Publications

reconstitution of translation from thermus thermophilus reveals a minimal set of components sufficient for protein synthesis at high temperatures and functional conservation of modern and ancient translation components.	thermus thermophilus is a thermophilic model organism distantly related to the mesophilic model organism e. coli. we reconstituted protein translation of thermus thermophilus in vitro from purified ribosomes, transfer ribonucleic acids (trnas) and 33 recombinant proteins. this reconstituted system was fully functional, capable of translating natural messenger rna (mrna) into active full-length proteins at temperatures up to 65°c and with yields up to 60 μg/ml. surprisingly, the synthesis of acti ...	2012	22723376
reverse structural genomics: an unusual flavin-binding site in a putative protease from bacteroides thetaiotaomicron.	the structure of a putative protease from bacteroides thetaiotaomicron features an unprecedented binding site for flavin mononucleotide. the flavin isoalloxazine ring is sandwiched between two tryptophan residues in the interface of the dimeric protein. we characterized the recombinant protein with regard to its affinity for naturally occurring flavin derivatives and several chemically modified flavin analogs. dissociation constants were determined by isothermal titration calorimetry. the protei ...	2012	22718753
structural constraints identified with covariation analysis in ribosomal rna.	covariation analysis is used to identify those positions with similar patterns of sequence variation in an alignment of rna sequences. these constraints on the evolution of two positions are usually associated with a base pair in a helix. while mutual information (mi) has been used to accurately predict an rna secondary structure and a few of its tertiary interactions, early studies revealed that phylogenetic event counting methods are more sensitive and provide extra confidence in the predictio ...	2012	22724009
identification and characterization of the thermus thermophilus 5-methylcytidine (m5c) methyltransferase modifying 23 s ribosomal rna (rrna) base c1942.	methylation of cytidines at carbon-5 is a common posttranscriptional rna modification encountered across all domains of life. here, we characterize the modifications of c1942 and c1962 in thermus thermophilus 23 s rrna as 5-methylcytidines (m(5)c) and identify the two associated methyltransferases. the methyltransferase modifying c1942, named rlmo, has not been characterized previously. rlmo modifies naked 23 s rrna, but not the assembled 50 s subunit or 70 s ribosomes. the x-ray crystal structu ...	2012	22711535
structure and activity of the pseudomonas aeruginosa hotdog-fold thioesterases pa5202 and pa2801.	the hotdog fold is one of the basic protein folds widely present in bacteria, archaea and eukaryotes. many of these proteins exhibit thioesterase activity against fatty acyl-coas and play important roles in lipid metabolism, cellular signalling and degradation of xenobiotics. the genome of the opportunistic pathogen pseudomonas aeruginosa contains over 20 genes encoding predicted hotdog-fold proteins, none of which have been experimentally characterized. we have found that two p. aeruginosa hotd ...	2012	22439787
dissociation of antibacterial activity and aminoglycoside ototoxicity in the 4-monosubstituted 2-deoxystreptamine apramycin.	aminoglycosides are potent antibacterials, but therapy is compromised by substantial toxicity causing, in particular, irreversible hearing loss. aminoglycoside ototoxicity occurs both in a sporadic dose-dependent and in a genetically predisposed fashion. we recently have developed a mechanistic concept that postulates a key role for the mitochondrial ribosome (mitoribosome) in aminoglycoside ototoxicity. we now report on the surprising finding that apramycin, a structurally unique aminoglycoside ...	2012	22699498
polymorphisms in the mitochondrial ribosome recycling factor ef-g2mt/mef2 compromise cell respiratory function and increase atorvastatin toxicity.	mitochondrial translation, essential for synthesis of the electron transport chain complexes in the mitochondria, is governed by nuclear encoded genes. polymorphisms within these genes are increasingly being implicated in disease and may also trigger adverse drug reactions. statins, a class of hmg-coa reductase inhibitors used to treat hypercholesterolemia, are among the most widely prescribed drugs in the world. however, a significant proportion of users suffer side effects of varying severity ...	2012	22719265
molecular dynamics of a thermostable multicopper oxidase from thermus thermophilus hb27: structural differences between the apo and holo forms.	molecular dynamic (md) simulations have been performed on tth-mco, a hyperthermophilic multicopper oxidase from thermus thermophilus hb27, in the apo as well as the holo form, with the aim of exploring the structural dynamic properties common to the two conformational states. according to structural comparison between this enzyme and other mcos, the substrate in process to electron transfer in an outer-sphere event seems to transiently occupy a shallow and overall hydrophobic cavity near the cu ...	2012	22808237
a lon-like protease with no atp-powered unfolding activity.	lon proteases are a family of atp-dependent proteases involved in protein quality control, with a unique proteolytic domain and an aaa(+) (atpases associated with various cellular activities) module accommodated within a single polypeptide chain. they were classified into two types as either the ubiquitous soluble lona or membrane-inserted archaeal lonb. in addition to the energy-dependent forms, a number of medically and ecologically important groups of bacteria encode a third type of lon-like ...	2012	22792246
mechanism of elongation factor-g-mediated fusidic acid resistance and fitness compensation in staphylococcus aureus.	antibiotic resistance in bacteria is often associated with fitness loss, which is compensated by secondary mutations. fusidic acid (fa), an antibiotic used against pathogenic bacteria staphylococcus aureus, locks elongation factor-g (ef-g) to the ribosome after gtp hydrolysis. to clarify the mechanism of fitness loss and compensation in relation to fa resistance, we have characterized three s. aureus ef-g mutants with fast kinetics and crystal structures. our results show that a significantly sl ...	2012	22767604
oxidants and not alkylating agents induce rapid mtdna loss and mitochondrial dysfunction.	mitochondrial dna (mtdna) is essential for proper mitochondrial function and encodes 22 trnas, 2 rrnas and 13 polypeptides that make up subunits of complex i, iii, iv, in the electron transport chain and complex v, the atp synthase. although mitochondrial dysfunction has been implicated in processes such as premature aging, neurodegeneration, and cancer, it has not been shown whether persistent mtdna damage causes a loss of oxidative phosphorylation. we addressed this question by treating mouse ...	2012	22766155
the scoi homologue senc is a copper binding protein that interacts directly with the cbb₃-type cytochrome oxidase in rhodobacter capsulatus.	sco proteins are widespread assembly factors for the cu(a) centre of aa₃-type cytochrome oxidases in eukaryotic and prokaryotic organisms. however, sco homologues are also found in bacteria like rhodobacter capsulatus which lack aa₃-type cytochrome oxidases and instead use a cbb₃-type cytochrome oxidase (cbb₃ cox) without a cu(a) centre as a terminal oxidase. in the current study, we have analyzed the role of sco (senc) during cbb₃ cox assembly in r. capsulatus. in agreement with earlier works, ...	2012	22771512
stoichiometry of proton pumping by the cbb3-type oxygen reductase in whole cells of rhodobacter capsulatus at ph 7 is about 0.5 h+ per electron.		2012	22761318
arrangement of subunits in intact mammalian mitochondrial atp synthase determined by cryo-em.	mitochondrial atp synthase is responsible for the synthesis of atp, a universal energy currency in cells. whereas x-ray crystallography has revealed the structure of the soluble region of the complex and the membrane-intrinsic c-subunits, little is known about the structure of the six other proteins (a, b, f, a6l, e, and g) that comprise the membrane-bound region of the complex in animal mitochondria. here, we present the structure of intact bovine mitochondrial atp synthase at ∼18 å resolution ...	2012	22753497
the elongation, termination, and recycling phases of translation in eukaryotes.	this work summarizes our current understanding of the elongation and termination/recycling phases of eukaryotic protein synthesis. we focus here on recent advances in the field. in addition to an overview of translation elongation, we discuss unique aspects of eukaryotic translation elongation including eef1 recycling, eef2 modification, and eef3 and eif5a function. likewise, we highlight the function of the eukaryotic release factors erf1 and erf3 in translation termination, and the functions o ...	2012	22751155
single-molecule analysis of inhibitory pausing states of v1-atpase.	v(1)-atpase, the hydrophilic v-atpase domain, is a rotary motor fueled by atp hydrolysis. here, we found that thermus thermophilus v(1)-atpase shows two types of inhibitory pauses interrupting continuous rotation: a short pause (sp, 4.2 s) that occurred frequently during rotation, and a long inhibitory pause (lp, >30 min) that terminated all active rotations. both pauses occurred at the same angle for atp binding and hydrolysis. kinetic analysis revealed that the time constants of inactivation i ...	2012	22736762
structural study on the architecture of the bacterial atp synthase fo motor.	we purified the f(o) complex from the ilyobacter tartaricus na(+)-translocating f(1)f(o)-atp synthase and performed a biochemical and structural study. laser-induced liquid bead ion desorption ms analysis demonstrates that all three subunits of the isolated f(o) complex were present and in native stoichiometry (ab(2)c(11)). cryoelectron microscopy of 2d crystals yielded a projection map at a resolution of 7.0 å showing electron densities from the c(11) rotor ring and up to seven adjacent helices ...	2012	22736796
control of electron transport routes through redox-regulated redistribution of respiratory complexes.	in cyanobacteria, respiratory electron transport takes place in close proximity to photosynthetic electron transport, because the complexes required for both processes are located within the thylakoid membranes. the balance of electron transport routes is crucial for cell physiology, yet the factors that control the predominance of particular pathways are poorly understood. here we use a combination of tagging with green fluorescent protein and confocal fluorescence microscopy in live cells of t ...	2012	22733774
structure of the signal transduction protein trap (target of rnaiii-activating protein).	the crystal structure of the signal transduction protein trap is reported at 1.85 å resolution. the structure of trap consists of a central eight-stranded β-barrel flanked asymmetrically by helices and is monomeric both in solution and in the crystal structure. a formate ion was found bound to trap identically in all four molecules in the asymmetric unit.	2012	22750855
the fragmented mitochondrial ribosomal rnas of plasmodium falciparum.	the mitochondrial genome in the human malaria parasite plasmodium falciparum is most unusual. over half the genome is composed of the genes for three classic mitochondrial proteins: cytochrome oxidase subunits i and iii and apocytochrome b. the remainder encodes numerous small rnas, ranging in size from 23 to 190 nt. previous analysis revealed that some of these transcripts have significant sequence identity with highly conserved regions of large and small subunit rrnas, and can form the expecte ...	2012	22761677
thermodynamic characterization of rna 2 × 3 nucleotide internal loops.	to better elucidate rna structure-function relationships and to improve the design of pharmaceutical agents that target specific rna motifs, an understanding of rna primary, secondary, and tertiary structure is necessary. the prediction of rna secondary structure from sequence is an intermediate step in predicting rna three-dimensional structure. rna secondary structure is typically predicted using a nearest neighbor model based on free energy parameters. the current free energy parameters for 2 ...	2012	22720720
thermostable dna polymerase from a viral metagenome is a potent rt-pcr enzyme.	viral metagenomic libraries are a promising but previously untapped source of new reagent enzymes. deep sequencing and functional screening of viral metagenomic dna from a near-boiling thermal pool identified clones expressing thermostable dna polymerase (pol) activity. among these, 3173 pol demonstrated both high thermostability and innate reverse transcriptase (rt) activity. we describe the biochemistry of 3173 pol and report its use in single-enzyme reverse transcription pcr (rt-pcr). wild-ty ...	2012	22675552
mass spectrometry--from peripheral proteins to membrane motors.	that membrane protein complexes could survive in the gas phase had always seemed impossible. the lack of chargeable residues, high hydrophobicity, and poor solubility and the vast excess of detergent contributed to the view that it would not be possible to obtain mass spectra of intact membrane complexes. with the recent success in recording mass spectra of these complexes, first from recombinant sources and later from the cellular environment, many surprising properties of these gas phase membr ...	2012	22750574
molecular evolution of dihydrouridine synthases.	dihydrouridine (d) is a modified base found in conserved positions in the d-loop of trna in bacteria, eukaryota, and some archaea. despite the abundant occurrence of d, little is known about its biochemical roles in mediating trna function. it is assumed that d may destabilize the structure of trna and thus enhance its conformational flexibility. d is generated post-transcriptionally by the reduction of the 5,6-double bond of a uridine residue in rna transcripts. the reaction is carried out by d ...	2012	22741570
dnak chaperone-dependent disaggregation by caseinolytic peptidase b (clpb) mutants reveals functional overlap in the n-terminal domain and nucleotide-binding domain-1 pore tyrosine.	protein disaggregation in escherichia coli is carried out by clpb, an aaa(+) (atpases associated with various cellular activities) molecular chaperone, together with the dnak chaperone system. conformational changes in clpb driven by atp binding and hydrolysis promote substrate binding, unfolding, and translocation. conserved pore tyrosines in both nucleotide-binding domain-1 (nbd-1) and -2 (nbd-2), which reside in flexible loops extending into the central pore of the clpb hexamer, bind substrat ...	2012	22745126
characterization of a conserved interaction between dna glycosylase and para in mycobacterium smegmatis and m. tuberculosis.	the chromosome partitioning proteins, parab, ensure accurate segregation of genetic materials into daughter cells and most bacterial species contain their homologs. however, little is known about the regulation of parab proteins. in this study, we found that 3-methyladenine dna glycosylase i mstag(ms5082) regulates bacterial growth and cell morphology by directly interacting with mspara (ms6939) and inhibiting its atpase activity in mycobacterium smegmatis. using bacterial two-hybrid and pull-do ...	2012	22675536
the c-terminal domain of 4-hydroxyphenylacetate 3-hydroxylase from acinetobacter baumannii is an autoinhibitory domain.	p-hydroxyphenylacetate (hpa) 3-hydroxylase from acinetobacter baumannii consists of a reductase component (c(1)) and an oxygenase component (c(2)). c(1) catalyzes the reduction of fmn by nadh to provide fmnh(-) as a substrate for c(2). the rate of reduction of flavin is enhanced ∼20-fold by binding hpa. the n-terminal domain of c(1) is homologous to other flavin reductases, whereas the c-terminal domain (residues 192-315) is similar to marr, a repressor protein involved in bacterial antibiotic r ...	2012	22661720
evaluation of biomass production, carotenoid level and antioxidant capacity produced by thermus filiformis using fractional factorial design.	a fractional factorial design 2(5-1) was used to evaluate the effect of temperature, ph, and concentrations of yeast extract, tryptone and nitsch's trace elements on the biomass, total carotenoids and protection against singlet oxygen by carotenoid extracts of the bacterium thermus filiformis. in addition, the carotenoid composition was determined by high-performance liquid chromatography connected to a diode array and mass spectrometer detectors (hplc-dad-ms/ms). the production of biomass range ...	2012	24031811
corallopyronin a specifically targets and depletes essential obligate wolbachia endobacteria from filarial nematodes in vivo.	doxycycline and rifampicin deplete essential wolbachia from filarial nematodes that cause lymphatic filariasis or onchocerciasis, resulting in blocked worm development and death. however, doxycycline is contraindicated for children and pregnant/breastfeeding women, as is rifampicin in the latter group with the additional specter of possible resistance development in mycobacterium spp. novel antibiotics with a narrower spectrum would aid in eliminating filarial diseases. corallococcus coralloides ...	2012	22586066
reconstitution of vacuolar-type rotary h+-atpase/synthase from thermus thermophilus.	vacuolar-type rotary h(+)-atpase/synthase (v(o)v(1)) from thermus thermophilus, composed of nine subunits, a, b, d, f, c, e, g, i, and l, has been reconstituted from individually isolated v(1) (a(3)b(3)d(1)f(1)) and v(o) (c(1)e(2)g(2)i(1)l(12)) subcomplexes in vitro. a(3)b(3)d and a(3)b(3) also reconstituted with v(o), resulting in a holoenzyme-like complexes. however, a(3)b(3)d-v(o) and a(3)b(3)-v(o) did not show atp synthesis and dicyclohexylcarbodiimide-sensitive atpase activity. the reconsti ...	2012	22582389
codon usage bias in prokaryotic pyrimidine-ending codons is associated with the degeneracy of the encoded amino acids.	synonymous codons are unevenly distributed among genes, a phenomenon termed codon usage bias. understanding the patterns of codon bias and the forces shaping them is a major step towards elucidating the adaptive advantage codon choice can confer at the level of individual genes and organisms. here, we perform a large-scale analysis to assess codon usage bias pattern of pyrimidine-ending codons in highly expressed genes in prokaryotes. we find a bias pattern linked to the degeneracy of the encode ...	2012	22581775
glucose(xylose) isomerase production by streptomyces sp. ch7 grown on agricultural residues.	streptomyces sp. ch7 was found to efficiently produce glucose(xylose) isomerase when grown on either xylan or agricultural residues. this strain produced a glucose(xylose) isomerase activity of roughly 1.8 u/mg of protein when it was grown in medium containing 1% xylose as a carbon source. maximal enzymatic activities of about 5 and 3 u/mg were obtained when 1% xylan and 2.5% corn husks were used, respectively. the enzyme was purified from a mycelial extract to 16-fold purity with only two conse ...	2012	24031932
deep sequencing reveals minor protease resistance mutations in patients failing a protease inhibitor regimen.	standard genotypic antiretroviral resistance testing, performed by bulk sequencing, does not readily detect variants that comprise <20% of the circulating hiv-1 rna population. nevertheless, it is valuable in selecting an antiretroviral regimen after antiretroviral failure. in patients with poor adherence, resistant variants may not reach this threshold. therefore, deep sequencing would be potentially valuable for detecting minority resistant variants. we compared bulk sequencing and deep sequen ...	2012	22457522
substrate-induced change in the quaternary structure of type 2 isopentenyl diphosphate isomerase from sulfolobus shibatae.	type 2 isopentenyl diphosphate isomerase catalyzes the interconversion between two active units for isoprenoid biosynthesis, i.e., isopentenyl diphosphate and dimethylallyl diphosphate, in almost all archaea and in some bacteria, including human pathogens. the enzyme is a good target for discovery of antibiotics because it is essential for the organisms that use only the mevalonate pathway to produce the active isoprene units and because humans possess a nonhomologous isozyme, type 1 isopentenyl ...	2012	22505674
life under multiple extreme conditions: diversity and physiology of the halophilic alkalithermophiles.	around the world, there are numerous alkaline, hypersaline environments that are heated either geothermally or through intense solar radiation. it was once thought that such harsh environments were inhospitable and incapable of supporting a variety of life. however, numerous culture-dependent and -independent studies revealed the presence of an extensive diversity of aerobic and anaerobic bacteria and archaea that survive and grow under these multiple harsh conditions. this diversity includes th ...	2012	22492435
site-directed mutagenesis of a family 42 β-galactosidase from an antarctic bacterium.	site directed mutagenesis was used to modify the active site of a cold active beta-galactosidase taken from an antarctic psychrotolerant planococcus bacterial isolate. the goal was to modify the active site such that there would be an increase in activity on certain substrates which showed little to no activity with the wild type enzyme. a total of 5 mutant enzymes were constructed with amino acid changes based on an analysis done via homology modeling. all 5 modified enzymes were assayed using ...	2012	22773960
rna in defense: crisprs protect prokaryotes against mobile genetic elements.	the crispr/cas system in prokaryotes provides resistance against invading viruses and plasmids. three distinct stages in the mechanism can be recognized. initially, fragments of invader dna are integrated as new spacers into the repetitive crispr locus. subsequently, the crispr is transcribed and the transcript is cleaved by a cas protein within the repeats, generating short rnas (crrnas) that contain the spacer sequence. finally, crrnas guide the cas protein machinery to a complementary invader ...	2012	21441598
structure and folding of a rare, natural kink turn in rna with an aa pair at the 2b2n position.	the kink turn (k-turn) is a frequently occurring motif, comprising a bulge followed by g•a and a•g pairs that introduces a sharp axial bend in duplex rna. natural k-turn sequences exhibit significant departures from the consensus, including the a•g pairs that form critical interactions stabilizing the core of the structure. kt-23 found in the small ribosomal subunit differs from the consensus in many organisms, particularly in the second a•g pair distal to the bulge (2b•2n). analysis of many kt- ...	2012	22539525
production of l-ribose from l-ribulose by a triple-site variant of mannose-6-phosphate isomerase from geobacillus thermodenitrificans.	a triple-site variant (w17q n90a l129f) of mannose-6-phosphate isomerase from geobacillus thermodenitrificans was obtained by combining variants with residue substitutions at different positions after random and site-directed mutagenesis. the specific activity and catalytic efficiency (k(cat)/k(m)) for l-ribulose isomerization of this variant were 3.1- and 7.1-fold higher, respectively, than those of the wild-type enzyme at ph 7.0 and 70°c in the presence of 1 mm co(2+). the triple-site variant ...	2012	22447612
a comparison of the crystal structures of eukaryotic and bacterial ssu ribosomal rnas reveals common structural features in the hypervariable regions.	while the majority of the ribosomal rna structure is conserved in the three major domains of life--archaea, bacteria, and eukaryotes, specific regions of the rrna structure are unique to at least one of these three primary forms of life. in particular, the comparative secondary structure for the eukaryotic ssu rrna contains several regions that are different from the analogous regions in the bacteria. our detailed analysis of two recently determined eukaryotic 40s ribosomal crystal structures, t ...	2012	22693601
phage-induced expression of crispr-associated proteins is revealed by shotgun proteomics in streptococcus thermophilus.	the crispr/cas system, comprised of clustered regularly interspaced short palindromic repeats along with their associated (cas) proteins, protects bacteria and archaea from viral predation and invading nucleic acids. while the mechanism of action for this acquired immunity is currently under investigation, the response of cas protein expression to phage infection has yet to be elucidated. in this study, we employed shotgun proteomics to measure the global proteome expression in a model system fo ...	2012	22666452
mobility of xe atoms within the oxygen diffusion channel of cytochrome ba(3) oxidase.	we use a form of "freeze-trap, kinetic crystallography" to explore the migration of xe atoms away from the dinuclear heme a(3)/cu(b) center in thermus thermophilus cytochrome ba(3) oxidase. this enzyme is a member of the heme-copper oxidase superfamily and is thus crucial for dioxygen-dependent life. the mechanisms involved in the migration of oxygen, water, electrons, and protons into and/or out of the specialized channels of the heme-copper oxidases are generally not well understood. pressuriz ...	2012	22607023
flexibility of the metal-binding region in apo-cupredoxins.	protein-mediated electron transfer is an essential event in many biochemical processes. efficient electron transfer requires the reorganization energy of the redox event to be minimized, which is ensured by the presence of rigid donor and acceptor sites. electron transfer copper sites are present in the ubiquitous cupredoxin fold, able to bind one or two copper ions. the low reorganization energy in these metal centers has been accounted for by assuming that the protein scaffold creates an entat ...	2012	22645370
engineering rotor ring stoichiometries in the atp synthase.	atp synthase membrane rotors consist of a ring of c-subunits whose stoichiometry is constant for a given species but variable across different ones. we investigated the importance of c/c-subunit contacts by site-directed mutagenesis of a conserved stretch of glycines (gxgxgxgxg) in a bacterial c(11) ring. structural and biochemical studies show a direct, specific influence on the c-subunit stoichiometry, revealing c(<11), c(12), c(13), c(14), and c(>14) rings. molecular dynamics simulations rati ...	2012	22628564
how hibernation factors rmf, hpf, and yfia turn off protein synthesis.	eubacteria inactivate their ribosomes as 100s dimers or 70s monomers upon entry into stationary phase. in escherichia coli, 100s dimer formation is mediated by ribosome modulation factor (rmf) and hibernation promoting factor (hpf), or alternatively, the yfia protein inactivates ribosomes as 70s monomers. here, we present high-resolution crystal structures of the thermus thermophilus 70s ribosome in complex with each of these stationary-phase factors. the binding site of rmf overlaps with that o ...	2012	22605777
absence of helicobacter pylori high tetracycline resistant 16s rdna aga926-928ttc genotype in gastric biopsy specimens from dyspeptic patients of a city in the interior of são paulo, brazil.	treatment effectiveness of helicobacter pylori varies regionally and is decreasing worldwide, principally as a result of antibiotic resistant bacterium. tetracycline is generally included in second line h. pylori eradication regimens. in brazil, a high level of tetracycline resistance (tetr) is mainly associated with aga926-928ttc 16 s rdna nucleotide substitutions. as h. pylori culture is fastidious, we investigated the primary occurrence of h. pylori 16 s rdna high level tetr genotype using a ...	2012	22594560
structure and conservation of the periplasmic targeting factor tic22 protein from plants and cyanobacteria.	mitochondria and chloroplasts are of endosymbiotic origin. their integration into cells entailed the development of protein translocons, partially by recycling bacterial proteins. we demonstrate the evolutionary conservation of the translocon component tic22 between cyanobacteria and chloroplasts. tic22 in anabaena sp. pcc 7120 is essential. the protein is localized in the thylakoids and in the periplasm and can be functionally replaced by a plant orthologue. tic22 physically interacts with the ...	2012	22593581
crystallization and preliminary x-ray analysis of the reductase component of p-hydroxyphenylacetate 3-hydroxylase from acinetobacter baumannii.	p-hydroxyphenylacetate 3-hydroxylase (hpah) from acinetobacter baumannii catalyzes the hydroxylation of p-hydroxyphenylacetate (hpa) at the ortho position to yield 3,4-dihydroxyphenylacetate (dhpa). hpah from a. baumannii is a two-component flavoprotein consisting of a smaller reductase (c(1)) component and a larger oxygenase (c(2)) component. the c(1) component supplies a reduced flavin in its free form to the c(2) counterpart for hydroxylation. in addition, hpa can bind to c(1) and enhance the ...	2012	22684080
a computational investigation on the connection between dynamics properties of ribosomal proteins and ribosome assembly.	assembly of the ribosome from its protein and rna constituents has been studied extensively over the past 50 years, and experimental evidence suggests that prokaryotic ribosomal proteins undergo conformational changes during assembly. however, to date, no studies have attempted to elucidate these conformational changes. the present work utilizes computational methods to analyze protein dynamics and to investigate the linkage between dynamics and binding of these proteins during the assembly of t ...	2012	22654657
real-time evidence for ef-g-induced dynamics of helix 44 in 16s rrna.	the penultimate stem-loop of 16s ribosomal rna (rrna), helix 44, plays a central role in ribosome function. using time-resolved dimethyl sulfate (dms) probing, we have analyzed time-dependent modifications that occur at specific bases in this helix near the decoding region, resulting from the binding of elongation factor g (ef-g) in various forms. when ef-g-gtp is bound to 70s ribosomes, bases a1492 and a1493 are immediately protected, while other bases in the region show either no change or enh ...	2012	22634282
crystal structure of the largest subunit of a bacterial rna-guided immune complex and its role in dna target binding.	prokaryotes make use of small rnas encoded by crispr (clustered regularly interspaced short palindromic repeat) loci to provide immunity against bacteriophage or plasmid invasion. in escherichia coli, the crispr-associated complex for antiviral defense (cascade) utilizes these rnas to target foreign dna for destruction. casa, the largest subunit of cascade, is essential for its function. here we report the crystal structure of thermus thermophilus casa. the structure is composed of two domains t ...	2012	22621933
a thermophilic ionic liquid-tolerant cellulase cocktail for the production of cellulosic biofuels.	generation of biofuels from sugars in lignocellulosic biomass is a promising alternative to liquid fossil fuels, but efficient and inexpensive bioprocessing configurations must be developed to make this technology commercially viable. one of the major barriers to commercialization is the recalcitrance of plant cell wall polysaccharides to enzymatic hydrolysis. biomass pretreatment with ionic liquids (ils) enables efficient saccharification of biomass, but residual ils inhibit both saccharificati ...	2012	22649505
proline oxidase promotes tumor cell survival in hypoxic tumor microenvironments.	proline is a readily released stress substrate that can be metabolized by proline oxidase (pox) to generate either reactive oxygen species (ros) to induce apoptosis or autophagy or atp during times of nutrient stress. however, the contribution of proline metabolism to tumorigenesis in hypoxic microenvironments has not been explored. in this study, we investigated the different functions of pox under hypoxia and glucose depletion. we found that hypoxia induced pox expression in cancer cells in vi ...	2012	22609800
structural insights into the substrate specificity of streptococcus pneumoniae β(1,3)-galactosidase bgac.	the surface-exposed β-galactosidase bgac from streptococcus pneumoniae was reported to be a virulence factor because of its specific hydrolysis activity toward the β(1,3)-linked galactose and n-acetylglucosamine (galβ(1,3)nag) moiety of oligosaccharides on the host molecules. here we report the crystal structure of bgac at 1.8 å and its complex with galactose at 1.95 å. at ph 5.5-8.0, bgac exists as a stable homodimer, each subunit of which consists of three distinct domains: a catalytic domain ...	2012	22593580
active site opening and closure control translocation of multisubunit rna polymerase.	multisubunit rna polymerase (rnap) is the central information-processing enzyme in all cellular life forms, yet its mechanism of translocation along the dna molecule remains conjectural. here, we report direct monitoring of bacterial rnap translocation following the addition of a single nucleotide. time-resolved measurements demonstrated that translocation is delayed relative to nucleotide incorporation and occurs shortly after or concurrently with pyrophosphate release. an investigation of tran ...	2012	22570421
structure based hypothesis of a mitochondrial ribosome rescue mechanism.	mtrf1 is a vertebrate mitochondrial protein with an unknown function that arose from a duplication of the mitochondrial release factor mtrf1a. to elucidate the function of mtrf1, we determined the positions that are conserved among mtrf1 sequences but that are different in their mtrf1a paralogs. we subsequently modeled the 3d structure of mtrf1a and mtrf1 bound to the ribosome, highlighting the structural implications of these differences to derive a hypothesis for the function of mtrf1.	2012	22569235
mycobacterium tuberculosis rbpa protein is a new type of transcriptional activator that stabilizes the σ a-containing rna polymerase holoenzyme.	rbpa is an rna polymerase (rnap)-binding protein whose presence increases the tolerance levels of mycobacteria to the first-line anti-tuberculosis drug rifampicin by an unknown mechanism. here, we show that the role of mycobacterium tuberculosis rbpa in resistance is indirect because it does not affect the sensitivity of rnap to rifampicin while it stimulates transcription controlled by the housekeeping σ(a)-factor. the transcription regulated by the stress-related σ(f) was not affected by rbpa. ...	2012	22570422
one ring to rule them all: trafficking of heme and heme synthesis intermediates in the metazoans.	the appearance of heme, an organic ring surrounding an iron atom, in evolution forever changed the efficiency with which organisms were able to generate energy, utilize gasses and catalyze numerous reactions. because of this, heme has become a near ubiquitous compound among living organisms. in this review we have attempted to assess the current state of heme synthesis and trafficking with a goal of identifying crucial missing information, and propose hypotheses related to trafficking that may g ...	2012	22575458
evolution of lysine biosynthesis in the phylum deinococcus-thermus.	thermus thermophilus biosynthesizes lysine through the α-aminoadipate (aaa) pathway: this observation was the first discovery of lysine biosynthesis through the aaa pathway in archaea and bacteria. genes homologous to the t. thermophilus lysine biosynthetic genes are widely distributed in bacteria of the deinococcus-thermus phylum. our phylogenetic analyses strongly suggest that a common ancestor of the deinococcus-thermus phylum had the ancestral genes for bacterial lysine biosynthesis through ...	2012	22645699
role of elongation and secondary pathways in s6 amyloid fibril growth.	the concerted action of a large number of individual molecular level events in the formation and growth of fibrillar protein structures creates a significant challenge for differentiating between the relative contributions of different self-assembly steps to the overall kinetics of this process. the characterization of the individual steps is, however, an important requirement for achieving a quantitative understanding of this general phenomenon which underlies many crucial functional and pathol ...	2012	22824281
stabilizing proteins from sequence statistics: the interplay of conservation and correlation in triosephosphate isomerase stability.	understanding the determinants of protein stability remains one of protein science's greatest challenges. there are still no computational solutions that calculate the stability effects of even point mutations with sufficient reliability for practical use. amino acid substitutions rarely increase the stability of native proteins; hence, large libraries and high-throughput screens or selections are needed to stabilize proteins using directed evolution. consensus mutations have proven effective fo ...	2012	22555051
the variable subdomain of escherichia coli seca functions to regulate seca atpase activity and adp release.	bacterial seca proteins can be categorized by the presence or absence of a variable subdomain (var) located within nucleotide-binding domain ii of the seca dead motor. here we show that var is dispensable for seca function, since the var deletion mutant secaδ519-547 displayed a wild-type rate of cellular growth and protein export. loss or gain of var is extremely rare in the history of bacterial evolution, indicating that it appears to contribute to seca function within the relevant species in t ...	2012	22389482
target- and resistance-based mechanistic studies with tp-434, a novel fluorocycline antibiotic.	tp-434 is a novel, broad-spectrum fluorocycline antibiotic with activity against bacteria expressing major antibiotic resistance mechanisms, including tetracycline-specific efflux and ribosomal protection. the mechanism of action of tp-434 was assessed using both cell-based and in vitro assays. in escherichia coli cells expressing recombinant tetracycline resistance genes, the mic of tp-434 (0.063 μg/ml) was unaffected by tet(m), tet(k), and tet(b) and increased to 0.25 and 4 μg/ml in the presen ...	2012	22354310
thermus thermophilus nucleoside phosphorylases active in the synthesis of nucleoside analogues.	cells extracts from thermus thermophilus hb27 express phosphorolytic activities on purines and pyrimidine nucleosides. five putative encoding genes were cloned and expressed in escherichia coli, and the corresponding recombinant proteins were purified and studied. two of these showed phosphorolytic activities against purine nucleosides, and third one showed phosphorolytic activity against pyrimidine nucleosides in vitro, and the three were named ttpnpi, ttpnpii, and ttpynp, respectively. the opt ...	2012	22344645
mycobacterium thermoresistibile as a source of thermostable orthologs of mycobacterium tuberculosis proteins.	the genus mycobacterium comprises major human pathogens such as the causative agent of tuberculosis, mycobacterium tuberculosis (mtb), and many environmental species. tuberculosis claims ~1.5 million lives every year, and drug resistant strains of mtb are rapidly emerging. to aid the development of new tuberculosis drugs, major efforts are currently under way to determine crystal structures of mtb drug targets and proteins involved in pathogenicity. however, a major obstacle to obtaining crystal ...	2012	22544630
crystal structure of dnak protein complexed with nucleotide exchange factor grpe in dnak chaperone system: insight into intermolecular communication.	the conserved, atp-dependent bacterial dnak chaperones process client substrates with the aid of the co-chaperones dnaj and grpe. however, in the absence of structural information, how these proteins communicate with each other cannot be fully delineated. for the study reported here, we solved the crystal structure of a full-length geobacillus kaustophilus hta426 grpe homodimer in complex with a nearly full-length g. kaustophilus hta426 dnak that contains the interdomain linker (acting as a pseu ...	2012	22544739
metallation state of human manganese superoxide dismutase expressed in saccharomyces cerevisiae.	human manganese superoxide dismutase (sod2p) has been expressed in yeast and the protein purified from isolated yeast mitochondria, yielding both the metallated protein and the less stable apoprotein in a single chromatographic step. at 30 °c growth temperature, more than half of the purified enzyme is apoprotein that can be fully activated following reconstitution, while the remainder contains a mixture of manganese and iron. in contrast, only fully metallated enzyme was isolated from a similar ...	2012	22561997
ispe inhibitors identified by a combination of in silico and in vitro high-throughput screening.	cdp-me kinase (ispe) contributes to the non-mevalonate or deoxy-xylulose phosphate (doxp) pathway for isoprenoid precursor biosynthesis found in many species of bacteria and apicomplexan parasites. ispe has been shown to be essential by genetic methods and since it is absent from humans it constitutes a promising target for antimicrobial drug development. using in silico screening directed against the substrate binding site and in vitro high-throughput screening directed against both, the substr ...	2012	22563402
a study on l-asparaginase of nocardia levis mk-vl_113.	an enzyme-based drug, l-asparaginase, was produced by nocardia levis mk-vl_113 isolated from laterite soils of guntur region. cultural parameters affecting the production of l-asparaginase by the strain were optimized. maximal yields of l-asparaginase were recorded from 3-day-old culture grown in modified asparagine-glycerol salts broth with initial ph 7.0 at temperature 30°c. glycerol (2%) and yeast extract (1.5%) served as good carbon and nitrogen sources for l-asparaginase production, respect ...	2012	22619604
crystallization and preliminary crystallographic analysis of the major capsid proteins vp16 and vp17 of bacteriophage p23-77.	members of the diverse double-β-barrel lineage of viruses are identified by the conserved structure of their major coat protein. new members of this lineage have been discovered based on structural analysis and we are interested in identifying relatives that utilize unusual versions of the double-β-barrel fold. one candidate for such studies is p23-77, an icosahedral dsdna bacteriophage that infects the extremophile thermus thermophilus. p23-77 has two major coat proteins, namely vp16 and vp17, ...	2012	22691792
preliminary crystallographic analysis of two hypothetical ribose-5-phosphate isomerases from streptococcus mutans.	study of the enzymes from sugar metabolic pathways may provide a better understanding of the pathogenesis of the human oral pathogen streptococcus mutans. bioinformatics, biochemical and crystallization methods were used to characterize and understand the function of two putative ribose-5-phosphate isomerases: smu1234 and smu2142. the proteins were cloned and constructed with n-terminal his tags. protein purification was performed by ni(2+)-chelating and size-exclusion chromatography. the crysta ...	2012	22691789
mechanism of foreign dna selection in a bacterial adaptive immune system.	in bacterial and archaeal crispr immune pathways, dna sequences from invading bacteriophage or plasmids are integrated into crispr loci within the host genome, conferring immunity against subsequent infections. the ribonucleoprotein complex cascade utilizes rnas generated from these loci to target complementary "nonself" dna sequences for destruction, while avoiding binding to "self" sequences within the crispr locus. here we show that casa, the largest protein subunit of cascade, is required fo ...	2012	22521690
crispr immunity relies on the consecutive binding and degradation of negatively supercoiled invader dna by cascade and cas3.	the prokaryotic crispr/cas immune system is based on genomic loci that contain incorporated sequence tags from viruses and plasmids. using small guide rna molecules, these sequences act as a memory to reject returning invaders. both the cascade ribonucleoprotein complex and the cas3 nuclease/helicase are required for crispr interference in escherichia coli, but it is unknown how natural target dna molecules are recognized and neutralized by their combined action. here we show that cascade effici ...	2012	22521689
the bacterial sec-translocase: structure and mechanism.	most bacterial secretory proteins pass across the cytoplasmic membrane via the translocase, which consists of a protein-conducting channel secyeg and an atp-dependent motor protein seca. the ancillary secdf membrane protein complex promotes the final stages of translocation. recent years have seen a major advance in our understanding of the structural and biochemical basis of protein translocation, and this has led to a detailed model of the translocation mechanism.	2012	22411975
solution nuclear magnetic resonance analyses of the anticodon arms of proteinogenic and nonproteinogenic trna(gly).	although the fate of most trna molecules in the cell is aminoacylation and delivery to the ribosome, some trnas are destined to fulfill other functional roles. in addition to their central role in translation, trna molecules participate in processes such as regulation of gene expression, bacterial cell wall biosynthesis, viral replication, antibiotic biosynthesis, and suppression of alternative splicing. in bacteria, glycyl-trna molecules with anticodon sequences gcc and ucc exhibit multiple ext ...	2012	22468768
hd-rnas: an automated hierarchical database of rna structures.	one of the important goals of most biological investigations is to classify and organize the experimental findings so that they are readily useful for deriving generalized rules. although there is a huge amount of information on rna structures in pdb, there are redundant files, ambiguous synthetic sequences etc. moreover, a systematic hierarchical organization, reflecting rna classification, is missing in pdb. in this investigation, we have classified all the available rna structures from pdb th ...	2012	22529851
dynamic, ligand-dependent conformational change triggers reaction of ribose-1,5-bisphosphate isomerase from thermococcus kodakarensis kod1.	ribose-1,5-bisphosphate isomerase (r15pi) is a novel enzyme recently identified as a member of an amp metabolic pathway in archaea. the enzyme converts d-ribose 1,5-bisphosphate into ribulose 1,5-bisphosphate, providing the substrate for archaeal ribulose-1,5-bisphosphate carboxylase/oxygenases. we here report the crystal structures of r15pi from thermococcus kodakarensis kod1 (tk-r15pi) with and without its substrate or product. tk-r15pi is a hexameric enzyme formed by the trimerization of dime ...	2012	22511789
molecular determinants of the cofactor specificity of ribitol dehydrogenase, a short-chain dehydrogenase/reductase.	ribitol dehydrogenase from zymomonas mobilis (zmrdh) catalyzes the conversion of ribitol to d-ribulose and concomitantly reduces nad(p)(+) to nad(p)h. a systematic approach involving an initial sequence alignment-based residue screening, followed by a homology model-based screening and site-directed mutagenesis of the screened residues, was used to study the molecular determinants of the cofactor specificity of zmrdh. a homologous conserved amino acid, ser156, in the substrate-binding pocket of ...	2012	22344653
the structure and function of the eukaryotic ribosome.	structures of the bacterial ribosome have provided a framework for understanding universal mechanisms of protein synthesis. however, the eukaryotic ribosome is much larger than it is in bacteria, and its activity is fundamentally different in many key ways. recent cryo-electron microscopy reconstructions and x-ray crystal structures of eukaryotic ribosomes and ribosomal subunits now provide an unprecedented opportunity to explore mechanisms of eukaryotic translation and its regulation in atomic ...	2012	22550233
ribosome engineering to promote new crystal forms.	crystallographic studies of the ribosome have provided molecular details of protein synthesis. however, the crystallization of functional complexes of ribosomes with gtpase translation factors proved to be elusive for a decade after the first ribosome structures were determined. analysis of the packing in different 70s ribosome crystal forms revealed that regardless of the species or space group, a contact between ribosomal protein l9 from the large subunit and 16s rrna in the shoulder of a neig ...	2012	22525755
the three-dimensional structural basis of type ii hyperprolinemia.	type ii hyperprolinemia is an autosomal recessive disorder caused by a deficiency in δ(1)-pyrroline-5-carboxylate dehydrogenase (p5cdh; also known as aldh4a1), the aldehyde dehydrogenase that catalyzes the oxidation of glutamate semialdehyde to glutamate. here, we report the first structure of human p5cdh (hsp5cdh) and investigate the impact of the hyperprolinemia-associated mutation of ser352 to leu on the structure and catalytic properties of the enzyme. the 2. 5-å-resolution crystal structure ...	2012	22516612
snapshot of virus evolution in hypersaline environments from the characterization of a membrane-containing salisaeta icosahedral phage 1.	the multitude of archaea and bacteria inhabiting extreme environments has only become evident during the last decades. as viruses apply a significant evolutionary force to their hosts, there is an inherent value in learning about viruses infecting these extremophiles. in this study, we have focused on one such unique virus-host pair isolated from a hypersaline environment: an icosahedral, membrane-containing double-stranded dna virus--salisaeta icosahedral phage 1 (ssip-1) and its halophilic hos ...	2012	22509017
riboswitch (t-box)-mediated control of trna-dependent amidation in clostridium acetobutylicum rationalizes gene and pathway redundancy for asparagine and asparaginyl-trnaasn synthesis.	analysis of the gram-positive clostridium acetobutylicum genome reveals an inexplicable level of redundancy for the genes putatively involved in asparagine (asn) and asn-trna(asn) synthesis. besides a duplicated set of gatcab trna-dependent amidotransferase genes, there is a triplication of aspartyl-trna synthetase genes and a duplication of asparagine synthetase b genes. this genomic landscape leads to the suspicion of the incoherent simultaneous use of the direct and indirect pathways of asn a ...	2012	22505715
substrate and enzyme functional groups contribute to translational quality control by bacterial prolyl-trna synthetase.	aminoacyl-trna synthetases activate specific amino acid substrates and attach them via an ester linkage to cognate trna molecules. in addition to cognate proline, prolyl-trna synthetase (prors) can activate cysteine and alanine and misacylate trna(pro). editing of the misacylated aminoacyl-trna is required for error-free protein synthesis. an editing domain (ins) appended to bacterial prors selectively hydrolyzes ala-trna(pro), whereas cys-trna(pro) is cleared by a freestanding editing domain, y ...	2012	22458656
rhodanese functions as sulfur supplier for key enzymes in sulfur energy metabolism.	how microorganisms obtain energy is a challenging topic, and there have been numerous studies on the mechanisms involved. here, we focus on the energy substrate traffic in the hyperthermophilic bacterium aquifex aeolicus. this bacterium can use insoluble sulfur as an energy substrate and has an intricate sulfur energy metabolism involving several sulfur-reducing and -oxidizing supercomplexes and enzymes. we demonstrate that the cytoplasmic rhodanese sbdp participates in this sulfur energy metabo ...	2012	22496367
para-like protein uses nonspecific chromosomal dna binding to partition protein complexes.	recent data have shown that plasmid partitioning par-like systems are used by some bacterial cells to control localization of protein complexes. here we demonstrate that one of these homologs, ppfa, uses nonspecific chromosome binding to separate cytoplasmic clusters of chemotaxis proteins upon division. using fluorescent microscopy and point mutations, we show dynamic chromosome binding and walker-type atpase activity are essential for cluster segregation. the n-terminal domain of a cytoplasmic ...	2012	22496588
structural and functional consequences of phosphate-arsenate substitutions in selected nucleotides: dna, rna, and atp.	a recent finding of a bacterial strain (gfaj-1) that can rely on arsenic instead of phosphorus raised the questions of if and how arsenate can replace phosphate in biomolecules that are essential to sustain cell life. apart from questions related to chemical stability, there are those of the structural and functional consequences of phosphate-arsenate substitutions in vital nucleotides in gfaj1-like cells. in this study we selected three types of molecules (atp/adp as energy source and replicati ...	2012	22480264
the type ii secretion system: biogenesis, molecular architecture and mechanism.	many gram-negative bacteria use the sophisticated type ii secretion system (t2ss) to translocate a wide range of proteins from the periplasm across the outer membrane. the inner-membrane platform of the t2ss is the nexus of the system and orchestrates the secretion process through its interactions with the periplasmic filamentous pseudopilus, the dodecameric outer-membrane complex and a cytoplasmic secretion atpase. here, recent structural and biochemical information is reviewed to describe our ...	2012	22466878
structural and mechanistic analysis of the membrane-embedded glycosyltransferase waaa required for lipopolysaccharide synthesis.	waaa is a key enzyme in the biosynthesis of lps, a critical component of the outer envelope of gram-negative bacteria. embedded in the cytoplasmic face of the inner membrane, waaa catalyzes the transfer of 3-deoxy-d-manno-oct-2-ulosonic acid (kdo) to the lipid a precursor of lps. here we present crystal structures of the free and cmp-bound forms of waaa from aquifex aeolicus, an ancient gram-negative hyperthermophile. these structures reveal details of the cmp-binding site and implicate a unique ...	2012	22474366
response to copper stress in streptomyces lividans extends beyond genes under direct control of a copper-sensitive operon repressor protein (csor).	a copper-sensitive operon repressor protein (csor) has been identified in streptomyces lividans (csor(sl)) and found to regulate copper homeostasis with attomolar affinity for cu(i). solution studies reveal apo- and cu(i)-csor(sl) to be a tetramer assembly, and a 1.7-å resolution crystal structure of apo-csor(sl) reveals that a significant conformational change is necessary to enable cu(i) binding. in silico prediction of the csor regulon was confirmed in vitro (emsa) and in vivo (rna-seq), whic ...	2012	22451651
role of trehalose in salinity and temperature tolerance in the model halophilic bacterium chromohalobacter salexigens.	the disaccharide trehalose is considered as a universal stress molecule, protecting cells and biomolecules from injuries imposed by high osmolarity, heat, oxidation, desiccation and freezing. chromohalobacter salexigens is a halophilic and extremely halotolerant γ-proteobacterium of the family halomonadaceae. in this work, we have investigated the role of trehalose as a protectant against salinity, temperature and desiccation in c. salexigens. a mutant deficient in the trehalose-6-phosphate synt ...	2012	22448254
folding without charges.	surface charges of proteins have in several cases been found to function as "structural gatekeepers," which avoid unwanted interactions by negative design, for example, in the control of protein aggregation and binding. the question is then if side-chain charges, due to their desolvation penalties, play a corresponding role in protein folding by avoiding competing, misfolded traps? to find out, we removed all 32 side-chain charges from the 101-residue protein s6 from thermus thermophilus. the re ...	2012	22454493
ultrafast excited-state deactivation of flavins bound to dodecin.	dodecins, a group of flavin-binding proteins with a dodecameric quaternary structure, are able to incorporate two flavins within each of their six identical binding pockets building an aromatic tetrade with two tryptophan residues. dodecin from the archaeal halobacterium salinarum is a riboflavin storage device. we demonstrate that unwanted side reactions induced by reactive riboflavin species and degradation of riboflavin are avoided by ultrafast depopulation of the reactive excited state of ri ...	2012	22451648
comparative analyses of homocitrate synthase genes of ascomycetous yeasts.	most ascomycetous yeasts have 2 homocitrate synthases (hcss). among the fungal lysine biosynthesis-related genes, only the hcs gene was duplicated in the course of evolution. it was recently reported that hcs of saccharomyces cerevisiae has an additional function in nuclear activities involving chromatin regulation related to dna damage repair, which is not related to lysine biosynthesis. thus, it is possible that the bifunctionality is associated with hcs gene duplication. phylogenetic analysis ...	2012	22518332
decoding in the absence of a codon by tmrna and smpb in the ribosome.	in bacteria, ribosomes stalled at the end of truncated messages are rescued by transfer-messenger rna (tmrna), a bifunctional molecule that acts as both a transfer rna (trna) and a messenger rna (mrna), and smpb, a small protein that works in concert with tmrna. here, we present the crystal structure of a tmrna fragment, smpb and elongation factor tu bound to the ribosome at 3.2 angstroms resolution. the structure shows how smpb plays the role of both the anticodon loop of trna and portions of m ...	2012	22422985
structural basis for the rescue of stalled ribosomes: structure of yaej bound to the ribosome.	in bacteria, the hybrid transfer-messenger rna (tmrna) rescues ribosomes stalled on defective messenger rnas (mrnas). however, certain gram-negative bacteria have evolved proteins that are capable of rescuing stalled ribosomes in a tmrna-independent manner. here, we report a 3.2 angstrom-resolution crystal structure of the rescue factor yaej bound to the thermus thermophilus 70s ribosome in complex with the initiator trna(i)(fmet) and a short mrna. the structure reveals that the c-terminal tail ...	2012	22422986
evolution of protein synthesis from an rna world.	because of the molecular complexity of the ribosome and protein synthesis, it is a challenge to imagine how translation could have evolved from a primitive rna world. two specific suggestions are made here to help to address this, involving separate evolution of the peptidyl transferase and decoding functions. first, it is proposed that translation originally arose not to synthesize functional proteins, but to provide simple (perhaps random) peptides that bound to rna, increasing its available s ...	2012	20610545
the open reading frame ttc1157 of thermus thermophilus hb27 encodes the methyltransferase forming n²-methylguanosine at position 6 in trna.	n(2)-methylguanosine (m(2)g) is found at position 6 in the acceptor stem of thermus thermophilus trna(phe). in this article, we describe the cloning, expression, and characterization of the t. thermophilus hb27 methyltransferase (mtase) encoded by the ttc1157 open reading frame that catalyzes the formation of this modified nucleoside. s-adenosyl-l-methionine is used as donor of the methyl group. the enzyme behaves as a monomer in solution. it contains an n-terminal thump domain predicted to bind ...	2012	22337946
regulation of expression and catalytic activity of escherichia coli rsmg methyltransferase.	rsmg is an adomet-dependent methyltransferase responsible for the synthesis of m(7)g527 in the 530 loop of bacterial 16s rrna. this loop is universally conserved, plays a key role in ribosomal accuracy, and is a target for streptomycin binding. loss of the m(7)g527 modification confers low-level streptomycin resistance and may affect ribosomal functioning. here, we explore the mechanisms controlling rsmg expression and activity, which may somehow respond to the demand set by the amount of rrna. ...	2012	22337945
a genome-wide view of the expression and processing patterns of thermus thermophilus hb8 crispr rnas.	the crispr-cas system represents an rna-based adaptive immune response system in prokaryotes and archaea. crisprs (clustered regularly interspaced short palindromic repeats) consist of arrays of short repeat-sequences interspaced by nonrepetitive short spacers, some of which show sequence similarity to foreign phage genetic elements. their cistronic transcripts are processed to produce the mature crispr rnas (crrnas), the elements that confer immunity by base-pairing with exogenous nucleic acids ...	2012	22355165