex is in an unfolded 2-fold symmet ...201323980027
structural phylogenomics retrodicts the origin of the genetic code and uncovers the evolutionary impact of protein flexibility.the genetic code shapes the genetic repository. its origin has puzzled molecular scientists for over half a century and remains a long-standing mystery. here we show that the origin of the genetic code is tightly coupled to the history of aminoacyl-trna synthetase enzymes and their interactions with trna. a timeline of evolutionary appearance of protein domain families derived from a structural census in hundreds of genomes reveals the early emergence of the 'operational' rna code and the late i ...201323991065
discovery of a potent benzoxaborole-based anti-pneumococcal agent targeting leucyl-trna synthetase.streptococcus pneumoniae causes bacterial pneumonia with high mortality and morbidity. the emergency of multidrug-resistant bacteria threatens the treatment of the disease. leucyl-trna synthetase (leurs) plays an essential role in cellular translation and is an attractive drug target for antimicrobial development. here we report the compound zcl039, a benzoxaborole-based derivative of an2690, as a potent anti-pneumococcal agent that inhibits s. pneumoniae leurs (spleurs) activity. we show using ...201323959225
the legionella pneumophila kai operon is implicated in stress response and confers fitness in competitive environments.legionella pneumophila uses aquatic protozoa as replication niche and protection from harsh environments. although l. pneumophila is not known to have a circadian clock, it encodes homologues of the kaibc proteins of cyanobacteria that regulate circadian gene expression. we show that l. pneumophila kaib, kaic and the downstream gene lpp1114, are transcribed as a unit under the control of the stress sigma factor rpos. kaic and kaib of l. pneumophila do not interact as evidenced by yeast and bacte ...201323957615
the legionella pneumophila kai operon is implicated in stress response and confers fitness in competitive environments.legionella pneumophila uses aquatic protozoa as replication niche and protection from harsh environments. although l. pneumophila is not known to have a circadian clock, it encodes homologues of the kaibc proteins of cyanobacteria that regulate circadian gene expression. we show that l. pneumophila kaib, kaic and the downstream gene lpp1114, are transcribed as a unit under the control of the stress sigma factor rpos. kaic and kaib of l. pneumophila do not interact as evidenced by yeast and bacte ...201323957615
serial femtosecond x-ray diffraction of 30s ribosomal subunit microcrystals in liquid suspension at ambient temperature using an x-ray free-electron laser.high-resolution ribosome structures determined by x-ray crystallography have provided important insights into the mechanism of translation. such studies have thus far relied on large ribosome crystals kept at cryogenic temperatures to reduce radiation damage. here, the application of serial femtosecond x-ray crystallography (sfx) using an x-ray free-electron laser (xfel) to obtain diffraction data from ribosome microcrystals in liquid suspension at ambient temperature is described. 30s ribosomal ...201323989164
identifying ligand-binding hot spots in proteins using brominated fragments.high-quality crystals of thermus thermophilus ef-tu in the gtp-bound conformation at 1.7-2.7 å resolution were used to test 18 small organic molecules, all brominated for confident identification in the anomalous difference maps. from this relatively small collection, it was possible to identify a small molecule bound in the functionally important trna cca-end binding pocket. the antibiotic ge2270 a is known to interact with the same pocket in ef-tu and to disrupt the association with trna. brom ...201323989163
hexamers of the type ii secretion atpase gspe from vibrio cholerae with increased atpase activity.the type ii secretion system (t2ss), a multiprotein machinery spanning two membranes in gram-negative bacteria, is responsible for the secretion of folded proteins from the periplasm across the outer membrane. the critical multidomain t2ss assembly atpase gspe(epse) had not been structurally characterized as a hexamer. here, four hexamers of vibrio cholerae gspe(epse) are obtained when fused to hcp1 as an assistant hexamer, as shown with native mass spectrometry. the enzymatic activity of the gs ...201323954505
mutants of phage bil67 ruvc with enhanced holliday junction binding selectivity and resolution symmetry.viral and bacterial holliday junction resolvases differ in specificity with the former typically being more promiscuous, acting on a variety of branched dna substrates, while the latter exclusively targets holliday junctions. we have determined the crystal structure of a ruvc resolvase from bacteriophage bil67 to help identify features responsible for dna branch discrimination. comparisons between phage and bacterial ruvc structures revealed significant differences in the number and position of ...201323888987
one number does not fit all: mapping local variations in resolution in cryo-em reconstructions.the resolution of density maps from single particle analysis is usually measured in terms of the highest spatial frequency to which consistent information has been obtained. this calculation represents an average over the entire reconstructed volume. in practice, however, substantial local variations in resolution may occur, either from intrinsic properties of the specimen or for technical reasons such as a non-isotropic distribution of viewing orientations. to address this issue, we propose the ...201323954653
an automated approach to network features of protein structure ensembles.network theory applied to protein structures provides insights into numerous problems of biological relevance. the explosion in structural data available from pdb and simulations establishes a need to introduce a standalone-efficient program that assembles network concepts/parameters under one hood in an automated manner. herein, we discuss the development/application of an exhaustive, user-friendly, standalone program package named psn-ensemble, which can handle structural ensembles generated t ...201323934896
novel extracellular phb depolymerase from streptomyces ascomycinicus: phb copolymers degradation in acidic conditions.the ascomycin-producer strain streptomyces ascomycinicus has been proven to be an extracellular poly(r)-3-hydroxybutyrate (phb) degrader. the fkbu gene, encoding a phb depolymerase (phaz sa ), has been cloned in e. coli and rhodococcus sp. t104 strains for gene expression. gram-positive host rhodococcus sp. t104 was able to produce and secrete to the extracellular medium an active protein form. phaz sa was purified by two hydrophobic interaction chromatographic steps, and afterwards was biochemi ...201323951224
rnase j participates in a pentatricopeptide repeat protein-mediated 5' end maturation of chloroplast mrnas.nucleus-encoded ribonucleases and rna-binding proteins influence chloroplast gene expression through their roles in rna maturation and stability. one mechanism for mrna 5' end maturation posits that sequence-specific pentatricopeptide repeat (ppr) proteins define termini by blocking the 5'→3' exonucleolytic activity of ribonuclease j (rnase j). to test this hypothesis in vivo, virus-induced gene silencing was used to reduce the expression of three ppr proteins and rnase j, both individually and ...201323921629
a comparison of structural and evolutionary attributes of escherichia coli and thermus thermophilus small ribosomal subunits: signatures of thermal adaptation.here we compare the structural and evolutionary attributes of thermus thermophilus and escherichia coli small ribosomal subunits (ssu). our results indicate that with few exceptions, thermophilic 16s ribosomal rna (16s rrna) is densely packed compared to that of mesophilic at most of the analogous spatial regions. in addition, we have located species-specific cavity clusters (ssccs) in both species. e. coli ssccs are numerous and larger compared to t. thermophilus ssccs, which again indicates de ...201323940533
confounders of mutation-rate estimators: selection and phenotypic lag in thermus thermophilus.in a recent description of the rate and character of spontaneous mutation in the hyperthermophilic bacterium thermus thermophilus, the mutation rate was observed to be substantially lower than seen in several mesophiles. subsequently, a report appeared indicating that this bacterium maintains an average of about 4.5 genomes per cell. this number of genomes might result in a segregation lag for the expression of a recessive mutation and might therefore lead to an underestimate of the rate of muta ...201323916418
protein biosynthesis in mitochondria.translation, that is biosynthesis of polypeptides in accordance with information encoded in the genome, is one of the most important processes in the living cell, and it has been in the spotlight of international research for many years. the mechanisms of protein biosynthesis in bacteria and in the eukaryotic cytoplasm are now understood in great detail. however, significantly less is known about translation in eukaryotic mitochondria, which is characterized by a number of unusual features. in t ...201324228873
mutations in mitochondrial ribosomal protein mrpl12 leads to growth retardation, neurological deterioration and mitochondrial translation deficiency.multiple respiratory chain deficiencies represent a common cause of mitochondrial diseases and are associated with a wide range of clinical symptoms. we report a subject, born to consanguineous parents, with growth retardation and neurological deterioration. multiple respiratory chain deficiency was found in muscle and fibroblasts of the subject as well as abnormal assembly of complexes i and iv. a microsatellite genotyping of the family members detected only one region of homozygosity on chromo ...201323603806
role of the ribosomal p-site elements of m²g966, m⁵c967, and the s9 c-terminal tail in maintenance of the reading frame during translational elongation in escherichia coli.the ribosomal p-site hosts the peptidyl-trnas during translation elongation. which p-site elements support these trna species to maintain codon-anticodon interactions has remained unclear. we investigated the effects of p-site features of methylations of g966, c967, and the conserved c-terminal tail sequence of ser, lys, and arg (skr) of the s9 ribosomal protein in maintenance of the translational reading frame of an mrna. we generated escherichia coli strains deleted for the skr sequence in s9 ...201323729652
argonaute protein as a linker to command center of physiological processes.micrornas (mirnas) post-transcriptionally regulate gene expression by binding to target mrnas with perfect or imperfect complementarity, recruiting an argonaute (ago) protein complex that usually results in degradation or translational repression of the target mrna. ago proteins function as the slicer enzyme in mirna and small interfering rna (sirna) pathways involved in human physiological and pathophysiological processes, such as antiviral responses and disease formation. although the past dec ...201323997530
pioglitazone leads to an inactivation and disassembly of complex i of the mitochondrial respiratory chain.thiazolidinediones are antidiabetic agents that increase insulin sensitivity but reduce glucose oxidation, state 3 respiration, and activity of complex i of the mitochondrial respiratory chain (mrc). the mechanisms of the latter effects are unclear. the aim of this study was to determine the mechanisms by which pioglitazone (pgz), a member of the thiazolidinedione class of antidiabetic agents, decreases the activity of the mrc. in isolated mitochondria from mouse liver, we measured the effects o ...201323915000
molecular basis of hhq biosynthesis: molecular dynamics simulations, enzyme kinetic and surface plasmon resonance studies.pqs (pseudomonasquinolone signal) and its precursor hhq are signal molecules of the p. aeruginosa quorum sensing system. they explicate their role in mammalian pathogenicity by binding to the receptor pqsr that induces virulence factor production and biofilm formation. the enzyme pqsd catalyses the biosynthesis of hhq.201323916145
distinction between the cfr methyltransferase conferring antibiotic resistance and the housekeeping rlmn methyltransferase.the cfr gene encodes the cfr methyltransferase that primarily methylates c-8 in a2503 of 23s rrna in the peptidyl transferase region of bacterial ribosomes. the methylation provides resistance to six classes of antibiotics of clinical and veterinary importance. the rlmn gene encodes the rlmn methyltransferase that methylates c-2 in a2503 in 23s rrna and a37 in trna, but rlmn does not significantly influence antibiotic resistance. the enzymes are homologous and use the same mechanism involving ra ...201323752511
an innate twist between crick's wobble and watson-crick base pairs.non-watson-crick pairs like the g·u wobble are frequent in rna duplexes. their geometric dissimilarity (nonisostericity) with the watson-crick base pairs and among themselves imparts structural variations decisive for biological functions. through a novel circular representation of base pairs, a simple and general metric scheme for quantification of base-pair nonisostericity, in terms of residual twist and radial difference that can also envisage its mechanistic effect, is proposed. the scheme i ...201323861536
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