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the wobble hypothesis revisited: uridine-5-oxyacetic acid is critical for reading of g-ending codons.according to crick's wobble hypothesis, trnas with uridine at the wobble position (position 34) recognize a- and g-, but not u- or c-ending codons. however, u in the wobble position is almost always modified, and salmonella enterica trnas containing the modified nucleoside uridine-5-oxyacetic acid (cmo(5)u34) at this position are predicted to recognize u- (but not c-) ending codons, in addition to a- and g-ending codons. we have constructed a set of s. enterica mutants with only the cmo(5)u-cont ...200717942742
interactions and dynamics of the shine dalgarno helix in the 70s ribosome.the crystal structure of an initiation-like 70s ribosome complex containing an 8-bp shine-dalgarno (sd) helix was determined at 3.8-a resolution. translation-libration-screw analysis showed that the inherent anisotropic motions of the sd helix were biased along its helical axis, suggesting that during the first step of translocation, the sd helix moves along its helical screw axis. contacts between the sd helix and the ribosome were primarily through interactions with helices 23a, 26, and 28 of ...200717940016
separating the effects of mutation and selection in producing dna skew in bacterial chromosomes.many bacterial chromosomes display nucleotide asymmetry, or skew, between the leading and lagging strands of replication. mutational differences between these strands result in an overall pattern of skew that is centered about the origin of replication. such a pattern could also arise from selection coupled with a bias for genes coded on the leading strand. the relative contributions of selection and mutation in producing compositional skew are largely unknown.200717935620
a cysteine-rich ccg domain contains a novel [4fe-4s] cluster binding motif as deduced from studies with subunit b of heterodisulfide reductase from methanothermobacter marburgensis.heterodisulfide reductase (hdr) of methanogenic archaea with its active-site [4fe-4s] cluster catalyzes the reversible reduction of the heterodisulfide (com-s-s-cob) of the methanogenic coenzyme m (com-sh) and coenzyme b (cob-sh). com-hdr, a mechanistic-based paramagnetic intermediate generated upon half-reaction of the oxidized enzyme with com-sh, is a novel type of [4fe-4s]3+ cluster with com-sh as a ligand. subunit hdrb of the methanothermobacter marburgensis hdrabc holoenzyme contains two cy ...200717929940
transfer rna in the hybrid p/e state: correlating molecular dynamics simulations with cryo-em data.transfer rna (trna) transiently occupies the hybrid p/e state (p/e-trna) when mrna-trna are translocated in the ribosome. in this study, we characterize the structure of p/e-trna and its interactions with the ribosome by correlating the results from molecular dynamics simulations on free trna with the cryo-em map of p/e-trna. in our approach, we show that the cryo-em map may be interpreted as a conformational average. along the molecular dynamics trajectories (44 ns, 18 ns, and 18 ns), some of t ...200717925437
genetic and phenotypic identification of fusidic acid-resistant mutants with the small-colony-variant phenotype in staphylococcus aureus.small-colony variants (scvs) of staphylococcus aureus are a slow-growing subpopulation whose phenotypes can include resistance to aminoglycosides, defects in electron transport, and enhanced persistence in mammalian cells. here we show that a subset of mutants selected as scvs by reduced susceptibility to aminoglycosides are resistant to the antibiotic fusidic acid (fa) and conversely that a subset of mutants selected for resistance to fa are scvs. mutation analysis reveals different genetic cla ...200717923494
insights into hsp70 chaperone activity from a crystal structure of the yeast hsp110 sse1.classic hsp70 chaperones assist in diverse processes of protein folding and translocation, and hsp110s had seemed by sequence to be distant relatives within an hsp70 superfamily. the 2.4 a resolution structure of sse1 with atp shows that hsp110s are indeed hsp70 relatives, and it provides insight into allosteric coupling between sites for atp and polypeptide-substrate binding in hsp70s. subdomain structures are similar in intact sse1(atp) and in the separate hsp70 domains, but conformational dis ...200717923091
orotate phosphoribosyltransferase from corynebacterium ammoniagenes lacking a conserved lysine.the pyre gene, encoding orotate phosphoribosyltransferase (oprtase), was cloned by nested pcr and colony blotting from corynebacterium ammoniagenes atcc 6872, which is widely used in nucleotide production. sequence analysis shows that there is a lack of an important conserved lysine (lys 73 in salmonella enterica serovar typhimurium oprtase) in the c. ammoniagenes oprtase. this lysine has been considered to contribute to the initiation of catalysis. the enzyme was overexpressed and purified from ...200717921291
evolution and functional characterization of the rh50 gene from the ammonia-oxidizing bacterium nitrosomonas europaea.the family of ammonia and ammonium channel proteins comprises the amt proteins, which are present in all three domains of life with the notable exception of vertebrates, and the homologous rh proteins (rh50 and rh30) that have been described thus far only in eukaryotes. the existence of an rh50 gene in bacteria was first revealed by the genome sequencing of the ammonia-oxidizing bacterium nitrosomonas europaea. here we have used a phylogenetic approach to study the evolution of the n. europaea r ...200717921289
the carboxy-terminal coiled-coil of the rna polymerase beta'-subunit is the main binding site for gre factors.bacterial gre transcript cleavage factors stimulate the intrinsic endonucleolytic activity of rna polymerase (rnap) to rescue stalled transcription complexes. they bind to rnap and extend their coiled-coil (cc) domains to the catalytic centre through the secondary channel. three existing models for the gre-rnap complex postulate congruent mechanisms of gre-assisted catalysis, while offering conflicting views of the gre-rnap interactions. here, we report the greb structure of escherichia coli. th ...200717917675
analysis of gene order data supports vertical inheritance of the leukotoxin operon and genome rearrangements in the 5' flanking region in genus mannheimia.the mannheimia subclades belong to the same bacterial genus, but have taken divergent paths toward their distinct lifestyles. for example, m. haemolytica + m. glucosida are potential pathogens of the respiratory tract in the mammalian suborder ruminantia, whereas m. ruminalis, the supposed sister group, lives as a commensal in the ovine rumen. we have tested the hypothesis that vertical inheritance of the leukotoxin (lktcabd) operon has occurred from the last common ancestor of genus mannheimia ...200717915007
an expanded set of amino acid analogs for the ribosomal translation of unnatural peptides.the application of in vitro translation to the synthesis of unnatural peptides may allow the production of extremely large libraries of highly modified peptides, which are a potential source of lead compounds in the search for new pharmaceutical agents. the specificity of the translation apparatus, however, limits the diversity of unnatural amino acids that can be incorporated into peptides by ribosomal translation. we have previously shown that over 90 unnatural amino acids can be enzymatically ...200717912351
functional smpb-ribosome interactions require tmrna.small protein b (smpb) is a requisite component of the transfer messenger rna (tmrna)-mediated bacterial translational quality control system known as trans-translation. the initial binding of tmrna and its subsequent accommodation into the ribosomal a-site are activities intimately linked to smpb protein function. from a mechanistic perspective, two key unanswered questions that require further investigation are: 1) what constitutes a stalled ribosome recognition complex and 2) does smpb pre-bi ...200717911096
human trna(gly) acceptor-stem microhelix: crystallization and preliminary x-ray diffraction analysis at 1.2 a resolution.the major dissimilarities between the eukaryotic/archaebacterial-type and eubacterial-type glycyl-trna synthetase systems (glyrs; class ii aminoacyl-trna synthetases) represent an intriguing example of evolutionarily divergent solutions to similar biological functions. the differences in the identity elements of the respective trna(gly) systems are located within the acceptor stem and include the discriminator base u73. in the present work, the human trna(gly) acceptor-stem microhelix was crysta ...200717909289
cocrystallizing natural rna with its unnatural mirror image: biochemical and preliminary x-ray diffraction analysis of a 5s rrna a-helix racemate.chemically synthesized rnas with the unnatural l-configuration possess enhanced in vivo stability and nuclease resistance, which is a highly desirable property for pharmacological applications. for a structural comparison, both l- and d-rna oligonucleotides of a shortened thermus flavus 5s rrna a-helix were chemically synthesized. the enantiomeric rna duplexes were stochiometrically cocrystallized as a racemate, which enabled analysis of the d- and l-rna enantiomers in the same crystals. in addi ...200717909284
x-ray structures of two proteins belonging to pfam duf178 revealed unexpected structural similarity to the duf191 pfam family.pfam is a comprehensive collection of protein domains and families, with a range of well-established information including genome annotation. pfam has two large series of functionally uncharacterized families, known as domains of unknown function (dufs) and uncharacterized protein families (upfs).200717908300
electron transport chain of saccharomyces cerevisiae mitochondria is inhibited by h2o2 at succinate-cytochrome c oxidoreductase level without lipid peroxidation involvement.the deleterious effects of h202 on the electron transport chain of yeast mitochondria and on mitochondrial lipid peroxidation were evaluated. exposure to h2o2 resulted in inhibition of the oxygen consumption in the uncoupled and phosphorylating states to 69% and 65%, respectively. the effect of h2o2 on the respiratory rate was associated with an inhibition of succinate-ubiquinone and succinate-dcip oxidoreductase activities. inhibitory effect of h2o2 on respiratory complexes was almost completel ...200717907001
identification of promoters recognized by rna polymerase-sigmae holoenzyme from thermus thermophilus hb8.thermus thermophilus sigma(e), an extracytoplasmic function sigma factor from the extremely thermophilic bacterium thermus thermophilus hb8, bound to the rna polymerase core enzyme and showed transcriptional activity. with the combination of in vitro transcription assay and genechip technology, we identified three promoters recognized by sigma(e). the predicted consensus promoter sequence for sigma(e) is 5'-ca(a/t)(a/c)c(a/c)-n(15)-ccgta-3'.200717905996
evaluation of the staphylococcus aureus class c nonspecific acid phosphatase (saps) as a reporter for gene expression and protein secretion in gram-negative and gram-positive bacteria.a phosphatase secreted by staphylococcus aureus strain 154 has previously been characterized and classified as a new member of the bacterial class c family of nonspecific acid phosphatases. as the acid phosphatase activity can be easily detected with a cost-effective plate screen assay, quantitatively measured by a simple enzyme assay, and detected by zymography, its potential use as a reporter system was investigated. the s. aureus acid phosphatase (saps) gene has been cloned and expressed from ...200717905879
functional analysis of the gtpases enga and yhbz encoded by salmonella typhimurium.the s. typhimurium genome encodes proteins, designated enga and yhbz, which have a high sequence identity with the gtpases enga/der and obge/cgtae of escherichia coli. the wild-type activity of the e. coli proteins is essential for normal ribosome maturation and cell viability. in order to characterize the potential involvement of the salmonella typhimurium enga and yhbz proteins in ribosome biology, we used high stringency affinity chromatography experiments to identify strongly binding ribosom ...200717905831
deinococcus geothermalis: the pool of extreme radiation resistance genes shrinks.bacteria of the genus deinococcus are extremely resistant to ionizing radiation (ir), ultraviolet light (uv) and desiccation. the mesophile deinococcus radiodurans was the first member of this group whose genome was completely sequenced. analysis of the genome sequence of d. radiodurans, however, failed to identify unique dna repair systems. to further delineate the genes underlying the resistance phenotypes, we report the whole-genome sequence of a second deinococcus species, the thermophile de ...200717895995
molecular architecture of strictosidine glucosidase: the gateway to the biosynthesis of the monoterpenoid indole alkaloid family.strictosidine beta-d-glucosidase (sg) follows strictosidine synthase (str1) in the production of the reactive intermediate required for the formation of the large family of monoterpenoid indole alkaloids in plants. this family is composed of approximately 2000 structurally diverse compounds. sg plays an important role in the plant cell by activating the glucoside strictosidine and allowing it to enter the multiple indole alkaloid pathways. here, we report detailed three-dimensional information d ...200717890378
flavin-dependent thymidylate synthase thyx activity: implications for the folate cycle in bacteria.although flavin-dependent thyx proteins show thymidylate synthase activity in vitro and functionally complement thya defects in heterologous systems, direct proof of their cellular functions is missing. using insertional mutagenesis of rhodobacter capsulatus thyx, we constructed the first defined thyx inactivation mutant. phenotypic analyses of the obtained mutant strain confirmed that r. capsulatus thyx is required for de novo thymidylate synthesis. full complementation of the r. capsulatus thy ...200717890305
dissection of 16s rrna methyltransferase (ksga) function in escherichia coli.a 16s rrna methyltransferase, ksga, identified originally in escherichia coli is highly conserved in all living cells, from bacteria to humans. ksga orthologs in eukaryotes possess functions in addition to their rrna methyltransferase activity. e. coli era is an essential gtp-binding protein. we recently observed that ksga functions as a multicopy suppressor for the cold-sensitive cell growth of an era mutant [era(e200k)] strain (q. lu and m. inouye, j. bacteriol. 180:5243-5246, 1998). here we o ...200717890303
saturation mutagenesis of a +1 programmed frameshift-inducing mrna sequence derived from a yeast retrotransposon.errors during the process of translating mrna information into protein products occur infrequently. frameshift errors occur less frequently than other types of errors, suggesting that the translational machinery has more robust mechanisms for precluding that kind of error. despite these mechanisms, mrna sequences have evolved that increase the frequency up to 10,000-fold. these sequences, termed programmed frameshift sites, usually consist of a heptameric nucleotide sequence, at which the change ...200717881742
role of hsp104 in the propagation and inheritance of the [het-s] prion.the chaperones of the clpb/hsp100 family play a central role in thermotolerance in bacteria, plants, and fungi by ensuring solubilization of heat-induced protein aggregates. in addition in yeast, hsp104 was found to be required for prion propagation. herein, we analyze the role of podospora anserina hsp104 (pahsp104) in the formation and propagation of the [het-s] prion. we show that deltapahsp104 strains propagate [het-s], making [het-s] the first native fungal prion to be propagated in the abs ...200717881723
human ribosomal protein s13 regulates expression of its own gene at the splicing step by a feedback mechanism.the expression of ribosomal protein (rp) genes is regulated at multiple levels. in yeast, two genes are autoregulated by feedback effects of the protein on pre-mrna splicing. here, we have investigated whether similar mechanisms occur in eukaryotes with more complicated and highly regulated splicing patterns. comparisons of the sequences of ribosomal protein s13 gene (rps13) among mammals and birds revealed that intron 1 is more conserved than the other introns. transfection of hek 293 cells wit ...200717881366
pathogenic mechanism of a human mitochondrial trnaphe mutation associated with myoclonic epilepsy with ragged red fibers syndrome.human mitochondrial trna (hmt-trna) mutations are associated with a variety of diseases including mitochondrial myopathies, diabetes, encephalopathies, and deafness. because the current understanding of the precise molecular mechanisms of these mutations is limited, there is no efficient method to treat their associated mitochondrial diseases. here, we use a variety of known mutations in hmt-trna(phe) to investigate the mechanisms that lead to malfunctions. we tested the impact of hmt-trna(phe) ...200717878308
phosphopantetheine adenylyltransferase from escherichia coli: investigation of the kinetic mechanism and role in regulation of coenzyme a biosynthesis.phosphopantetheine adenylyltransferase (ppat) from escherichia coli is an essential hexameric enzyme that catalyzes the penultimate step in coenzyme a (coa) biosynthesis and is a target for antibacterial drug discovery. the enzyme utilizes mg-atp and phosphopantetheine (php) to generate dephospho-coa (dpcoa) and pyrophosphate. when overexpressed in e. coli, ppat copurifies with tightly bound coa, suggesting a feedback inhibitory role for this cofactor. using an enzyme-coupled assay for the forwa ...200717873050
conformational energy and structure in canonical and noncanonical forms of trna determined by temperature analysis of the rate of s(4)u8-c13 photocrosslinking.bacterial trnas frequently have 4-thiouridine (s(4)u) modification at position 8, which is adjacent to the c13-g22-m(7)g46 base triple in the elbow region of the trna tertiary structure. irradiation with light in the uva range induces an efficient photocrosslink between s(4)u8 and c13. the temperature dependence of the rate constants for photocrosslinking between the s(4)u8 and c13 has been used to investigate the trna conformational energy and structure in escherichia coli trna(val), trna(phe), ...200717872510
substrate specificity and properties of the escherichia coli 16s rrna methyltransferase, rsme.the small ribosome subunit of escherichia coli contains 10 base-methylated sites distributed in important functional regions. at present, seven enzymes responsible for methylation of eight bases are known, but most of them have not been well characterized. one of these enzymes, rsme, was recently identified and shown to specifically methylate u1498. here we describe the enzymatic properties and substrate specificity of rsme. the enzyme forms dimers in solution and is most active in the presence ...200717872509
structural and kinetic characterization of quinolinate phosphoribosyltransferase (hqprtase) from homo sapiens.human quinolinate phosphoribosyltransferase (ec 2.4.2.19) (hqprtase) is a member of the type ii phosphoribosyltransferase family involved in the catabolism of quinolinic acid (qa). it catalyses the formation of nicotinic acid mononucleotide from quinolinic acid, which involves a phosphoribosyl transfer reaction followed by decarboxylation. hqprtase has been implicated in a number of neurological conditions and in order to study it further, we have carried out structural and kinetic studies on re ...200717868694
a quantitative kinetic scheme for 70 s translation initiation complex formation.association of the 30 s initiation complex (30sic) and the 50 s ribosomal subunit, leading to formation of the 70 s initiation complex (70sic), is a critical step of the translation initiation pathway. the 70sic contains initiator trna, fmet-trna(fmet), bound in the p (peptidyl)-site in response to the aug start codon. we have formulated a quantitative kinetic scheme for the formation of an active 70sic from 30sic and 50 s subunits on the basis of parallel rapid kinetics measurements of gtp hydr ...200717868692
virus-encoded aminoacyl-trna synthetases: structural and functional characterization of mimivirus tyrrs and metrs.aminoacyl-trna synthetases are pivotal in determining how the genetic code is translated in amino acids and in providing the substrate for protein synthesis. as such, they fulfill a key role in a process universally conserved in all cellular organisms from their most complex to their most reduced parasitic forms. in contrast, even complex viruses were not found to encode much translation machinery, with the exception of isolated components such as trnas. in this context, the discovery of four am ...200717855524
elongation factor 1a mediates the specificity of mitochondrial trna import in t. brucei.mitochondrial trna import is widespread in eukaryotes. yet, the mechanism that determines its specificity is unknown. previous in vivo experiments using the trnas(met), trna(ile) and trna(lys) have suggested that the t-stem nucleotide pair 51:63 is the main localization determinant of trnas in trypanosoma brucei. in the cytosol-specific initiator trna(met), this nucleotide pair is identical to the main antideterminant that prevents interaction with cytosolic elongation factor (eef1a). here we sh ...200717853889
nmr analysis of [methyl-13c]methionine uvrb from bacillus caldotenax reveals uvrb-domain 4 heterodimer formation in solution.uvrb is a central dna damage recognition protein involved in bacterial nucleotide excision repair. structural information has been limited by the apparent disorder of the c-terminal domain 4 in crystal structures of intact uvrb; in solution, the isolated domain 4 is found to form a helix-loop-helix dimer. in order to gain insight into the behavior of uvrb in solution, we have performed nmr studies on [methyl-13c]methionine-labeled uvrb from bacillus caldotenax (molecular mass=75 kda). the 13 met ...200717822711
ribosome biogenesis and the translation process in escherichia coli.translation, the decoding of mrna into protein, is the third and final element of the central dogma. the ribosome, a nucleoprotein particle, is responsible and essential for this process. the bacterial ribosome consists of three rrna molecules and approximately 55 proteins, components that are put together in an intricate and tightly regulated way. when finally matured, the quality of the particle, as well as the amount of active ribosomes, must be checked. the focus of this review is ribosome b ...200717804668
first experimental evidence for the preferential stabilization of the natural d- over the nonnatural l-configuration in nucleic acids.the homochirality of biomolecules is a prerequisite for the origin and evolution of terrestrial life. the unique selection of d-monosaccharides, in particular, d-ribose in rna and d-deoxyribose in dna, leads to the construction of proteins by l-amino acids. this points to the exclusive role of stereoselectivity in the most important physiological processes. so far, there is no experimental confirmation for the theoretical calculations of the energy differences between enantiomers used for the ex ...200717804644
phenylalanyl-trna synthetase editing defects result in efficient mistranslation of phenylalanine codons as tyrosine.translational quality control is monitored at several steps, including substrate selection by aminoacyl-trna synthetases (aarss), and discrimination of aminoacyl-trnas by elongation factor tu (ef-tu) and the ribosome. phenylalanyl-trna synthetase (phers) misactivates tyr but is able to correct the mistake using a proofreading activity named editing. previously we found that overproduction of editing-defective phers resulted in tyr incorporation at phe-encoded positions in vivo, although the misr ...200717804641
purification, crystallization and preliminary x-ray analysis of the fumarylacetoacetase family member ttha0809 from thermus thermophilus hb8.fumarylacetoacetase catalyzes the final step of tyrosine and phenylalanine catabolism. a recombinant form of the fumarylacetoacetase family member ttha0809 from thermus thermophilus hb8 has been crystallized by the oil-microbatch method using sodium chloride as a precipitating agent. the crystals belong to the monoclinic space group p2(1), with unit-cell parameters a = 93.3, b = 73.4, c = 122.6 a, beta = 111.8 degrees. the crystals are most likely to contain two dimers in the asymmetric unit, wi ...200717768357
crystallization and preliminary x-ray crystallographic analysis of a 40 kda n-terminal fragment of the yeast prion-remodeling factor hsp104.a 40 kda n-terminal fragment of saccharomyces cerevisiae hsp104 was crystallized in two different crystal forms. native 1 diffracted to 2.6 a resolution and belonged to space group p2(1)2(1)2(1), with unit-cell parameters a = 66.6, b = 75.8, c = 235.7 a. native 2 diffracted to 2.9 a resolution and belonged to space group p6(1)22 or p6(5)22, with unit-cell parameters a = 179.1, b = 179.1, c = 69.7 a. this is the first report of the crystallization of a eukaryotic member of the hsp100 family of mo ...200717768355
purification, crystallization and preliminary x-ray characterization of a human mitochondrial phenylalanyl-trna synthetase.human monomeric mitochondrial phenylalanyl-trna synthetase (mitphers) is an enzyme that catalyzes the charging of trna with the cognate amino acid phenylalanine. human mitphers is a chimera of the bacterial alpha-subunit of phers and the b8 domain of its beta-subunit. together, the alpha-subunit and the 'rnp-domain' (b8 domain) at the c-terminus form the minimal structural set to construct an enzyme with phenylalanylation activity. the recombinant human mitphers was purified to homogeneity and c ...200717768348
the primordial metabolism: an ancestral interconnection between leucine, arginine, and lysine biosynthesis.it is generally assumed that primordial cells had small genomes with simple genes coding for enzymes able to react with a wide range of chemically related substrates, interconnecting different metabolic routes. new genes coding for enzymes with a narrowed substrate specificity arose by paralogous duplication(s) of ancestral ones and evolutionary divergence. in this way new metabolic pathways were built up by primordial cells. useful hints to disclose the origin and evolution of ancestral metabol ...200717767731
an aminoacyl-trna synthetase:elongation factor complex for substrate channeling in archaeal translation.translation requires the specific attachment of amino acids to trnas by aminoacyl-trna synthetases (aarss) and the subsequent delivery of aminoacyl-trnas to the ribosome by elongation factor 1 alpha (ef-1alpha). interactions between ef-1alpha and various aarss have been described in eukaryotes, but the role of these complexes remains unclear. to investigate possible interactions between ef-1alpha and other cellular components, a yeast two-hybrid screen was performed for the archaeon methanotherm ...200717766929
analysis of promoter targets for escherichia coli transcription elongation factor grea in vivo and in vitro.transcription elongation factor grea induces nucleolytic activity of bacterial rna polymerase (rnap). in vitro, transcript cleavage by grea contributes to transcription efficiency by (i) suppressing pauses and arrests, (ii) stimulating rnap promoter escape, and (iii) enhancing transcription fidelity. however, it is unclear which of these functions is (are) most relevant in vivo. by comparing global gene expression profiles of escherichia coli strains lacking gre factors and strains expressing ei ...200717766423
structure-based design of robust glucose biosensors using a thermotoga maritima periplasmic glucose-binding protein.we report the design and engineering of a robust, reagentless fluorescent glucose biosensor based on the periplasmic glucose-binding protein obtained from thermotoga maritima (tmgbp). the gene for this protein was cloned from genomic dna and overexpressed in escherichia coli, the identity of its cognate sugar was confirmed, ligand binding was studied, and the structure of its glucose complex was solved to 1.7 angstrom resolution by x-ray crystallography. tmgbp is specific for glucose and exhibit ...200717766373
redox-dependent change of nucleotide affinity to the active site of the mammalian complex i.a very potent and specific inhibitor of mitochondrial nadh:ubiquinone oxidoreductase (complex i), a derivative of nadh (nadh-oh) has recently been discovered (kotlyar, a. b., karliner, j. s., and cecchini, g. (2005) febs lett. 579, 4861-4866). here we present a quantitative analysis of the interaction of nadh-oh and other nucleotides with oxidized and reduced complex i in tightly coupled submitochondrial particles. both the rate of the nadh-oh binding and its affinity to complex i are strongly d ...200717760425
anticodon recognition and discrimination by the alpha-helix cage domain of class i lysyl-trna synthetase.aminoacyl-trna synthetases are normally found in one of two mutually exclusive structural classes, the only known exception being lysyl-trna synthetase which exists in both classes i (lysrs1) and ii (lysrs2). differences in trna acceptor stem recognition between lysrs1 and lysrs2 do not drastically impact cellular aminoacylation levels, focusing attention on the mechanism of trna anticodon recognition by lysrs1. on the basis of structure-based sequence alignments, seven trnalys anticodon variant ...200717760422
human tryptophanyl-trna synthetase is switched to a trna-dependent mode for tryptophan activation by mutations at v85 and i311.for most aminoacyl-trna synthetases (aars), their cognate trna is not obligatory to catalyze amino acid activation, with the exception of four class i (aars): arginyl-trna synthetase, glutamyl-trna synthetase, glutaminyl-trna synthetase and class i lysyl-trna synthetase. furthermore, for arginyl-, glutamyl- and glutaminyl-trna synthetase, the integrated 3' end of the trna is necessary to activate the atp-ppi exchange reaction. tryptophanyl-trna synthetase is a class i aars that catalyzes tryptop ...200717726052
messenger rna conformations in the ribosomal e site revealed by x-ray crystallography.a comparison of messenger rna in x-ray crystal structures of 70s ribosomal complexes in the initiation, post-initiation and elongation states of translation shows distinct conformational differences in the exit (e) codon. here, we present structural evidence indicating that, after the initiation event, the e codon nucleotides relax and form a classical a-helical conformation. this conformation is similar to that of the p and a codons, and is favourable for establishing watson-crick interactions ...200717721443
conserved lipid-binding sites in membrane proteins: a focus on cytochrome c oxidase.specific interactions between lipids and membrane proteins have been observed in recent high-resolution crystal structures of membrane proteins. a number of cytochrome oxidase structures were analyzed, along with many amino acid sequences of membrane-spanning regions aligned according to their location in the membrane. the results reveal conservation of lipid-binding sites and of the residues that form them. these studies imply that bound lipids have important roles that are crucial to the assem ...200717719219
allosteric control of the rna polymerase by the elongation factor rfah.efficient transcription of long polycistronic operons in bacteria frequently relies on accessory proteins but their molecular mechanisms remain obscure. rfah is a cellular elongation factor that acts as a polarity suppressor by increasing rna polymerase (rnap) processivity. in this work, we provide evidence that rfah acts by reducing transcriptional pausing at certain positions rather than by accelerating rnap at all sites. we show that 'fast' rnap variants are characterized by pause-free rna ch ...200717711918
enzymes involved in dna ligation and end-healing in the radioresistant bacterium deinococcus radiodurans.enzymes involved in dna metabolic events of the highly radioresistant bacterium deinococcus radiodurans are currently examined to understand the mechanisms that protect and repair the deinococcus radiodurans genome after extremely high doses of gamma-irradiation. although several deinococcus radiodurans dna repair enzymes have been characterised, no biochemical data is available for dna ligation and dna endhealing enzymes of deinococcus radiodurans so far. dna ligases are necessary to seal broke ...200717705817
conformations of flanking bases in hiv-1 rna dis kissing complexes studied by molecular dynamics.explicit solvent molecular dynamics simulations (in total almost 800 ns including locally enhanced sampling runs) were applied with different ion conditions and with two force fields (amber and charmm) to characterize typical geometries adopted by the flanking bases in the rna kissing-loop complexes. we focus on flanking base positions in multiple x-ray and nmr structures of hiv-1 dis kissing complexes and kissing complex from the large ribosomal subunit of haloarcula marismortui. an initial x-r ...200717704156
a functional interaction of smpb with tmrna for determination of the resuming point of trans-translation.in trans-translation, transfer-messenger rna (tmrna), possessing a dual function as a trna and an mrna, relieves a stalled translation on the ribosome with the help of smpb. here, we established an in vitro system using escherichia coli translation and trans-translation factors to evaluate two steps of trans-translation, peptidyl transfer from peptidyl-trna to alanyl-tmrna and translation of the resume codon on tmrna. using this system, the effects of several mutations upstream of the tag-encodi ...200717698641
type i and type ii fatty acid biosynthesis in eimeria tenella: enoyl reductase activity and structure.apicomplexan parasites of the genus eimeria are the major causative agent of avian coccidiosis, leading to high economic losses in the poultry industry. recent results show that eimeria tenella harbours an apicoplast organelle, and that a key biosynthetic enzyme, enoyl reductase, is located in this organelle. in related parasites, enoyl reductase is one component of a type ii fatty acid synthase (fas) and has proven to be an attractive target for antimicrobial compounds. we cloned and expressed ...200717697396
fr3d: finding local and composite recurrent structural motifs in rna 3d structures.new methods are described for finding recurrent three-dimensional (3d) motifs in rna atomic-resolution structures. recurrent rna 3d motifs are sets of rna nucleotides with similar spatial arrangements. they can be local or composite. local motifs comprise nucleotides that occur in the same hairpin or internal loop. composite motifs comprise nucleotides belonging to three or more different rna strand segments or molecules. we use a base-centered approach to construct efficient, yet exhaustive sea ...200717694311
fr3d: finding local and composite recurrent structural motifs in rna 3d structures.new methods are described for finding recurrent three-dimensional (3d) motifs in rna atomic-resolution structures. recurrent rna 3d motifs are sets of rna nucleotides with similar spatial arrangements. they can be local or composite. local motifs comprise nucleotides that occur in the same hairpin or internal loop. composite motifs comprise nucleotides belonging to three or more different rna strand segments or molecules. we use a base-centered approach to construct efficient, yet exhaustive sea ...200717694311
identification of indole derivatives as self-growth inhibitors of symbiobacterium thermophilum, a unique bacterium whose growth depends on coculture with a bacillus sp.symbiobacterium thermophilum is a syntrophic bacterium whose growth depends on coculture with a bacillus sp. recently, we discovered that co(2) generated by bacillus is the major inducer for the growth of s. thermophilum; however, the evidence suggested that an additional element is required for its full growth. here, we studied the self-growth-inhibitory substances produced by s. thermophilum. we succeeded in purifying two substances from an ether extract of the culture supernatant of s. thermo ...200717693561
hitting bacteria at the heart of the central dogma: sequence-specific inhibition.abstract: an important objective in developing new drugs is the achievement of high specificity to maximize curing effect and minimize side-effects, and high specificity is an integral part of the antisense approach. the antisense techniques have been extensively developed from the application of simple long, regular antisense rna (asrna) molecules to highly modified versions conferring resistance to nucleases, stability of hybrid formation and other beneficial characteristics, though still pres ...200717692125
analysis of a nuclease activity of catalytic domain of thermus thermophilus muts2 by high-accuracy mass spectrometry.electrospray ionization with fourier-transform ion cyclotron resonance mass spectrometry (esi-ft icr ms) is a powerful tool for analyzing the precise structural features of biopolymers, including oligonucleotides. here, we described the detailed characterization of a newly discovered nuclease activity of the c-terminal domain of thermus thermophilus muts2 (ttmuts2). using this method, the length, nucleotide content and nature of the 5'- and 3'-termini of the product oligonucleotides were accurat ...200717686785
discovery of novel dna gyrase inhibitors by high-throughput virtual screening.the bacterial type ii topoisomerases dna gyrase and topoisomerase iv are validated targets for clinically useful quinolone antimicrobial drugs. a significant limitation to widely utilized quinolone inhibitors is the emergence of drug-resistant bacteria due to an altered dna gyrase. to address this problem, we have used structure-based molecular docking to identify novel drug-like small molecules that target sites distinct from those targeted by quinolone inhibitors. a chemical ligand database co ...200717682095
crystal structure of the escherichia coli regulator of sigma70, rsd, in complex with sigma70 domain 4.the escherichia coli rsd protein binds tightly and specifically to the rna polymerase (rnap) sigma(70) factor. rsd plays a role in alternative sigma factor-dependent transcription by biasing the competition between sigma(70) and alternative sigma factors for the available core rnap. here, we determined the 2.6 a-resolution x-ray crystal structure of rsd bound to sigma(70) domain 4 (sigma(70)(4)), the primary determinant for rsd binding within sigma(70). the structure reveals that rsd binding int ...200717681541
sympatric drosophila simulans flies with distinct mtdna show age related differences in mitochondrial metabolism.the primary causes of age-related changes in mitochondrial metabolism are not known. the goal of this study is to document the influence of naturally occurring mtdna variation on age dependent changes in mitochondrial respiration, hydrogen peroxide (h(2)o(2)) generation and antioxidant defenses in the fly drosophila simulans. possible changes include an increase in rates of reactive oxygen species production with age and/or an age dependent decrease in antioxidant response. for this study we hav ...200717681231
the archaeon methanosarcina acetivorans contains a protein disulfide reductase with an iron-sulfur cluster.methanosarcina acetivorans, a strictly anaerobic methane-producing species belonging to the domain archaea, contains a gene cluster annotated with homologs encoding oxidative stress proteins. one of the genes (ma3736) is annotated as a gene encoding an uncharacterized carboxymuconolactone decarboxylase, an enzyme required for aerobic growth with aromatic compounds by species in the domain bacteria. methane-producing species are not known to utilize aromatic compounds, suggesting that ma3736 is i ...200717675382
complete genomic sequence and mass spectrometric analysis of highly diverse, atypical bacillus thuringiensis phage 0305phi8-36.to investigate the apparent genomic complexity of long-genome bacteriophages, we have sequenced the 218,948-bp genome (6479-bp terminal repeat), and identified the virion proteins (55), of bacillus thuringiensis bacteriophage 0305phi8-36. phage 0305phi8-36 is an atypical myovirus with three large curly tail fibers. an accurate mode of dna pyrosequencing was used to sequence the genome and mass spectrometry was used to accomplish the comprehensive virion protein survey. advanced informatic techni ...200717673272
identification of genes encoding trna modification enzymes by comparative genomics.as the molecular adapters between codons and amino acids, transfer-rnas are pivotal molecules of the genetic code. the coding properties of a trna molecule do not reside only in its primary sequence. posttranscriptional nucleoside modifications, particularly in the anticodon loop, can modify cognate codon recognition, affect aminoacylation properties, or stabilize the codon-anticodon wobble base pairing to prevent ribosomal frameshifting. despite a wealth of biophysical and structural knowledge ...200717673083
crystallization and preliminary crystallographic analysis of hygromycin b phosphotransferase from escherichia coli.aminoglycoside antibiotics, such as hygromycin, kanamycin, neomycin, spectinomycin and streptomycin, inhibit protein synthesis by acting on bacterial and eukaryotic ribosomes. hygromycin b phosphotransferase (hph; ec 2.7.1.119) converts hygromycin b to 7''-o-phosphohygromycin using a phosphate moiety from atp, resulting in the loss of its cell-killing activity. the hph protein has been crystallized for the first time using a thermostable mutant and the hanging-drop vapour-diffusion method. the c ...200717671368
crystallization and preliminary x-ray diffraction analysis of the full-length mg2+ transporter mgte.the mgte family of mg(2+) transporters are ubiquitously conserved in all three domains. the genes encoding full-length mgte from seven different species were cloned. three of the seven mgte transporters were overexpressed and purified for use in crystallization trials. only thermus thermophilus mgte was successfully crystallized using the sitting-drop vapour-diffusion method. selenomethionine-substituted (semet) crystals were obtained by cross-microseeding using the native microcrystals. the sem ...200717671367
crystallization and preliminary x-ray diffraction analysis of the cytosolic domain of the mg2+ transporter mgte.the mgte family of mg(2+) transporters is ubiquitously conserved in all domains of life. the cytosolic domains of the mgte mg(2+) transporters include a cystathionine-beta-synthase (cbs) domain which is known to play a regulatory function in transporter proteins. the cytosolic domain of mgte from thermus thermophilus was overexpressed, purified and crystallized in the presence and absence of mg(2+). the crystals formed in the presence of mg(2+) diffracted x-rays to 2.3 a resolution using synchro ...200717671366
crystallization and preliminary x-ray diffraction analysis of a soluble domain of the putative zinc transporter czrb from thermus thermophilus.czrb is a putative zinc transporter from thermus thermophilus. the protein is proposed to consist of a hexahelical transmembrane domain with a cytosolic extramembranal c-terminus. the latter 92-residue fragment may be expressed free and may function independently of the full-length integral membrane protein. a 6xhis-tagged form of the water-soluble fragment has been overexpressed in escherichia coli and diffraction-quality crystals of the tagged and tag-free variants have been grown. preliminary ...200717671365
negamycin binds to the wall of the nascent chain exit tunnel of the 50s ribosomal subunit.negamycin, a small-molecule inhibitor of protein synthesis, binds the haloarcula marismortui 50s ribosomal subunit at a single site formed by highly conserved rna nucleotides near the cytosolic end of the nascent chain exit tunnel. the mechanism of antibiotic action and the function of this unexplored tunnel region remain intriguingly elusive.200717664317
tertiary structure and function of an rna motif required for plant vascular entry to initiate systemic trafficking.vascular entry is a decisive step for the initiation of long-distance movement of infectious and endogenous rnas, silencing signals and developmental/defense signals in plants. however, the mechanisms remain poorly understood. we used potato spindle tuber viroid (pstvd) as a model to investigate the direct role of the rna itself in vascular entry. we report here the identification of an rna motif that is required for pstvd to traffic from nonvascular into the vascular tissue phloem to initiate s ...200717660743
biosynthesis of isoprenoids in plants: structure of the 2c-methyl-d-erithrytol 2,4-cyclodiphosphate synthase from arabidopsis thaliana. comparison with the bacterial enzymes.the x-ray crystal structure of the 2c-methyl-d-erythritol 2,4-cyclodiphosphate synthase (mcs) from arabidopsis thaliana has been solved at 2.3 a resolution in complex with a cytidine-5-monophosphate (cmp) molecule. this is the first structure determined of an mcs enzyme from a plant. major differences between the a. thaliana and bacterial mcs structures are found in the large molecular cavity that forms between subunits and involve residues that are highly conserved among plants. in some bacteri ...200717660251
a comparative analysis of the triloops in all high-resolution rna structures reveals sequence structure relationships.despite an increasing number of experimentally determined rna structures, the gap between the number of structures and that of rna families is still growing. to overcome this limitation, efficient and reliable rna modeling methodologies must be developed. in order to reach this goal, here, we show how triloop sequence-structure relationships have been inferred through a systematic analysis of all triloops found in available high-resolution structures. the structural annotation of all triloops al ...200717652406
changes in the conformation of 5s rrna cause alterations in principal functions of the ribosomal nanomachine.5s rrna is an integral component of the large ribosomal subunit in virtually all living organisms. polyamine binding to 5s rrna was investigated by cross-linking of n1-azidobenzamidino (aba)-spermine to naked 5s rrna or 50s ribosomal subunits and whole ribosomes from escherichia coli cells. aba-spermine cross-linking sites were kinetically measured and their positions in 5s rrna were localized by primer extension analysis. helices iii and v, and loops a, c, d and e in naked 5s rrna were found to ...200717652323
mg2+ binding and archaeosine modification stabilize the g15 c48 levitt base pair in trnas.the g15-c48 levitt base pair, located at a crucial position in the core of canonical trnas, assumes a reverse watson-crick (rwc) geometry. by means of bioinformatics analysis and quantum mechanics calculations we show here that such a geometry is moderately more stable than an alternative bifurcated trans geometry, involving the guanine watson-crick face and the cytosine keto group, which we have also found in known rna structures. however we also demonstrate that the rwc geometry can take advan ...200717652139
effects of the deletion of the escherichia coli frataxin homologue cyay on the respiratory nadh:ubiquinone oxidoreductase.frataxin is discussed as involved in the biogenesis of iron-sulfur clusters. recently it was discovered that a frataxin homologue is a structural component of the respiratory nadh:ubiquinone oxidoreductase (complex i) in thermus thermophilus. it was not clear whether frataxin is in general a component of complex i from bacteria. the escherichia coli homologue of frataxin is coined cyay.200717650323
molecular breeding of polymerases for amplification of ancient dna.in the absence of repair, lesions accumulate in dna. thus, dna persisting in specimens of paleontological, archaeological or forensic interest is inevitably damaged. we describe a strategy for the recovery of genetic information from damaged dna. by molecular breeding of polymerase genes from the genus thermus (taq (thermus aquaticus), tth (thermus thermophilus) and tfl (thermus flavus)) and compartmentalized self-replication selection, we have evolved polymerases that can extend single, double ...200717632524
human endonuclease viii-like (neil) proteins in the giant dna mimivirus.endonuclease viii (nei), which recognizes and repairs oxidized pyrimidines in the base excision repair (ber) pathway, is sparsely distributed among both the prokaryotes and eukaryotes. recently, we and others identified three homologs of escherichia coli endonuclease viii-like (neil) proteins in humans. here, we report identification of human neil homologs in mimivirus, a giant dna virus that infects acanthamoeba. characterization of the two mimiviral homologs, mvnei1 and mvnei2, showed that the ...200717627905
crystal structure of hyperthermophilic esterase este1 and the relationship between its dimerization and thermostability properties.este1 is a hyperthermophilic esterase belonging to the hormone-sensitive lipase family and was originally isolated by functional screening of a metagenomic library constructed from a thermal environmental sample. dimers and oligomers may have been evolutionally selected in thermophiles because intersubunit interactions can confer thermostability on the proteins. the molecular mechanisms of thermostabilization of this extremely thermostable esterase are not well understood due to the lack of stru ...200717625021
crystallization and preliminary x-ray crystallographic study of a putative aspartyl-trna synthetase from the crenarchaeon sulfolobus tokodaii strain 7.genome analysis suggests that the aspartyl-trna synthetase of the crenarchaeon sulfolobus tokodaii strain 7 belongs to the nondiscriminating type that is believed to catalyze aspartylation of trna(asp) and trna(asn). this protein has been overexpressed in escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method from 100 mm sodium hepes buffer ph 7.5 containing 100 mm nacl and 1.6 m (nh4)2so4 as the crystallizing reagent. diffraction data were collected to 2.3 a ...200717620724
crystallization and preliminary x-ray analysis of the oxygenase component (hpab) of 4-hydroxyphenylacetate 3-monooxygenase from thermus thermophilus hb8.the 4-hydroxyphenylacetate (4hpa) 3-monooxygenase enzyme catalyzes the hydroxylation of 4hpa to 3,4-dihydroxyphenylacetate in the initial step of the degradation pathway of 4hpa. this enzyme consists of two components: an oxygenase (hpab) and a reductase (hpac). hpab hydroxylates 4hpa using an oxygen molecule and a reduced flavin, which is supplied by hpac. hpab from thermus thermophilus hb8 was overexpressed in escherichia coli and crystallized. crystals of hpab were grown in 0.4 m 1,6-hexanedi ...200717620709
a computational pipeline for high- throughput discovery of cis-regulatory noncoding rna in prokaryotes.noncoding rnas (ncrnas) are important functional rnas that do not code for proteins. we present a highly efficient computational pipeline for discovering cis-regulatory ncrna motifs de novo. the pipeline differs from previous methods in that it is structure-oriented, does not require a multiple-sequence alignment as input, and is capable of detecting rna motifs with low sequence conservation. we also integrate rna motif prediction with rna homolog search, which improves the quality of the rna mo ...200717616982
use of an escherichia coli recombinant producing thermostable polyphosphate kinase as an atp regenerator to produce fructose 1,6-diphosphate.heat-treated escherichia coli producing thermus polyphosphate kinase regenerated atp by using exogenous polyphosphate. this recombinant could be used as a platform to produce valuable compounds in combination with thermostable phosphorylating or energy-requiring enzymes. in this work, we demonstrated the production of fructose 1,6-diphosphate from fructose and polyphosphate.200717616610
structural characterization of the ribosome maturation protein, rimm.the rimm protein has been implicated in the maturation of the 30s ribosomal subunit. it binds to ribosomal protein s19, located in the head domain of the 30s subunit. multiple sequence alignments predicted that rimm possesses two domains in its n- and c-terminal regions. in the present study, we have produced thermus thermophilus rimm in both the full-length form (162 residues) and its n-terminal fragment, spanning residues 1 to 85, as soluble proteins in escherichia coli and have performed stru ...200717616598
nanoscale dewetting transition in protein complex folding.in a previous study, a surprising drying transition was observed to take place inside the nanoscale hydrophobic channel in the tetramer of the protein melittin. the goal of this paper is to determine if there are other protein complexes capable of displaying a dewetting transition during their final stage of folding. we searched the entire protein data bank (pdb) for all possible candidates, including protein tetramers, dimers, and two-domain proteins, and then performed the molecular dynamics ( ...200717608515
small but versatile: the extraordinary functional and structural diversity of the beta-grasp fold.the beta-grasp fold (beta-gf), prototyped by ubiquitin (ub), has been recruited for a strikingly diverse range of biochemical functions. these functions include providing a scaffold for different enzymatic active sites (e.g. nudix phosphohydrolases) and iron-sulfur clusters, rna-soluble-ligand and co-factor-binding, sulfur transfer, adaptor functions in signaling, assembly of macromolecular complexes and post-translational protein modification. to understand the basis for the functional versatil ...200717605815
thermostability of proteins: role of metal binding and ph on the stability of the dinuclear cua site of thermus thermophilus.the dinuclear copper center (ttcua) forming the electron entry site in the subunit ii of the cytochrome c oxidase in thermus thermophilus shows high stability toward thermal as well as denaturant-induced unfolding of the protein at ambient ph. we have studied the effect of ph on the stability of the holo-protein as well as of the apo-protein by uv-visible absorption, far-uv, and visible circular dichroism spectroscopy. the results show that the holo-protein both in the native mixed-valence state ...200717604317
mechanistic studies of the long chain acyl-coa synthetase faa1p from saccharomyces cerevisiae.long chain acyl-coa synthetase (acsl; fatty acid coa ligase: amp forming; ec 6.2.1.3) catalyzes the formation of acyl-coa through a process, which requires fatty acid, atp and coenzymea as substrates. in the yeast saccharomyces cerevisiae the principal acsl is faa1p (encoded by the faa1 gene). the preferred substrates for this enzyme are cis-monounsaturated long chain fatty acids. our previous work has shown faa1p is a principal component of a fatty acid transport/activation complex that also in ...200717604220
use of a dominant rpsl allele conferring streptomycin dependence for positive and negative selection in thermus thermophilus.a spontaneous rpsl mutant of thermus thermophilus was isolated in a search for new selection markers for this organism. this new allele, named rpsl1, encodes a k47r/k57e double mutant s12 ribosomal protein that confers a streptomycin-dependent (sd) phenotype to t. thermophilus. models built on the available three-dimensional structures of the 30s ribosomal subunit revealed that the k47r mutation directly affects the streptomycin binding site on s12, whereas the k57e does not apparently affect th ...200717601820
paradigm development: comparative and predictive 3d modeling of hiv-1 virion infectivity factor (vif).obtaining structural information about vif is of interest for several reasons that include the study of the interaction of vif with apobec3g, a resistance factor. vif is a potential drug target and its function is essential for the hiv-1 infectivity process. to study vif mechanism of action, we need to decipher its structure. pivotal in this approach is the painstaking prediction of its protein structure. the three-dimensional (3d) crystal structure for vif has not been established. in order to ...200617597910
modeling of phosphomethyl pyrimidine kinase from leptospira interrogans serovar lai strain 56601.many microorganisms, as well as plants and fungi, synthesize thiamin, but vertebrates do not produce it. phosphomethyl pyrimidine kinase is an enzyme involved in an intermediary step of thiamin biosynthesis from purine molecules. this enzyme is absent in humans. thus, it is a potential chemotherapeutic target for antileptospiral treatment. structure of this enzyme from leptospira interrogans serovar lai strain 56601 has not yet been elucidated. we used the structural template of phosphomethyl py ...200617597880
protein subunit interfaces: heterodimers versus homodimers.protein dimers are either homodimers (complexation of identical monomers) or heterodimers (complexation of non-identical monomers). these dimers are common in catalysis and regulation. however, the molecular principles of protein dimer interactions are difficult to understand mainly due to the geometrical and chemical characteristics of proteins. nonetheless, the principles of protein dimer interactions are often studied using a dataset of 3d structural complexes determined by x-ray crystallogra ...200517597849
advancements in the pathophysiology of friedreich's ataxia and new prospects for treatments.on november 9-12, 2006, the friedreich's ataxia research alliance (fara) and the national institutes of health (nih) hosted the third international friedreich's ataxia (frda) scientific conference at the nih in bethesda, maryland, highlighting the exciting research leading now to a variety of clinical trials that show promise of effective treatments for this devastating disorder. nearly 150 leading frda scientists from around the world discussed their new insights and findings. the presence of s ...200717596984
geranylgeranyl diphosphate synthase in fission yeast is a heteromer of farnesyl diphosphate synthase (fps), fps1, and an fps-like protein, spo9, essential for sporulation.both farnesyl diphosphate synthase (fps) and geranylgeranyl diphosphate synthase (ggps) are key enzymes in the synthesis of various isoprenoid-containing compounds and proteins. here, we describe two novel schizosaccharomyces pombe genes, fps1(+) and spo9(+), whose products are similar to fps in primary structure, but whose functions differ from one another. fps1 is essential for vegetative growth, whereas, a spo9 null mutant exhibits temperature-sensitive growth. expression of fps1(+), but not ...200717596513
testing constraints on rrna bases that make nonsequence-specific contacts with the codon-anticodon complex in the ribosomal a site.during protein synthesis, interactions between the decoding center of the ribosome and the codon-anticodon complexes maintain translation accuracy. correct aminoacyl-trnas induce the ribosome to shift into a "closed" conformation that both blocks trna dissociation and accelerates the process of trna acceptance. as part of the ribosomal recognition of cognate trnas, the rrna nucleotides g530 and a1492 form a hydrogen-bonded pair that interacts with the middle position of the codon.anticodon compl ...200717592040
direct localization of the 51 and 24 kda subunits of mitochondrial complex i by three-dimensional difference imaging.complex i is the largest complex in the respiratory chain, and the least understood. we have determined the 3d structure of complex i from yarrowia lipolytica lacking the flavoprotein part of the n-module, which consists of the 51 kda (nubm) and the 24 kda (nuhm) subunits. the reconstruction was determined by 3d electron microscopy of single particles. a comparison to our earlier reconstruction of the complete y. lipolytica complex i clearly assigns the two flavoprotein subunits to an outer lobe ...200717591445
crystal structures of tm0549 and ne1324--two orthologs of e. coli ahas isozyme iii small regulatory subunit.crystal structures of two orthologs of the regulatory subunit of acetohydroxyacid synthase iii (ahas, ec 2.2.1.6) from thermotoga maritima (tm0549) and nitrosomonas europea (ne1324) were determined by single-wavelength anomalous diffraction methods with the use of selenomethionine derivatives at 2.3 a and 2.5 a, respectively. tm0549 and ne1324 share the same fold, and in both proteins the polypeptide chain contains two separate domains of a similar size. each protein contains a c-terminal domain ...200717586771
the 2.2 a resolution crystal structure of bacillus cereus nif3-family protein yqfo reveals a conserved dimetal-binding motif and a regulatory domain.yqfo of bacillus cereus is a member of the widespread nif3 family of proteins, which has been highlighted as an important target for structural genomics. the n- and c-terminal domains are conserved across the family and contain a dimetal-binding motif in a putative active site. yqfo contains an insert in the middle of the protein, present in a minority of bacterial family members. the structure of yqfo was determined at a resolution of 2.2 a and reveals conservation of the putative active site. ...200717586767
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