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rotary atpases: models, machine elements and technical specifications.rotary atpases are molecular rotary motors involved in biological energy conversion. they either synthesize or hydrolyze the universal biological energy carrier adenosine triphosphate. recent work has elucidated the general architecture and subunit compositions of all three sub-types of rotary atpases. composite models of the intact f-, v- and a-type atpases have been constructed by fitting high-resolution x-ray structures of individual subunits or sub-complexes into low-resolution electron dens ...201323369889
morphological development and cytochrome c oxidase activity in streptomyces lividans are dependent on the action of a copper bound sco protein.copper has an important role in the life cycle of many streptomycetes, stimulating the developmental switch between vegetative mycelium and aerial hyphae concomitant with the production of antibiotics. in streptomycetes, a gene encoding for a putative sco-like protein has been identified and is part of an operon that contains two other genes predicted to handle cellular copper. we report on the sco-like protein from streptomyces lividans (sco(sl)) and present a series of experiments that firmly ...201323345541
dna repair by reversal of dna damage.endogenous and exogenous factors constantly challenge cellular dna, generating cytotoxic and/or mutagenic dna adducts. as a result, organisms have evolved different mechanisms to defend against the deleterious effects of dna damage. among these diverse repair pathways, direct dna-repair systems provide cells with simple yet efficient solutions to reverse covalent dna adducts. in this review, we focus on recent advances in the field of direct dna repair, namely, photolyase-, alkyltransferase-, an ...201323284047
a structural basis for streptomycin-induced misreading of the genetic code.during protein synthesis, the ribosome selects aminoacyl-transfer rnas with anticodons matching the messenger rna codon present in the a site of the small ribosomal subunit. the aminoglycoside antibiotic streptomycin disrupts decoding by binding close to the site of codon recognition. here we use x-ray crystallography to define the impact of streptomycin on the decoding site of the thermus thermophilus 30s ribosomal subunit in complexes with cognate or near-cognate anticodon stem-loop analogues ...201323322043
a mutation of the rna polymerase β' subunit (rpoc) confers cephalosporin resistance in bacillus subtilis.in bacteria, mutations affecting the major catalytic subunits of rna polymerase (encoded by rpob and rpoc) emerge in response to a variety of selective pressures. here we isolated a bacillus subtilis strain with high-level resistance to cefuroxime (cef). whole-genome resequencing revealed only one missense mutation affecting an invariant residue in close proximity to the c-terminal dna-binding domain of rpoc (g1122d). genetic reconstruction experiments demonstrate that this substitution is suffi ...201323070162
characterization of recombinant fluoroquinolone-resistant pneumococcus-like isolates.fourteen fluoroquinolone-resistant streptococcal isolates with recombinant dna topoisomerase genes, preliminarily identified as pneumococci, were further characterized using phenotypic and genotypic approaches. phenotypic tests classified them as atypical pneumococci. phylogenetic relationships were analyzed by using the sequences of seven housekeeping alleles from these isolates and from isolates of streptococcus pneumoniae, streptococcus mitis, streptococcus oralis, and streptococcus pseudopne ...201323114769
molecular detection of bacteria in plant tissues, using universal 16s ribosomal dna degenerated primers.highly specific, sensitive and rapid tests are required for the detection and identification of covert bacterial contaminations in plant tissue cultures. current methods available for this purpose are tedious, time consuming, highly error prone, expensive, require advanced technical expertise and are sometimes ineffective. we report here the development of a sensitive polymerase chain reaction (pcr) based method for the rapid detection and identification of bacteria occurring in plant tissue cul ...201426019546
evolution of mitochondria reconstructed from the energy metabolism of living bacteria.the ancestors of mitochondria, or proto-mitochondria, played a crucial role in the evolution of eukaryotic cells and derived from symbiotic α-proteobacteria which merged with other microorganisms - the basis of the widely accepted endosymbiotic theory. however, the identity and relatives of proto-mitochondria remain elusive. here we show that methylotrophic α-proteobacteria could be the closest living models for mitochondrial ancestors. we reached this conclusion after reconstructing the possibl ...201424804722
biotechnology of polyketides: new breath of life for the novel antibiotic genetic pathways discovery through metagenomics.the discovery of secondary metabolites produced by microorganisms (e.g., penicillin in 1928) and the beginning of their industrial application (1940) opened new doors to what has been the main medication source for the treatment of infectious diseases and tumors. in fact, approximately 80 years after the discovery of the first antibiotic compound, and despite all of the warnings about the failure of the "goose that laid the golden egg," the potential of this wealth is still inexorable: simply ad ...201424688489
biotechnology of polyketides: new breath of life for the novel antibiotic genetic pathways discovery through metagenomics.the discovery of secondary metabolites produced by microorganisms (e.g., penicillin in 1928) and the beginning of their industrial application (1940) opened new doors to what has been the main medication source for the treatment of infectious diseases and tumors. in fact, approximately 80 years after the discovery of the first antibiotic compound, and despite all of the warnings about the failure of the "goose that laid the golden egg," the potential of this wealth is still inexorable: simply ad ...201424688489
the dynamic nature of genomes across the tree of life.genomes are dynamic in lineages across the tree of life. among bacteria and archaea, for example, dna content varies throughout life cycles, and nonbinary cell division in diverse lineages indicates the need for coordination of the inheritance of genomes. these observations contrast with the textbook view that bacterial and archaeal genomes are monoploid (i.e., single copied) and fixed both within species and throughout an individual's lifetime. here, we synthesize information on three aspects o ...201424500971
conserved evolutionary units in the heme-copper oxidase superfamily revealed by novel homologous protein families.the heme-copper oxidase (hco) superfamily includes hcos in aerobic respiratory chains and nitric oxide reductases (nors) in the denitrification pathway. the hco/nor catalytic subunit has a core structure consisting of 12 transmembrane helices (tmhs) arranged in three-fold rotational pseudosymmetry, with six conserved histidines for heme and metal binding. using sensitive sequence similarity searches, we detected a number of novel hco/nor homologs and named them hco homology (hcoh) proteins. seve ...201424931479
convergent evolution of aua decoding in bacteria and archaea.deciphering aua codons is a difficult task for organisms, because aua and aug specify isoleucine (ile) and methionine (met), separately. each of the other purine-ending sense co-don sets (nnr) specifies a single amino acid in the universal genetic code. in bacteria and archaea, the cytidine derivatives, 2-lysylcytidine (l or lysidine) and 2-agmatinylcytidine (agm(2)c or agmatidine), respectively, are found at the first letter of the anticodon of trna(ile) responsible for aua codons. these modifi ...201425629511
convergent evolution of aua decoding in bacteria and archaea.deciphering aua codons is a difficult task for organisms, because aua and aug specify isoleucine (ile) and methionine (met), separately. each of the other purine-ending sense co-don sets (nnr) specifies a single amino acid in the universal genetic code. in bacteria and archaea, the cytidine derivatives, 2-lysylcytidine (l or lysidine) and 2-agmatinylcytidine (agm(2)c or agmatidine), respectively, are found at the first letter of the anticodon of trna(ile) responsible for aua codons. these modifi ...201425629511
two-subunit enzymes involved in eukaryotic post-transcriptional trna modification.trna modifications are crucial for efficient and accurate protein translation, with defects often linked to disease. there are 7 cytoplasmic trna modifications in the yeast saccharomyces cerevisiae that are formed by an enzyme consisting of a catalytic subunit and an auxiliary protein, 5 of which require only a single subunit in bacteria, and 2 of which are not found in bacteria. these enzymes include the deaminase tad2-tad3, and the methyltransferases trm6-trm61, trm8-trm82, trm7-trm732, and tr ...201425625329
two-subunit enzymes involved in eukaryotic post-transcriptional trna modification.trna modifications are crucial for efficient and accurate protein translation, with defects often linked to disease. there are 7 cytoplasmic trna modifications in the yeast saccharomyces cerevisiae that are formed by an enzyme consisting of a catalytic subunit and an auxiliary protein, 5 of which require only a single subunit in bacteria, and 2 of which are not found in bacteria. these enzymes include the deaminase tad2-tad3, and the methyltransferases trm6-trm61, trm8-trm82, trm7-trm732, and tr ...201425625329
the identification and characterization of non-coding and coding rnas and their modified nucleosides by mass spectrometry.the analysis of ribonucleic acids (rna) by mass spectrometry has been a valuable analytical approach for more than 25 years. in fact, mass spectrometry has become a method of choice for the analysis of modified nucleosides from rna isolated out of biological samples. this review summarizes recent progress that has been made in both nucleoside and oligonucleotide mass spectral analysis. applications of mass spectrometry in the identification, characterization and quantification of modified nucleo ...201425616408
the identification and characterization of non-coding and coding rnas and their modified nucleosides by mass spectrometry.the analysis of ribonucleic acids (rna) by mass spectrometry has been a valuable analytical approach for more than 25 years. in fact, mass spectrometry has become a method of choice for the analysis of modified nucleosides from rna isolated out of biological samples. this review summarizes recent progress that has been made in both nucleoside and oligonucleotide mass spectral analysis. applications of mass spectrometry in the identification, characterization and quantification of modified nucleo ...201425616408
pseudomonas aeruginosa isolates from dental unit waterlines can be divided in two distinct groups, including one displaying phenotypes similar to isolates from cystic fibrosis patients.pseudomonas aeruginosa displays broad genetic diversity, giving it an astonishing capacity to adapt to a variety of environments and to infect a wide range of hosts. while many p. aeruginosa isolates of various origins have been analyzed, isolates from cystic fibrosis (cf) patients have received the most attention. less is known about the genetic and phenotypic diversity of p. aeruginosa isolates that colonize other environments where flourishing biofilms can be found. in the present study, 29 p ...201425653647
pseudomonas aeruginosa isolates from dental unit waterlines can be divided in two distinct groups, including one displaying phenotypes similar to isolates from cystic fibrosis patients.pseudomonas aeruginosa displays broad genetic diversity, giving it an astonishing capacity to adapt to a variety of environments and to infect a wide range of hosts. while many p. aeruginosa isolates of various origins have been analyzed, isolates from cystic fibrosis (cf) patients have received the most attention. less is known about the genetic and phenotypic diversity of p. aeruginosa isolates that colonize other environments where flourishing biofilms can be found. in the present study, 29 p ...201425653647
an mrps12 mutation modifies aminoglycoside sensitivity caused by 12s rrna mutations.several homoplasmic pathologic mutations in mitochondrial dna, such as those causing leber hereditary optic neuropathy or non-syndromic hearing loss, show incomplete penetrance. therefore, other elements must modify their pathogenicity. discovery of these modifying factors is not an easy task because in multifactorial diseases conventional genetic approaches may not always be informative. here, we have taken an evolutionary approach to unmask putative modifying factors for a particular homoplasm ...201425642242
an mrps12 mutation modifies aminoglycoside sensitivity caused by 12s rrna mutations.several homoplasmic pathologic mutations in mitochondrial dna, such as those causing leber hereditary optic neuropathy or non-syndromic hearing loss, show incomplete penetrance. therefore, other elements must modify their pathogenicity. discovery of these modifying factors is not an easy task because in multifactorial diseases conventional genetic approaches may not always be informative. here, we have taken an evolutionary approach to unmask putative modifying factors for a particular homoplasm ...201425642242
trigger loop folding determines transcription rate of escherichia coli's rna polymerase.two components of the rna polymerase (rnap) catalytic center, the bridge helix and the trigger loop (tl), have been linked with changes in elongation rate and pausing. here, single molecule experiments with the wt and two tl-tip mutants of the escherichia coli enzyme reveal that tip mutations modulate rnap's pause-free velocity, identifying tl conformational changes as one of two rate-determining steps in elongation. consistent with this observation, we find a direct correlation between helix pr ...201425552559
trigger loop folding determines transcription rate of escherichia coli's rna polymerase.two components of the rna polymerase (rnap) catalytic center, the bridge helix and the trigger loop (tl), have been linked with changes in elongation rate and pausing. here, single molecule experiments with the wt and two tl-tip mutants of the escherichia coli enzyme reveal that tip mutations modulate rnap's pause-free velocity, identifying tl conformational changes as one of two rate-determining steps in elongation. consistent with this observation, we find a direct correlation between helix pr ...201425552559
a structural determinant in the uracil dna glycosylase superfamily for the removal of uracil from adenine/uracil base pairs.the uracil dna glycosylase superfamily consists of several distinct families. family 2 mismatch-specific uracil dna glycosylase (mug) from escherichia coli is known to exhibit glycosylase activity on three mismatched base pairs, t/u, g/u and c/u. family 1 uracil n-glycosylase (ung) from e. coli is an extremely efficient enzyme that can remove uracil from any uracil-containing base pairs including the a/u base pair. here, we report the identification of an important structural determinant that un ...201425550433
a structural determinant in the uracil dna glycosylase superfamily for the removal of uracil from adenine/uracil base pairs.the uracil dna glycosylase superfamily consists of several distinct families. family 2 mismatch-specific uracil dna glycosylase (mug) from escherichia coli is known to exhibit glycosylase activity on three mismatched base pairs, t/u, g/u and c/u. family 1 uracil n-glycosylase (ung) from e. coli is an extremely efficient enzyme that can remove uracil from any uracil-containing base pairs including the a/u base pair. here, we report the identification of an important structural determinant that un ...201425550433
structure of the pseudomonas aeruginosa transamidosome reveals unique aspects of bacterial trna-dependent asparagine biosynthesis.many prokaryotes lack a trna synthetase to attach asparagine to its cognate trna(asn), and instead synthesize asparagine from trna(asn)-bound aspartate. this conversion involves two enzymes: a nondiscriminating aspartyl-trna synthetase (nd-asprs) that forms asp-trna(asn), and a heterotrimeric amidotransferase gatcab that amidates asp-trna(asn) to form asn-trna(asn) for use in protein synthesis. nd-asprs, gatcab, and trna(asn) may assemble in an ∼400-kda complex, known as the asn-transamidosome, ...201425548166
structure of the pseudomonas aeruginosa transamidosome reveals unique aspects of bacterial trna-dependent asparagine biosynthesis.many prokaryotes lack a trna synthetase to attach asparagine to its cognate trna(asn), and instead synthesize asparagine from trna(asn)-bound aspartate. this conversion involves two enzymes: a nondiscriminating aspartyl-trna synthetase (nd-asprs) that forms asp-trna(asn), and a heterotrimeric amidotransferase gatcab that amidates asp-trna(asn) to form asn-trna(asn) for use in protein synthesis. nd-asprs, gatcab, and trna(asn) may assemble in an ∼400-kda complex, known as the asn-transamidosome, ...201425548166
conformational preferences underlying reduced activity of a thermophilic ribonuclease h.the conformational basis for reduced activity of the thermophilic ribonuclease hi enzyme from thermus thermophilus, compared to its mesophilic homolog from escherichia coli, is elucidated using a combination of nmr spectroscopy and molecular dynamics (md) simulations. explicit-solvent all-atom md simulations of the two wild-type proteins and an e. coli mutant in which a glycine residue is inserted after position 80 to mimic the t. thermophilus protein reproduce the differences in conformational ...201425550198
conformational preferences underlying reduced activity of a thermophilic ribonuclease h.the conformational basis for reduced activity of the thermophilic ribonuclease hi enzyme from thermus thermophilus, compared to its mesophilic homolog from escherichia coli, is elucidated using a combination of nmr spectroscopy and molecular dynamics (md) simulations. explicit-solvent all-atom md simulations of the two wild-type proteins and an e. coli mutant in which a glycine residue is inserted after position 80 to mimic the t. thermophilus protein reproduce the differences in conformational ...201425550198
crystal structure of thermobifida fusca cse1 reveals target dna binding site.the clustered regularly interspaced short palindromic repeats (crispr)-crispr-associated (cas) defense system is the only adaptive and inheritable immunity found in prokaryotes. the immunity is achieved through a multistep process of adaptation, expression, and interference. in the type i-e system, interference is mediated by the crispr-associated complex for antiviral defense (cascade), which recognizes invading double-stranded dna (dsdna) through the protospacer adjacent motif (pam) by one of ...201425420472
crystal structure of thermobifida fusca cse1 reveals target dna binding site.the clustered regularly interspaced short palindromic repeats (crispr)-crispr-associated (cas) defense system is the only adaptive and inheritable immunity found in prokaryotes. the immunity is achieved through a multistep process of adaptation, expression, and interference. in the type i-e system, interference is mediated by the crispr-associated complex for antiviral defense (cascade), which recognizes invading double-stranded dna (dsdna) through the protospacer adjacent motif (pam) by one of ...201425420472
trnas as antibiotic targets.transfer rnas (trnas) are central players in the protein translation machinery and as such are prominent targets for a large number of natural and synthetic antibiotics. this review focuses on the role of trnas in bacterial antibiosis. we will discuss examples of antibiotics that target multiple stages in trna biology from trna biogenesis and modification, mature trnas, aminoacylation of trna as well as prevention of proper trna function by small molecules binding to the ribosome. finally, the r ...201425547494
trnas as antibiotic targets.transfer rnas (trnas) are central players in the protein translation machinery and as such are prominent targets for a large number of natural and synthetic antibiotics. this review focuses on the role of trnas in bacterial antibiosis. we will discuss examples of antibiotics that target multiple stages in trna biology from trna biogenesis and modification, mature trnas, aminoacylation of trna as well as prevention of proper trna function by small molecules binding to the ribosome. finally, the r ...201425547494
cmr4 is the slicer in the rna-targeting cmr crispr complex.clustered regularly interspaced short palindromic repeat (crispr) loci and crispr-associated (cas) proteins form an adaptive immune system that protects prokaryotes against plasmids and viruses. the cmr complex, a type iii-b effector complex, uses the crispr rna (crrna) as a guide to target rna. here, we show that the cmr complex of pyrococcus furiosus cleaves rna at multiple sites that are 6 nt apart and are positioned relative to the 5'-end of the crrna. we identified cmr4 as the slicer and de ...201425541196
cmr4 is the slicer in the rna-targeting cmr crispr complex.clustered regularly interspaced short palindromic repeat (crispr) loci and crispr-associated (cas) proteins form an adaptive immune system that protects prokaryotes against plasmids and viruses. the cmr complex, a type iii-b effector complex, uses the crispr rna (crrna) as a guide to target rna. here, we show that the cmr complex of pyrococcus furiosus cleaves rna at multiple sites that are 6 nt apart and are positioned relative to the 5'-end of the crrna. we identified cmr4 as the slicer and de ...201425541196
transfer rna methyltransferases from thermoplasma acidophilum, a thermoacidophilic archaeon.we investigated trna methyltransferase activities in crude cell extracts from the thermoacidophilic archaeon thermoplasma acidophilum. we analyzed the modified nucleosides in native initiator and elongator trnamet, predicted the candidate genes for the trna methyltransferases on the basis of the trnamet and trnaleu sequences, and characterized trm5, trm1 and trm56 by purifying recombinant proteins. we found that the ta0997, ta0931, and ta0836 genes of t. acidophilum encode trm1, trm56 and trm5, ...201425546389
transfer rna methyltransferases from thermoplasma acidophilum, a thermoacidophilic archaeon.we investigated trna methyltransferase activities in crude cell extracts from the thermoacidophilic archaeon thermoplasma acidophilum. we analyzed the modified nucleosides in native initiator and elongator trnamet, predicted the candidate genes for the trna methyltransferases on the basis of the trnamet and trnaleu sequences, and characterized trm5, trm1 and trm56 by purifying recombinant proteins. we found that the ta0997, ta0931, and ta0836 genes of t. acidophilum encode trm1, trm56 and trm5, ...201425546389
a novel cytosolic nadh:quinone oxidoreductase from methanothermobacter marburgensis.methanothermobacter marburgensis is a strictly anaerobic, thermophilic methanogenic archaeon that uses methanogenesis to convert h2 and co2 to energy. m. marburgensis is one of the best-studied methanogens, and all genes required for methanogenic metabolism have been identified. nonetheless, the present study describes a gene (gene id 9704440) coding for a putative201425372605
ttomp85, a β-barrel assembly protein, functions by barrel augmentation.outer membrane proteins are vital for gram-negative bacteria and organisms that inherited organelles from them. proteins from the omp85/bama family conduct the insertion of membrane proteins into the outer membrane. we show that an eight-stranded outer membrane β-barrel protein, ttoa, is inserted and folded into liposomes by an omp85 homologue. furthermore, we recorded the channel conductance of this omp85 protein in black lipid membranes, alone and in the presence of peptides comprising the seq ...201425537637
ttomp85, a β-barrel assembly protein, functions by barrel augmentation.outer membrane proteins are vital for gram-negative bacteria and organisms that inherited organelles from them. proteins from the omp85/bama family conduct the insertion of membrane proteins into the outer membrane. we show that an eight-stranded outer membrane β-barrel protein, ttoa, is inserted and folded into liposomes by an omp85 homologue. furthermore, we recorded the channel conductance of this omp85 protein in black lipid membranes, alone and in the presence of peptides comprising the seq ...201425537637
enhancing allosteric inhibition in thermus thermophilus phosphofructokinase.the coupling between the binding of the substrate fru-6-p and the inhibitor phospho(enol)pyruvate (pep) in phosphofructokinase (pfk) from the extreme thermophile thermus thermophilus is much weaker than that seen in a pfk from bacillus stearothermophilus. from the crystal structures of bacillus stearothermophilus pfk (bspfk) the residues at positions 59, 158, and 215 in bspfk are located on the path leading from the allosteric site to the nearest active site and are part of the intricate hydroge ...201425531642
enhancing allosteric inhibition in thermus thermophilus phosphofructokinase.the coupling between the binding of the substrate fru-6-p and the inhibitor phospho(enol)pyruvate (pep) in phosphofructokinase (pfk) from the extreme thermophile thermus thermophilus is much weaker than that seen in a pfk from bacillus stearothermophilus. from the crystal structures of bacillus stearothermophilus pfk (bspfk) the residues at positions 59, 158, and 215 in bspfk are located on the path leading from the allosteric site to the nearest active site and are part of the intricate hydroge ...201425531642
the bacterial flagellar protein export apparatus processively transports flagellar proteins even with extremely infrequent atp hydrolysis.for self-assembly of the bacterial flagellum, a specific protein export apparatus utilizes atp and proton motive force (pmf) as the energy source to transport component proteins to the distal growing end. the export apparatus consists of a transmembrane pmf-driven export gate and a cytoplasmic atpase complex composed of flih, flii and flij. the flii(6)flij complex is structurally similar to the α(3)β(3)γ complex of f(o)f(1)-atpase. flij allows the gate to efficiently utilize pmf to drive flagell ...201425531309
molecular details of a starch utilization pathway in the human gut symbiont eubacterium rectale.eubacterium rectale is a prominent human gut symbiont yet little is known about the molecular strategies this bacterium has developed to acquire nutrients within the competitive gut ecosystem. starch is one of the most abundant glycans in the human diet, and e. rectale increases in vivo when the host consumes a diet rich in resistant starch, although it is not a primary degrader of this glycan. here we present the results of a quantitative proteomics study in which we identify two glycoside hydr ...201425388295
molecular details of a starch utilization pathway in the human gut symbiont eubacterium rectale.eubacterium rectale is a prominent human gut symbiont yet little is known about the molecular strategies this bacterium has developed to acquire nutrients within the competitive gut ecosystem. starch is one of the most abundant glycans in the human diet, and e. rectale increases in vivo when the host consumes a diet rich in resistant starch, although it is not a primary degrader of this glycan. here we present the results of a quantitative proteomics study in which we identify two glycoside hydr ...201425388295
evolution of oligomeric state through allosteric pathways that mimic ligand binding.evolution and design of protein complexes are almost always viewed through the lens of amino acid mutations at protein interfaces. we showed previously that residues not involved in the physical interaction between proteins make important contributions to oligomerization by acting indirectly or allosterically. in this work, we sought to investigate the mechanism by which allosteric mutations act, using the example of the pyrr family of pyrimidine operon attenuators. in this family, a perfectly s ...201425525255
metatranscriptomics of n2-fixing cyanobacteria in the amazon river plume.biological n2 fixation is an important nitrogen source for surface ocean microbial communities. however, nearly all information on the diversity and gene expression of organisms responsible for oceanic n2 fixation in the environment has come from targeted approaches that assay only a small number of genes and organisms. using genomes of diazotrophic cyanobacteria to extract reads from extensive meta-genomic and -transcriptomic libraries, we examined diazotroph diversity and gene expression from ...201425514535
metatranscriptomics of n2-fixing cyanobacteria in the amazon river plume.biological n2 fixation is an important nitrogen source for surface ocean microbial communities. however, nearly all information on the diversity and gene expression of organisms responsible for oceanic n2 fixation in the environment has come from targeted approaches that assay only a small number of genes and organisms. using genomes of diazotrophic cyanobacteria to extract reads from extensive meta-genomic and -transcriptomic libraries, we examined diazotroph diversity and gene expression from ...201425514535
structural basis for the interaction of protein s1 with the escherichia coli ribosome.in gram-negative bacteria, the multi-domain protein s1 is essential for translation initiation, as it recruits the mrna and facilitates its localization in the decoding centre. in sharp contrast to its functional importance, s1 is still lacking from the high-resolution structures available for escherichia coli and thermus thermophilus ribosomes and thus the molecular mechanism governing the s1-ribosome interaction has still remained elusive. here, we present the structure of the n-terminal s1 do ...201425510494
structural basis for the interaction of protein s1 with the escherichia coli ribosome.in gram-negative bacteria, the multi-domain protein s1 is essential for translation initiation, as it recruits the mrna and facilitates its localization in the decoding centre. in sharp contrast to its functional importance, s1 is still lacking from the high-resolution structures available for escherichia coli and thermus thermophilus ribosomes and thus the molecular mechanism governing the s1-ribosome interaction has still remained elusive. here, we present the structure of the n-terminal s1 do ...201425510494
crystal structure of subunits d and f in complex gives insight into energy transmission of the eukaryotic v-atpase from saccharomyces cerevisiae.eukaryotic v1vo-atpases hydrolyze atp in the v1 domain coupled to ion pumping in vo. a unique mode of regulation of v-atpases is the reversible disassembly of v1 and vo, which reduces atpase activity and causes silencing of ion conduction. the subunits d and f are proposed to be key in these enzymatic processes. here, we describe the structures of two conformations of the subunit df assembly of saccharomyces cerevisiae (scdf) v-atpase at 3.1 å resolution. subunit d (scd) consists of a long pair ...201425505269
crystal structure of subunits d and f in complex gives insight into energy transmission of the eukaryotic v-atpase from saccharomyces cerevisiae.eukaryotic v1vo-atpases hydrolyze atp in the v1 domain coupled to ion pumping in vo. a unique mode of regulation of v-atpases is the reversible disassembly of v1 and vo, which reduces atpase activity and causes silencing of ion conduction. the subunits d and f are proposed to be key in these enzymatic processes. here, we describe the structures of two conformations of the subunit df assembly of saccharomyces cerevisiae (scdf) v-atpase at 3.1 å resolution. subunit d (scd) consists of a long pair ...201425505269
thermoadaptation-directed enzyme evolution in an error-prone thermophile derived from geobacillus kaustophilus hta426.thermostability is an important property of enzymes utilized for practical applications because it allows long-term storage and use as catalysts. in this study, we constructed an error-prone strain of the thermophile geobacillus kaustophilus hta426 and investigated thermoadaptation-directed enzyme evolution using the strain. a mutation frequency assay using the antibiotics rifampin and streptomycin revealed that g. kaustophilus had substantially higher mutability than escherichia coli and bacill ...201425326311
thermoadaptation-directed enzyme evolution in an error-prone thermophile derived from geobacillus kaustophilus hta426.thermostability is an important property of enzymes utilized for practical applications because it allows long-term storage and use as catalysts. in this study, we constructed an error-prone strain of the thermophile geobacillus kaustophilus hta426 and investigated thermoadaptation-directed enzyme evolution using the strain. a mutation frequency assay using the antibiotics rifampin and streptomycin revealed that g. kaustophilus had substantially higher mutability than escherichia coli and bacill ...201425326311
a mechanistic model of cor15 protein function in plant freezing tolerance: integration of structural and functional characteristics.plants as sessile organisms are strongly challenged by environmental stresses. many plants species are able to cold-acclimate, acquiring higher freezing tolerance upon exposure to low but non-freezing temperatures. among a plethora of adaptational processes, this involves the accumulation of cold regulated (cor) proteins that are assumed to stabilize and protect cellular structures during freezing. however, their molecular functions are largely unknown. we recently reported a comprehensive study ...201425496049
on the validation of crystallographic symmetry and the quality of structures.in 2008, zwart and colleagues observed that the fraction of the structures deposited in the pdb alleged to have "pseudosymmetry" or "special noncrystallographic symmetry" (ncs) was about 6%, and that this percentage was rising annually. a few years later, poon and colleagues found that 2% of all the crystal structures in the pdb belonged to higher symmetry space groups than those assigned to them. here, i report an analysis of the x-ray diffraction data deposited for this class of structures, wh ...201425352397
on the validation of crystallographic symmetry and the quality of structures.in 2008, zwart and colleagues observed that the fraction of the structures deposited in the pdb alleged to have "pseudosymmetry" or "special noncrystallographic symmetry" (ncs) was about 6%, and that this percentage was rising annually. a few years later, poon and colleagues found that 2% of all the crystal structures in the pdb belonged to higher symmetry space groups than those assigned to them. here, i report an analysis of the x-ray diffraction data deposited for this class of structures, wh ...201425352397
first evidence for substrate channeling between proline catabolic enzymes: a validation of domain fusion analysis for predicting protein-protein interactions.proline dehydrogenase (prodh) and δ(1)-pyrroline-5-carboxylate (p5c) dehydrogenase (p5cdh) catalyze the four-electron oxidation of proline to glutamate via the intermediates p5c and l-glutamate-γ-semialdehyde (gsa). in gram-negative bacteria, prodh and p5cdh are fused together in the bifunctional enzyme proline utilization a (puta) whereas in other organisms prodh and p5cdh are expressed as separate monofunctional enzymes. substrate channeling has previously been shown for bifunctional putas, bu ...201425492892
first evidence for substrate channeling between proline catabolic enzymes: a validation of domain fusion analysis for predicting protein-protein interactions.proline dehydrogenase (prodh) and δ(1)-pyrroline-5-carboxylate (p5c) dehydrogenase (p5cdh) catalyze the four-electron oxidation of proline to glutamate via the intermediates p5c and l-glutamate-γ-semialdehyde (gsa). in gram-negative bacteria, prodh and p5cdh are fused together in the bifunctional enzyme proline utilization a (puta) whereas in other organisms prodh and p5cdh are expressed as separate monofunctional enzymes. substrate channeling has previously been shown for bifunctional putas, bu ...201425492892
crispr rna binding and dna target recognition by purified cascade complexes from escherichia coli.clustered regularly interspaced short palindromic repeats (crisprs) and their associated cas proteins comprise a prokaryotic rna-guided adaptive immune system that interferes with mobile genetic elements, such as plasmids and phages. the type i-e crispr interference complex cascade from escherichia coli is composed of five different cas proteins and a 61-nt-long guide rna (crrna). crrnas contain a unique 32-nt spacer flanked by a repeat-derived 5' handle (8 nt) and a 3' handle (21 nt). the space ...201425488810
crispr rna binding and dna target recognition by purified cascade complexes from escherichia coli.clustered regularly interspaced short palindromic repeats (crisprs) and their associated cas proteins comprise a prokaryotic rna-guided adaptive immune system that interferes with mobile genetic elements, such as plasmids and phages. the type i-e crispr interference complex cascade from escherichia coli is composed of five different cas proteins and a 61-nt-long guide rna (crrna). crrnas contain a unique 32-nt spacer flanked by a repeat-derived 5' handle (8 nt) and a 3' handle (21 nt). the space ...201425488810
the biosynthesis of thiol- and thioether-containing cofactors and secondary metabolites catalyzed by radical s-adenosylmethionine enzymes.sulfur atoms are present as thiol and thioether functional groups in amino acids, coenzymes, cofactors, and various products of secondary metabolic pathways. the biosynthetic pathways for several sulfur-containing biomolecules require the substitution of sulfur for hydrogen at unreactive aliphatic or electron-rich aromatic carbon atoms. examples discussed in this review include biotin, lipoic acid, methylthioether modifications found in some nucleic acids and proteins, and thioether cross-links ...201425477512
the biosynthesis of thiol- and thioether-containing cofactors and secondary metabolites catalyzed by radical s-adenosylmethionine enzymes.sulfur atoms are present as thiol and thioether functional groups in amino acids, coenzymes, cofactors, and various products of secondary metabolic pathways. the biosynthetic pathways for several sulfur-containing biomolecules require the substitution of sulfur for hydrogen at unreactive aliphatic or electron-rich aromatic carbon atoms. examples discussed in this review include biotin, lipoic acid, methylthioether modifications found in some nucleic acids and proteins, and thioether cross-links ...201425477512
fada5 a thiolase from mycobacterium tuberculosis: a steroid-binding pocket reveals the potential for drug development against tuberculosis.with the exception of hiv, tuberculosis (tb) is the leading cause of mortality among infectious diseases. the urgent need to develop new antitubercular drugs is apparent due to the increasing number of drug-resistant mycobacterium tuberculosis (mtb) strains. proteins involved in cholesterol import and metabolism have recently been discovered as potent targets against tb. fada5, a thiolase from mtb, is catalyzing the last step of the β-oxidation reaction of the cholesterol side-chain degradation ...201425482540
fada5 a thiolase from mycobacterium tuberculosis: a steroid-binding pocket reveals the potential for drug development against tuberculosis.with the exception of hiv, tuberculosis (tb) is the leading cause of mortality among infectious diseases. the urgent need to develop new antitubercular drugs is apparent due to the increasing number of drug-resistant mycobacterium tuberculosis (mtb) strains. proteins involved in cholesterol import and metabolism have recently been discovered as potent targets against tb. fada5, a thiolase from mtb, is catalyzing the last step of the β-oxidation reaction of the cholesterol side-chain degradation ...201425482540
essential structural and functional roles of the cmr4 subunit in rna cleavage by the cmr crispr-cas complex.the cmr complex is the multisubunit effector complex of the type iii-b clustered regularly interspaced short palindromic repeats (crispr)-cas immune system. the cmr complex recognizes a target rna through base pairing with the integral crispr rna (crrna) and cleaves the target at multiple regularly spaced locations within the complementary region. to understand the molecular basis of the function of this complex, we have assembled information from electron microscopic and x-ray crystallographic ...201425482566
a novel mechanism of protein thermostability: a unique n-terminal domain confers heat resistance to fe/mn-sods.superoxide dismutases (sods), especially thermostable sods, are widely applied in medical treatments, cosmetics, food, agriculture, and other industries given their excellent antioxidant properties. a novel thermostable cambialistic sod from geobacillus thermodenitrificans ng80-2 exhibits maximum activity at 70 °c and high thermostability over a broad range of temperatures (20-80 °c). unlike other reported sods, this enzyme contains an extra repeat-containing n-terminal domain (ntd) of 244 resid ...201425445927
a new window into the molecular physiology of membrane proteins.integral membrane proteins comprise ∼25% of the human proteome. yet, our understanding of their molecular physiology is still in its infancy. this can be attributed to two factors: the experimental challenges that arise from the difficult chemical nature of membrane proteins, and the unclear relationship between their activity and their native environment. new approaches are therefore required to address these challenges. recent developments in mass spectrometry have shown that it is possible to ...201425630257
a new window into the molecular physiology of membrane proteins.integral membrane proteins comprise ∼25% of the human proteome. yet, our understanding of their molecular physiology is still in its infancy. this can be attributed to two factors: the experimental challenges that arise from the difficult chemical nature of membrane proteins, and the unclear relationship between their activity and their native environment. new approaches are therefore required to address these challenges. recent developments in mass spectrometry have shown that it is possible to ...201425630257
interface matters: the stiffness route to stability of a thermophilic tetrameric malate dehydrogenase.in this work we investigate by computational means the behavior of two orthologous bacterial proteins, a mesophilic and a thermophilic tetrameric malate dehydrogenase (maldh), at different temperatures. namely, we quantify how protein mechanical rigidity at different length- and time-scales correlates to protein thermophilicity as commonly believed. in particular by using a clustering analysis strategy to explore the conformational space of the folded proteins, we show that at ambient conditions ...201425437494
substrate-specific development of thermophilic bacterial consortia by using chemically pretreated switchgrass.microbial communities that deconstruct plant biomass have broad relevance in biofuel production and global carbon cycling. biomass pretreatments reduce plant biomass recalcitrance for increased efficiency of enzymatic hydrolysis. we exploited these chemical pretreatments to study how thermophilic bacterial consortia adapt to deconstruct switchgrass (sg) biomass of various compositions. microbial communities were adapted to untreated, ammonium fiber expansion (afex)-pretreated, and ionic-liquid ( ...201425261509
structural insights into translational recoding by frameshift suppressor trnasufj.the three-nucleotide mrna reading frame is tightly regulated during translation to ensure accurate protein expression. translation errors that lead to aberrant protein production can result from the uncoupled movement of the trna in either the 5' or 3' direction on mrna. here, we report the biochemical and structural characterization of +1 frameshift suppressor trna(sufj), a trna known to decode four, instead of three, nucleotides. frameshift suppressor trna(sufj) contains an insertion 5' to its ...201425352689
protein acetylation affects acetate metabolism, motility and acid stress response in escherichia coli.although protein acetylation is widely observed, it has been associated with few specific regulatory functions making it poorly understood. to interrogate its functionality, we analyzed the acetylome in escherichia coli knockout mutants of cobb, the only known sirtuin-like deacetylase, and patz, the best-known protein acetyltransferase. for four growth conditions, more than 2,000 unique acetylated peptides, belonging to 809 proteins, were identified and differentially quantified. nearly 65% of t ...201425518064
double-sieving-defective aminoacyl-trna synthetase causes protein mistranslation and affects cellular physiology and development.aminoacyl-trna synthetases (aarss) constitute a family of ubiquitously expressed essential enzymes that ligate amino acids to their cognate trnas for protein synthesis. recently, aars mutations have been linked to various human diseases; however, how these mutations lead to diseases has remained unclear. in order to address the importance of aminoacylation fidelity in multicellular organisms, we generated an amino-acid double-sieving model in drosophila melanogaster using phenylalanyl-trna synth ...201425427601
structure and transport mechanism of the sodium/proton antiporter mjnhap1.sodium/proton antiporters are essential for sodium and ph homeostasis and play a major role in human health and disease. we determined the structures of the archaeal sodium/proton antiporter mjnhap1 in two complementary states. the inward-open state was obtained by x-ray crystallography in the presence of sodium at ph 8, where the transporter is highly active. the outward-open state was obtained by electron crystallography without sodium at ph 4, where mjnhap1 is inactive. comparison of both str ...201425426803
structure and substrate ion binding in the sodium/proton antiporter panhap.sodium/proton antiporters maintain intracellular ph and sodium levels. detailed structures of antiporters with bound substrate ions are essential for understanding how they work. we have resolved the substrate ion in the dimeric, electroneutral sodium/proton antiporter panhap from pyrococcus abyssi at 3.2 å, and have determined its structure in two different conformations at ph 8 and ph 4. the ion is coordinated by three acidic sidechains, a water molecule, a serine and a main-chain carbonyl in ...201425426802
structural principles of crispr rna processing.the cas6 superfamily, the cas5d subclass, and the host rnase iii endoribonucleases are responsible for producing small rnas (crrna) that function in the crispr-cas immunity. the three enzymes may also interact with the crrna-associated nucleic acid interference complexes. recent development in structural biology of cas6 and cas5d and their complexes with rna substrates has lent new insights on principles of crrna processing and the structural basis for linking crrna processing to interference. b ...201425435327
structural principles of crispr rna processing.the cas6 superfamily, the cas5d subclass, and the host rnase iii endoribonucleases are responsible for producing small rnas (crrna) that function in the crispr-cas immunity. the three enzymes may also interact with the crrna-associated nucleic acid interference complexes. recent development in structural biology of cas6 and cas5d and their complexes with rna substrates has lent new insights on principles of crrna processing and the structural basis for linking crrna processing to interference. b ...201425435327
structure of putrescine aminotransferase from escherichia coli provides insights into the substrate specificity among class iii aminotransferases.ygjg is a putrescine aminotransferase enzyme that transfers amino groups from compounds with terminal primary amines to compounds with an aldehyde group using pyridoxal-5'-phosphate (plp) as a cofactor. previous biochemical data show that the enzyme prefers primary diamines, such as putrescine, over ornithine as a substrate. to better understand the enzyme's substrate specificity, crystal structures of ygjg from escherichia coli were determined at 2.3 and 2.1 å resolutions for the free and putre ...201425423189
tangled web of interactions among proteins involved in iron-sulfur cluster assembly as unraveled by nmr, saxs, chemical crosslinking, and functional studies.proteins containing iron-sulfur (fe-s) clusters arose early in evolution and are essential to life. organisms have evolved machinery consisting of specialized proteins that operate together to assemble fe-s clusters efficiently so as to minimize cellular exposure to their toxic constituents: iron and sulfide ions. to date, the best studied system is the iron-sulfur cluster (isc) operon of escherichia coli, and the eight isc proteins it encodes. our investigations over the past five years have id ...201425450980
tangled web of interactions among proteins involved in iron-sulfur cluster assembly as unraveled by nmr, saxs, chemical crosslinking, and functional studies.proteins containing iron-sulfur (fe-s) clusters arose early in evolution and are essential to life. organisms have evolved machinery consisting of specialized proteins that operate together to assemble fe-s clusters efficiently so as to minimize cellular exposure to their toxic constituents: iron and sulfide ions. to date, the best studied system is the iron-sulfur cluster (isc) operon of escherichia coli, and the eight isc proteins it encodes. our investigations over the past five years have id ...201425450980
recq helicase and recj nuclease provide complementary functions to resect dna for homologous recombination.recombinational dna repair by the recf pathway of escherichia coli requires the coordinated activities of reca, recfor, recq, recj, and single-strand dna binding (ssb) proteins. these proteins facilitate formation of homologously paired joint molecules between linear double-stranded (dsdna) and supercoiled dna. repair starts with resection of the broken dsdna by recq, a 3'→5' helicase, recj, a 5'→3' exonuclease, and ssb protein. the ends of a dsdna break can be blunt-ended, or they may possess e ...201425411316
argonaute piwi domain and microrna duplex structure regulate small rna sorting in arabidopsis.small rnas (srnas) are loaded into argonaute (ago) proteins to induce gene silencing. in plants, the 5'-terminal nucleotide is important for srna sorting into different agos. here we show that microrna (mirna) duplex structure also contributes to mirna sorting. base pairing at the 15th nucleotide of a mirna duplex is important for mirna sorting in both arabidopsis ago1 and ago2. ago2 favours mirna duplexes with no middle mismatches, whereas ago1 tolerates, or prefers, duplexes with central misma ...201425406978
structural and functional characterization of a ketosteroid transcriptional regulator of mycobacterium tuberculosis.catabolism of host cholesterol is critical to the virulence of mycobacterium tuberculosis and is a potential target for novel therapeutics. kstr2, a tetr family repressor (tfr), regulates the expression of 15 genes encoding enzymes that catabolize the last half of the cholesterol molecule, represented by 3aα-h-4α(3'-propanoate)-7aβ-methylhexahydro-1,5-indane-dione (hip). binding of kstr2 to its operator sequences is relieved upon binding of hip-coa. a 1.6-å resolution crystal structure of the ks ...201425406313
structural and functional characterization of a ketosteroid transcriptional regulator of mycobacterium tuberculosis.catabolism of host cholesterol is critical to the virulence of mycobacterium tuberculosis and is a potential target for novel therapeutics. kstr2, a tetr family repressor (tfr), regulates the expression of 15 genes encoding enzymes that catabolize the last half of the cholesterol molecule, represented by 3aα-h-4α(3'-propanoate)-7aβ-methylhexahydro-1,5-indane-dione (hip). binding of kstr2 to its operator sequences is relieved upon binding of hip-coa. a 1.6-å resolution crystal structure of the ks ...201425406313
movement of elongation factor g between compact and extended conformations.previous structural studies suggested that ribosomal translocation is accompanied by large interdomain rearrangements of elongation factor g (ef-g). here, we follow the movement of domain iv of ef-g relative to domain ii of ef-g using ensemble and single-molecule förster resonance energy transfer. our results indicate that ribosome-free ef-g predominantly adopts a compact conformation that can also, albeit infrequently, transition into a more extended conformation in which domain iv moves away f ...201425463439
movement of elongation factor g between compact and extended conformations.previous structural studies suggested that ribosomal translocation is accompanied by large interdomain rearrangements of elongation factor g (ef-g). here, we follow the movement of domain iv of ef-g relative to domain ii of ef-g using ensemble and single-molecule förster resonance energy transfer. our results indicate that ribosome-free ef-g predominantly adopts a compact conformation that can also, albeit infrequently, transition into a more extended conformation in which domain iv moves away f ...201425463439
viruses of haloarchaea.in hypersaline environments, haloarchaea (halophilic members of the archaea) are the dominant organisms, and the viruses that infect them, haloarchaeoviruses are at least ten times more abundant. since their discovery in 1974, described haloarchaeoviruses include head-tailed, pleomorphic, spherical and spindle-shaped morphologies, representing myoviridae, siphoviridae, podoviridae, pleolipoviridae, sphaerolipoviridae and fuselloviridae families. this review overviews current knowledge of haloarc ...201425402735
phylogenetically driven sequencing of extremely halophilic archaea reveals strategies for static and dynamic osmo-response.organisms across the tree of life use a variety of mechanisms to respond to stress-inducing fluctuations in osmotic conditions. cellular response mechanisms and phenotypes associated with osmoadaptation also play important roles in bacterial virulence, human health, agricultural production and many other biological systems. to improve understanding of osmoadaptive strategies, we have generated 59 high-quality draft genomes for the haloarchaea (a euryarchaeal clade whose members thrive in hypersa ...201425393412
structural insight into amino group-carrier protein-mediated lysine biosynthesis: crystal structure of the lysz·lysw complex from thermus thermophilus.in the biosynthesis of lysine by thermus thermophilus, the metabolite α-ketoglutarate is converted to the intermediate α-aminoadipate (aaa), which is protected by the 54-amino acid acidic protein lysw. in this study, we determined the crystal structure of lysz from t. thermophilus (ttlysz), an amino acid kinase that catalyzes the second step in the aaa to lysine conversion, which was in a complex with lysw at a resolution of 1.85 å. a crystal analysis coupled with isothermal titration calorimetr ...201425392000
structural insight into amino group-carrier protein-mediated lysine biosynthesis: crystal structure of the lysz·lysw complex from thermus thermophilus.in the biosynthesis of lysine by thermus thermophilus, the metabolite α-ketoglutarate is converted to the intermediate α-aminoadipate (aaa), which is protected by the 54-amino acid acidic protein lysw. in this study, we determined the crystal structure of lysz from t. thermophilus (ttlysz), an amino acid kinase that catalyzes the second step in the aaa to lysine conversion, which was in a complex with lysw at a resolution of 1.85 å. a crystal analysis coupled with isothermal titration calorimetr ...201425392000
cecafdb: a curated database for the documentation, visualization and comparative analysis of central carbon metabolic flux distributions explored by 13c-fluxomics.the central carbon metabolic flux database (cecafdb, available at http://www.cecafdb.org) is a manually curated, multipurpose and open-access database for the documentation, visualization and comparative analysis of the quantitative flux results of central carbon metabolism among microbes and animal cells. it encompasses records for more than 500 flux distributions among 36 organisms and includes information regarding the genotype, culture medium, growth conditions and other specific information ...201425392417
cecafdb: a curated database for the documentation, visualization and comparative analysis of central carbon metabolic flux distributions explored by 13c-fluxomics.the central carbon metabolic flux database (cecafdb, available at http://www.cecafdb.org) is a manually curated, multipurpose and open-access database for the documentation, visualization and comparative analysis of the quantitative flux results of central carbon metabolism among microbes and animal cells. it encompasses records for more than 500 flux distributions among 36 organisms and includes information regarding the genotype, culture medium, growth conditions and other specific information ...201425392417
thermodynamic system drift in protein evolution.proteins from thermophiles are generally more thermostable than their mesophilic homologs, but little is known about the evolutionary process driving these differences. here we attempt to understand how the diverse thermostabilities of bacterial ribonuclease h1 (rnh) proteins evolved. rnh proteins from thermus thermophilus (ttrnh) and escherichia coli (ecrnh) share similar structures but differ in melting temperature (t(m)) by 20 °c. ttrnh's greater stability is caused in part by the presence of ...201425386647
interacting cytoplasmic loops of subunits a and c of escherichia coli f1f0 atp synthase gate h+ transport to the cytoplasm.h(+)-transporting f1f0 atp synthase catalyzes the synthesis of atp via coupled rotary motors within f0 and f1. h(+) transport at the subunit a-c interface in transmembranous f0 drives rotation of a cylindrical c10 oligomer within the membrane, which is coupled to rotation of subunit γ within the α3β3 sector of f1 to mechanically drive atp synthesis. f1f0 functions in a reversible manner, with atp hydrolysis driving h(+) transport. atp-driven h(+) transport in a select group of cysteine mutants i ...201425385585
phenotyping the quality of complex medium components by simple online-monitored shake flask experiments.media containing yeast extracts and other complex raw materials are widely used for the cultivation of microorganisms. however, variations in the specific nutrient composition can occur, due to differences in the complex raw material ingredients and in the production of these components. these lot-to-lot variations can affect growth rate, product yield and product quality in laboratory investigations and biopharmaceutical production processes. in the fda's process analytical technology (pat) ini ...201425376163
the σ enigma: bacterial σ factors, archaeal tfb and eukaryotic tfiib are homologs.structural comparisons of initiating rna polymerase complexes and structure-based amino acid sequence alignments of general transcription initiation factors (eukaryotic tfiib, archaeal tfb and bacterial σ factors) show that these proteins are homologs. tfiib and tfb each have two-five-helix cyclin-like repeats (clrs) that include a c-terminal helix-turn-helix (hth) motif (clr/hth domains). four homologous hth motifs are present in bacterial σ factors that are relics of clr/hth domains. sequence ...201425483602
rna targeting by the type iii-a crispr-cas csm complex of thermus thermophilus.crispr-cas is a prokaryotic adaptive immune system that provides sequence-specific defense against foreign nucleic acids. here we report the structure and function of the effector complex of the type iii-a crispr-cas system of thermus thermophilus: the csm complex (ttcsm). ttcsm is composed of five different protein subunits (csm1-csm5) with an uneven stoichiometry and a single crrna of variable size (35-53 nt). the ttcsm crrna content is similar to the type iii-b cmr complex, indicating that cr ...201425457165
silencing by h-ns potentiated the evolution of salmonella.the bacterial h-ns protein silences expression from sequences with higher at-content than the host genome and is believed to buffer the fitness consequences associated with foreign gene acquisition. loss of h-ns results in severe growth defects in salmonella, but the underlying reasons were unclear. an experimental evolution approach was employed to determine which secondary mutations could compensate for the loss of h-ns in salmonella. six independently derived s. typhimurium hns mutant strains ...201425375226
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