Publications
| Title | Abstract | Year(sorted ascending) Filter | PMID Filter |
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| structural basis of the interaction between the aaa atpase p97/vcp and its adaptor protein p47. | the aaa atpase p97/vcp is involved in many cellular events including ubiquitin-dependent processes and membrane fusion. in the latter, the p97 adaptor protein p47 is of central importance. in order to provide insight into the molecular basis of p97 adaptor binding, we have determined the crystal structure of p97 nd1 domains complexed with p47 c-terminal domain at 2.9 a resolution. the structure reveals that the p47 ubiquitin regulatory x domain (ubx) domain interacts with the p97 n domain via a ... | 2004 | 14988733 |
| the kink-turn motif in rna is dimorphic, and metal ion-dependent. | the kink-turn (k-turn) is a new motif in rna structure that was identified by examination of the crystal structures of the ribosome. we examined the structural and dynamic properties of this element in free solution. the k-turn rna exists in a dynamic equilibrium between a tightly kinked conformation and a more open structure similar to a simple bulge bend. the highly kinked form is stabilized by the noncooperative binding of metal ions, but a significant population of the less-kinked form is pr ... | 2004 | 14730024 |
| unscrambling an egg: protein disaggregation by aaa+ proteins. | aprotein quality control system, consisting of molecular chaperones and proteases, controls the folding status of proteins and prevents the aggregation of misfolded proteins by either refolding or degrading aggregation-prone species. during severe stress conditions this protection system can be overwhelmed by high substrate load, resulting in the formation of protein aggregates. in such emergency situations, hsp104/clpb becomes a key player for cell survival, as it has the extraordinary capacity ... | 2004 | 14728719 |
| efficiency and pattern of uv pulse laser-induced rna-rna cross-linking in the ribosome. | escherichia coli ribosomes were irradiated with a krf excimer laser (248 nm, 22 ns pulse) with incident pulse energies in the range of 10-40 mj for a 1 cm2 area, corresponding to fluences of 4.5 to 18 x 10(9) w m(-2), to determine strand breakage yields and the frequency and pattern of rna-rna cross- linking in the 16s rrna. samples were irradiated in a cuvette with one laser pulse or in a flow cell with an average of 4.6 pulses per sample. the yield of strand breaks per photon was intensity dep ... | 2004 | 14999094 |
| crystal structure of rlmai: implications for understanding the 23s rrna g745/g748-methylation at the macrolide antibiotic-binding site. | the rlma class of enzymes (rlma(i) and rlma(ii)) catalyzes n1-methylation of a guanine base (g745 in gram-negative and g748 in gram-positive bacteria) of hairpin 35 of 23s rrna. we have determined the crystal structure of escherichia coli rlma(i) at 2.8-a resolution, providing 3d structure information for the rlma class of rna methyltransferases. the dimeric protein structure exhibits features that provide new insights into its molecular function. each rlma(i) molecule has a zn-binding domain, r ... | 2004 | 14999102 |
| definition of bases in 23s rrna essential for ribosomal subunit association. | the ribosome is a two-subunit molecular machine, sporting a working cycle that involves coordinated movements of the subunits. recent structural studies of the 70s ribosome describe a rather large number of intersubunit contacts, some of which are dynamic during translocation. we set out to determine which intersubunit contacts are functionally indispensable for the association of ribosome subunits by using a modification interference approach. modification of the n-1 position of a715, a1912, or ... | 2004 | 15037769 |
| probing the q-proton pathway of ba3-cytochrome c oxidase by time-resolved fourier transform infrared spectroscopy. | in cytochrome c oxidase, the terminal respiratory enzyme, electron transfers are strongly coupled to proton movements within the enzyme. two proton pathways (k and d) containing water molecules and hydrophobic amino acids have been identified and suggested to be involved in the proton translocation from the mitochondrial matrix or the bacterial cytoplasm into the active site. in addition to the k and d proton pathways, a third proton pathway (q) has been identified only in ba3-cytochrome c oxida ... | 2004 | 15041681 |
| dna damage recognition of mutated forms of uvrb proteins in nucleotide excision repair. | the dna repair protein uvrb plays an indispensable role in the stepwise and sequential damage recognition of nucleotide excision repair in escherichia coli. our previous studies suggested that uvrb is responsible for the chemical damage recognition only upon a strand opening mediated by uvra. difficulties were encountered in studying the direct interaction of uvrb with adducts due to the presence of uvra. we report herein that a single point mutation of y95w in which a tyrosine is replaced by a ... | 2004 | 15065863 |
| thermodynamic characterization of the interaction of mutant uvrb protein with damaged dna. | during the dna damage recognition of nucleotide excision repair in escherichia coli the interaction of uvrb protein with damaged dna ensures the recognition of differences in the intrinsic chemical structures of a variety of adduct molecules in dna double helix. our earlier study indicated that a single tyrosine-to-tryptophan mutation at residue 95 converted the uvrb to a protein [uvrb(y95w)] that is able to bind to a structure-specific bubble dna substrate, even in the absence of uvra. fluoresc ... | 2004 | 15065864 |
| an aminoacyl-trna synthetase-like protein encoded by the escherichia coli yadb gene glutamylates specifically trnaasp. | the product of the escherichia coli yadb gene is homologous to the n-terminal part of bacterial glutamyl-trna synthetases (glurss), including the rossmann fold with the acceptor-binding domain and the stem-contact fold. this glurs-like protein, which lacks the anticodon-binding domain, does not use trna(glu) as substrate in vitro nor in vivo, but aminoacylates trna(asp) with glutamate. the yadb gene is expressed in wild-type e. coli as an operon with the dksa gene, which encodes a protein involv ... | 2004 | 15096594 |
| a truncated aminoacyl-trna synthetase modifies rna. | aminoacyl-trna synthetases are modular enzymes composed of a central active site domain to which additional functional domains were appended in the course of evolution. analysis of bacterial genome sequences revealed the presence of many shorter aminoacyl-trna synthetase paralogs. here we report the characterization of a well conserved glutamyl-trna synthetase (glurs) paralog (yadb in escherichia coli) that is present in the genomes of >40 species of proteobacteria, cyanobacteria, and actinobact ... | 2004 | 15096612 |
| specific recognition of rpso mrna and 16s rrna by escherichia coli ribosomal protein s15 relies on both mimicry and site differentiation. | the ribosomal protein s15 binds to 16s rrna, during ribosome assembly, and to its own mrna (rpso mrna), affecting autocontrol of its expression. in both cases, the rna binding site is bipartite with a common subsite consisting of a g*u/g-c motif. the second subsite is located in a three-way junction in 16s rrna and in the distal part of a stem forming a pseudoknot in escherichia coli rpso mrna. to determine the extent of mimicry between these two rna targets, we determined which amino acids inte ... | 2004 | 15101974 |
| crystal structures of escherichia coli topoisomerase iv pare subunit (24 and 43 kilodaltons): a single residue dictates differences in novobiocin potency against topoisomerase iv and dna gyrase. | topoisomerase iv and dna gyrase are related bacterial type ii topoisomerases that utilize the free energy from atp hydrolysis to catalyze topological changes in the bacterial genome. the essential function of dna gyrase is the introduction of negative dna supercoils into the genome, whereas the essential function of topoisomerase iv is to decatenate daughter chromosomes following replication. here, we report the crystal structures of a 43-kda n-terminal fragment of escherichia coli topoisomerase ... | 2004 | 15105144 |
| thermal and conformational stability of ssh10b protein from archaeon sulfolobus shibattae. | the secondary structure of the dna binding protein ssh10b is largely unaffected by change in temperature between 25 degrees c and 85 degrees c, indicating that the protein is highly thermostable. here, we report the temperature-dependent equilibrium denaturation of ssh10b in the presence of guanidine hydrochloride (gdnhcl). it was found that the transition midpoint values of the temperature (t(m)), and changes of enthalpy (deltah(m)) and entropy (deltas(m)) of ssh10b unfolding were linearly decr ... | 2004 | 15107015 |
| ring-shaped architecture of recr: implications for its role in homologous recombinational dna repair. | recr, together with recf and reco, facilitates reca loading in the recf pathway of homologous recombinational dna repair in procaryotes. the human rad52 protein is a functional counterpart of recfor. we present here the crystal structure of recr from deinococcus radiodurans (dr recr). a monomer of dr recr has a two-domain structure: the n-terminal domain with a helix-hairpin-helix (hhh) motif and the c-terminal domain with a cys4 zinc-finger motif, a toprim domain and a walker b motif. four such ... | 2004 | 15116069 |
| distinctive protein signatures provide molecular markers and evidence for the monophyletic nature of the deinococcus-thermus phylum. | the deinococcus-thermus group of species is currently recognized as a distinct phylum solely on the basis of their branching in 16s rrna trees. no unique biochemical or molecular characteristics that can distinguish this group from all other bacteria are known at present. in this work, we describe eight conserved indels (viz., inserts or deletions) in seven widely distributed proteins that are distinctive characteristics of the deinococcus-thermus phylum but are not found in any other group of b ... | 2004 | 15126471 |
| comparison of two real-time quantitative assays for detection of severe acute respiratory syndrome coronavirus. | the new severe acute respiratory syndrome (sars) coronavirus (cov), described in february 2003, infected a total of 8,439 people. a total of 812 people died due to respiratory insufficiency. close contact with symptomatic patients appeared to be the main route of transmission. however, potential transmission by blood transfusion could not be definitely excluded. two real-time sars-specific pcr assays were assessed for their sensitivities, agreement of test results, and intra-assay variabilities. ... | 2004 | 15131175 |
| thermus thermophilus genome analysis: benefits and implications. | the genome sequence analysis of thermus thermophilus hb27, a microorganism with high biotechnological potential, has recently been published. in that report, the chromosomal and the megaplasmid sequence were compared to those of other organisms and discussed on the basis of their physiological and metabolic features. out of the 2,218 putative genes identified through the large genome sequencing project, a significant number has potential interest for biotechnology. the present communication will ... | 2004 | 15134584 |
| mycobacterium tuberculosis dnaa initiator protein: purification and dna-binding requirements. | the mycobacterium tuberculosis oric (the origin of chromosomal replication) region contains 13 non-perfect dnaa boxes. the m. tuberculosis initiator protein, dnaa, was overexpressed in escherichia coli as a soluble his-tagged fusion protein. the purified protein his6mtdnaa was investigated for its binding properties to dnaa boxes from the oric region. gel retardation demonstrated that the dnaa from m. tuberculosis requires two dnaa boxes for efficient binding. electron microscopy as well as dnas ... | 2004 | 15137907 |
| resistance of rumen bacteria murein to bovine gastric lysozyme. | lysozymes, enzymes mostly associated with defence against bacterial infections, are mureinolytic. ruminants have evolved a gastric c type lysozyme as a digestive enzyme, and profit from digestion of foregut bacteria, after most dietary components, including protein, have been fermented in the rumen. in this work we characterized the biological activities of bovine gastric secretions against membranes, purified murein and bacteria. | 2004 | 15137912 |
| mapping of the second tetracycline binding site on the ribosomal small subunit of e.coli. | tetracycline blocks stable binding of aminoacyl-trna to the bacterial ribosomal a-site. various tetracycline binding sites have been identified in crystals of the 30s ribosomal small subunit of thermus thermophilus. here we describe a direct photo- affinity modification of the ribosomal small subunits of escherichia coli with 7-[3h]-tetracycline. to select for specific interactions, an excess of the 30s subunits over tetracycline has been used. primer extension analysis of the 16s rrna revealed ... | 2004 | 15141029 |
| the structure of a ribosomal protein s8/spc operon mrna complex. | in bacteria, translation of all the ribosomal protein cistrons in the spc operon mrna is repressed by the binding of the product of one of them, s8, to an internal sequence at the 5' end of the l5 cistron. the way in which the first two genes of the spc operon are regulated, retroregulation, is mechanistically distinct from translational repression by s8 of the genes from l5 onward. a 2.8 a resolution crystal structure has been obtained of escherichia coli s8 bound to this site. despite sequence ... | 2004 | 15146079 |
| a minimalist glutamyl-trna synthetase dedicated to aminoacylation of the trnaasp quc anticodon. | escherichia coli encodes yadb, a protein displaying 34% identity with the catalytic core of glutamyl-trna synthetase but lacking the anticodon-binding domain. we show that yadb is a trna modifying enzyme that evidently glutamylates the queuosine residue, a modified nucleoside at the wobble position of the trna(asp) quc anticodon. this conclusion is supported by a variety of biochemical data and by the inability of the enzyme to glutamylate trna(asp) isolated from an e.coli trna-guanosine transgl ... | 2004 | 15150343 |
| simulation, experiment, and evolution: understanding nucleation in protein s6 folding. | in this study, we explore nucleation and the transition state ensemble of the ribosomal protein s6 using a monte carlo (mc) go model in conjunction with restraints from experiment. the results are analyzed in the context of extensive experimental and evolutionary data. the roles of individual residues in the folding nucleus are identified, and the order of events in the s6 folding mechanism is explored in detail. interpretation of our results agrees with, and extends the utility of, experiments ... | 2004 | 15150413 |
| new enzymes from environmental cassette arrays: functional attributes of a phosphotransferase and an rna-methyltransferase. | by targeting gene cassettes by polymerase chain reaction (pcr) directly from environmentally derived dna, we are able to amplify entire open reading frames (orfs) independently of prior sequence knowledge. approximately 10% of the mobile genes recovered by these means can be attributed to known protein families. here we describe the characterization of two orfs which show moderate homology to known proteins: (1) an aminoglycoside phosphotransferase displaying 25% sequence identity with aph(7") f ... | 2004 | 15152095 |
| crystal structure of escherichia coli cytidine triphosphate synthetase, a nucleotide-regulated glutamine amidotransferase/atp-dependent amidoligase fusion protein and homologue of anticancer and antiparasitic drug targets. | cytidine triphosphate synthetases (ctpss) produce ctp from utp and glutamine, and regulate intracellular ctp levels through interactions with the four ribonucleotide triphosphates. we solved the 2.3-a resolution crystal structure of escherichia coli ctps using hg-mad phasing. the structure reveals a nearly symmetric 222 tetramer, in which each bifunctional monomer contains a dethiobiotin synthetase-like amidoligase n-terminal domain and a type 1 glutamine amidotransferase c-terminal domain. for ... | 2004 | 15157079 |
| crystal structure of the deinococcus radiodurans single-stranded dna-binding protein suggests a mechanism for coping with dna damage. | single-stranded dna (ssdna)-binding (ssb) proteins are uniformly required to bind and protect single-stranded intermediates in dna metabolic pathways. all bacterial and eukaryotic ssb proteins studied to date oligomerize to assemble four copies of a conserved domain, called an oligonucleotide/oligosaccharide-binding (ob) fold, that cooperate in nonspecific ssdna binding. the vast majority of bacterial ssb family members function as homotetramers, with each monomer contributing a single ob fold. ... | 2004 | 15159541 |
| the ribosomal protein rps15p is required for nuclear exit of the 40s subunit precursors in yeast. | we have conducted a genetic screen in order to identify ribosomal proteins of saccharomyces cerevisiae involved in nuclear export of the small subunit precursors. this has led us to distinguish rps15p as a protein dispensable for maturation of the pre-40s particles, but whose assembly into the pre-ribosomes is a prerequisite to their nuclear exit. upon depletion of rps15p, 20s pre-rrna is released from the nucleolus and retained in the nucleus, without alteration of the pre-rrna early cleavages. ... | 2004 | 15167894 |
| identification of active site residues in mevalonate diphosphate decarboxylase: implications for a family of phosphotransferases. | a combination of sequence homology analyses of mevalonate diphosphate decarboxylase (mdd) proteins and structural information for mdd leads to the hypothesis that asp 302 and lys 18 are active site residues in mdd. these residues were mutated to replace acidic/basic side chains and the mutant proteins were isolated and characterized. binding and competitive displacement studies using trinitrophenyl-atp, a fluorescent analog of substrate atp, indicate that these mutant enzymes (d302a, d302n, k18m ... | 2004 | 15169949 |
| visualization of ribosome-recycling factor on the escherichia coli 70s ribosome: functional implications. | after the termination step of protein synthesis, a deacylated trna and mrna remain associated with the ribosome. the ribosome-recycling factor (rrf), together with elongation factor g (ef-g), disassembles this posttermination complex into mrna, trna, and the ribosome. we have obtained a three-dimensional cryo-electron microscopic map of a complex of the escherichia coli 70s ribosome and rrf. we find that rrf interacts mainly with the segments of the large ribosomal subunit's (50s) rrna helices t ... | 2004 | 15178758 |
| cloning, sequencing, and characterization of a heat- and alkali-stable type i pullulanase from anaerobranca gottschalkii. | the gene encoding a type i pullulanase was identified from the genome sequence of the anaerobic thermoalkaliphilic bacterium anaerobranca gottschalkii. in addition, the homologous gene was isolated from a gene library of anaerobranca horikoshii and sequenced. the proteins encoded by these two genes showed 39% amino acid sequence identity to the pullulanases from the thermophilic anaerobic bacteria fervidobacterium pennivorans and thermotoga maritima. the pullulanase gene from a. gottschalkii (en ... | 2004 | 15184138 |
| interaction of horse heart and thermus thermophilus type c cytochromes with phospholipid vesicles and hydrophobic surfaces. | the binding of horse heart cytochrome c (cyt-c) and thermus thermophilus cytochrome c(552) (cyt-c(552)) to dioleoyl phosphatidylglycerol (dopg) vesicles was investigated using fourier transform infrared (ftir) spectroscopy and turbidity measurements. ftir spectra revealed that the tertiary structures of both cytochromes became more open when bound to dopg vesicles, but this was more pronounced for cyt-c. their secondary structures were unchanged. turbidity measurements showed important differenc ... | 2004 | 15189883 |
| interactions between uvra and uvrb: the role of uvrb's domain 2 in nucleotide excision repair. | nucleotide excision repair (ner) is a highly conserved dna repair mechanism present in all kingdoms of life. uvrb is a central component of the bacterial ner system, participating in damage recognition, strand excision and repair synthesis. none of the three presently available crystal structures of uvrb has defined the structure of domain 2, which is critical for the interaction with uvra. we have solved the crystal structure of the uvrb y96a variant, which reveals a new fold for domain 2 and i ... | 2004 | 15192705 |
| thiostrepton-resistant mutants of thermus thermophilus. | ribosomal protein l11 and its associated binding site on 23s rrna together comprise one of the principle components that mediate interactions of translation factors with the ribosome. this site is also the target of the antibiotic thiostrepton, which has been proposed to act by preventing important structural transitions that occur in this region of the ribosome during protein synthesis. here, we describe the isolation and characterization of spontaneous thiostrepton-resistant mutants of the ext ... | 2004 | 15199170 |
| the role of bacterial antizyme: from an inhibitory protein to atoc transcriptional regulator. | this review considers the role of bacterial antizyme in the regulation of polyamine biosynthesis and gives new perspectives on the involvement of antizyme in other significant cellular mechanisms. antizyme is a protein molecule induced by the end product of the enzymic reaction that it inhibits, in a non-competitive manner. the bacterial ornithine decarboxylase is regulated by nucleotides, phosphorylation and antizyme. the inhibition of ornithine decarboxylase by antizyme can be relieved to diff ... | 2004 | 15200682 |
| a gene from the mesophilic bacterium dehalococcoides ethenogenes encodes a novel mannosylglycerate synthase. | mannosylglycerate (mg) is a common compatible solute found in thermophilic and hyperthermophilic prokaryotes. in this study we characterized a mesophilic and bifunctional mannosylglycerate synthase (mgsd) encoded in the genome of the bacterium dehalococcoides ethenogenes. mgsd encodes two domains with extensive homology to mannosyl-3-phosphoglycerate synthase (mpgs, ec 2.4.1.217) and to mannosyl-3-phosphoglycerate phosphatase (mpgp, ec 3.1.3.70), which catalyze the consecutive synthesis and deph ... | 2004 | 15205409 |
| two distinct domains of the beta subunit of aquifex aeolicus leucyl-trna synthetase are involved in trna binding as revealed by a three-hybrid selection. | the aquifex aeolicus alphabeta-leurs is the only known heterodimeric class ia aminoacyl-trna synthetase. in this study, we investigated the function of the beta subunit which is believed to bind trna(leu). a yeast three-hybrid system was constructed on the basis of the interaction of the beta subunit with its cognate trna(leu). then, seven mutated beta subunits exhibiting impaired trna binding capacities were selected out from a randomly mutated library. two mutations were identified in the clas ... | 2004 | 15208367 |
| direct evidence that a conserved arginine in ruvb aaa+ atpase acts as an allosteric effector for the atpase activity of the adjacent subunit in a hexamer. | the escherichia coli ruva and ruvb protein complex promotes branch migration of holliday junctions during recombinational repair and homologous recombination and at stalled replication forks. the ruvb protein belongs to the aaa(+) (atpase associated with various cellular activities) atpase family and forms a hexameric ring in an atp-dependent manner. studies on the oligomeric aaa(+) class atpases suggest that a conserved arginine residue is located in close proximity to the atpase site of the ad ... | 2004 | 15210950 |
| crystal structure of elongation factor p from thermus thermophilus hb8. | translation elongation factor p (ef-p) stimulates ribosomal peptidyltransferase activity. ef-p is conserved in bacteria and is essential for cell viability. eukarya and archaea have an ef-p homologue, eukaryotic initiation factor 5a (eif-5a). in the present study, we determined the crystal structure of ef-p from thermus thermophilus hb8 at a 1.65-a resolution. ef-p consists of three beta-barrel domains (i, ii, and iii), whereas eif-5a has only two domains (n and c domains). domain i of ef-p is t ... | 2004 | 15210970 |
| molecular characterization of benzimidazole resistance in helicobacter pylori. | a family of benzimidazole derivatives (bi) was shown to possess potent and selective activity against helicobacter pylori, although the precise cellular target of the bis is unknown. spontaneous h. pylori mutants were isolated as resistant to a representative bi (compound a). genomic dna was isolated from a bi-resistant mutant, transformed into a bi-sensitive strain, and found to be sufficient to confer bi resistance. the resistance determinant was localized to a 17-kb clone after screening a la ... | 2004 | 15215104 |
| the crystal structure of human endonuclease viii-like 1 (neil1) reveals a zincless finger motif required for glycosylase activity. | in prokaryotes, two dna glycosylases recognize and excise oxidized pyrimidines: endonuclease iii (nth) and endonuclease viii (nei). the oxidized purine 8-oxoguanine, on the other hand, is recognized by fpg (also known as mutm), a glycosylase that belongs to the same family as nei. the recent availability of the human genome sequence allowed the identification of three human homologs of escherichia coli nei. we report here the crystal structure of a human nei-like (neil) enzyme, neil1. the struct ... | 2004 | 15232006 |
| thermus thermophilus as a cell factory for the production of a thermophilic mn-dependent catalase which fails to be synthesized in an active form in escherichia coli. | thermostable mn-dependent catalases are promising enzymes in biotechnological applications as h(2)o(2)-detoxifying systems. we cloned the genes encoding mn-dependent catalases from thermus thermophilus hb27 and hb8 and a less thermostable mutant carrying two amino acid replacements (m129v and e293g). when the wild-type and mutant genes were overexpressed in escherichia coli, unmodified or six-his-tagged proteins of the expected size were overproduced as inactive proteins. several attempts to obt ... | 2004 | 15240253 |
| identification of a bifunctional enzyme mnmc involved in the biosynthesis of a hypermodified uridine in the wobble position of trna. | the gene encoding the bifunctional enzyme mnmc that catalyzes the two last steps in the biosynthesis of 5-methylaminomethyl-2-thiouridine (mnm5s2u) in trna has been previously mapped at about 50 min on the escherichia coli k12 chromosome, but to date the identity of the corresponding enzyme has not been correlated with any of the known open reading frames (orfs). using the protein fold-recognition approach, we predicted that the 74-kda product of the yfck orf located at 52.6 min and annotated as ... | 2004 | 15247431 |
| the phosphoenolpyruvate carboxylase from methanothermobacter thermautotrophicus has a novel structure. | in methanothermobacter thermautotrophicus, oxaloacetate synthesis is a major and essential co(2)-fixation reaction. this methanogenic archaeon possesses two oxaloacetate-synthesizing enzymes, pyruvate carboxylase and phosphoenolpyruvate carboxylase. the phosphoenolpyruvate carboxylase from this organism was purified to homogeneity. the subunit size of this homotetrameric protein was 55 kda, which is about half that of all known bacterial and eukaryotic phosphoenolpyruvate carboxylases (ppcs). th ... | 2004 | 15262949 |
| discrimination against deoxyribonucleotide substrates by bacterial rna polymerase. | nucleic acid polymerases have evolved elaborate mechanisms that prevent incorporation of the non-cognate substrates, which are distinguished by both the base and the sugar moieties. while the mechanisms of substrate selection have been studied in single-subunit dna and rna polymerases (dnaps and rnaps, respectively), the determinants of substrate binding in the multisubunit rnaps are not yet known. molecular modeling of thermus thermophilus rnap-substrate ntp complex identified a conserved beta' ... | 2004 | 15262972 |
| heteronuclear nmr investigations of dynamic regions of intact escherichia coli ribosomes. | 15n-(1)h nmr spectroscopy has been used to probe the dynamic properties of uniformly (15)n labeled escherichia coli ribosomes. despite the high molecular weight of the complex ( approximately 2.3 mda), [(1)h-(15)n] heteronuclear single-quantum correlation spectra contain approximately 100 well resolved resonances, the majority of which arise from two of the four c-terminal domains of the stalk proteins, l7/l12. heteronuclear pulse-field gradient nmr experiments show that the resonances arise fro ... | 2004 | 15263071 |
| formation and characterization of an all-ferrous rieske cluster and stabilization of the [2fe-2s]0 core by protonation. | the all-ferrous rieske cluster, [2fe-2s](0), has been produced in solution and characterized by protein-film voltammetry and uv-visible, epr, and mössbauer spectroscopies. the [2fe-2s](0) cluster, in the overexpressed soluble domain of the rieske protein from the bovine cytochrome bc(1) complex, is formed at -0.73 v at ph 7. therefore, at ph 7, the [2fe-2s](1+/0) couple is 1.0 v below the [2fe-2s](2+/1+) couple. the two cluster-bound ferrous irons are both high spin (s = 2), and they are coupled ... | 2004 | 15263097 |
| insights into the dna repair process by the formamidopyrimidine-dna glycosylase investigated by molecular dynamics. | formamidopyrimidine-dna glycosylase (fpg) identifies and removes 8-oxoguanine from dna. all of the x-ray structures of fpg complexed to an abasic site containing dna exhibit a common disordered region present in the c-terminal domain of the enzyme. however, this region is believed to be involved in the damaged base binding site when the initial protein/dna complex is formed. the dynamic behavior of the disordered polypeptide (named loop) in relation to the supposed scenario for the dna repair me ... | 2004 | 15273302 |
| staphylococcus aureus dna ligase: characterization of its kinetics of catalysis and development of a high-throughput screening compatible chemiluminescent hybridization protection assay. | dna ligases are key enzymes involved in the repair and replication of dna. prokaryotic dna ligases uniquely use nad+ as the adenylate donor during catalysis, whereas eukaryotic enzymes use atp. this difference in substrate specificity makes the bacterial enzymes potential targets for therapeutic intervention. we have developed a homogeneous chemiluminescence-based hybridization protection assay for staphylococcus aureus dna ligase that uses novel acridinium ester technology and demonstrate that ... | 2004 | 15283677 |
| a single-amino-acid lid renders a gas-tight compartment within a membrane-bound transporter. | proteins undergo structural fluctuations between nearly isoenergetic substates. such fluctuations are often intimately linked with the functional properties of proteins. however, in some cases, such as in transmembrane ion transporters, the control of the ion transport requires that the protein is designed to restrict the motions in specific regions. in this study, we have investigated the dynamics of a membrane-bound respiratory oxidase, which acts both as an enzyme catalyzing reduction of o(2) ... | 2004 | 15289603 |
| targeting the a site rna of the escherichia coli ribosomal 30 s subunit by 2'-o-methyl oligoribonucleotides: a quantitative equilibrium dialysis binding assay and differential effects of aminoglycoside antibiotics. | the bacterial ribosome comprises 30 s and 50 s ribonucleoprotein subunits, contains a number of binding sites for known antibiotics and is an attractive target for selection of novel antibacterial agents. on the 30 s subunit, for example, the a site (aminoacyl site) close to the 3'-end of 16 s rrna is highly important in the decoding process. binding by some aminoglycoside antibiotics to the a site leads to erroneous protein synthesis and is lethal for bacteria. we targeted the a site on purifie ... | 2004 | 15294017 |
| construction of a chimeric thermostable pyrophosphatase to facilitate its purification and immobilization by using the choline-binding tag. | the thermophilic inorganic pyrophosphatase (pyr) from thermus thermophilus has been produced in escherichia coli fused to the c terminus of the choline-binding tag (chb tag) derived from the choline-binding domain (chbd) of pneumococcal lyta autolysin. the chimeric chbd-pyr protein retains its thermostable activity and can be purified in a single step by deae-cellulose affinity chromatography. pyr can be further released from the chbd by thrombin, using the specific protease recognition site inc ... | 2004 | 15294797 |
| stabilizing roles of residual structure in the empty heme binding pockets and unfolded states of microsomal and mitochondrial apocytochrome b5. | the microsomal (mc) and mitochondrial (om) isoforms of mammalian cytochrome b5 are the products of different genes, which likely arose via duplication of a primordial gene and subsequent functional divergence. despite sharing essentially identical folds, heme-polypeptide interactions are stronger in om b5s than in mc b5s due to the presence of two conserved patches of hydrophobic amino acid side chains in the om heme binding pockets. this is of fundamental interest in terms of understanding heme ... | 2004 | 15295112 |
| creating ribosomes with an all-rna 30s subunit p site. | ribosome crystal structures have revealed that two small subunit proteins, s9 and s13, have c-terminal tails, which, together with several features of 16s rrna, contact the anticodon stem-loop of p-site trna. to test the functional importance of these protein tails, we created genomic deletions of the c-terminal regions of s9 and s13. all of the tail deletions, including double mutants containing deletions in both s9 and s13, were viable, showing that escherichia coli cells can synthesize all of ... | 2004 | 15308780 |
| thermus thermophilus l11 methyltransferase, prma, is dispensable for growth and preferentially modifies free ribosomal protein l11 prior to ribosome assembly. | the ribosomal protein l11 in bacteria is posttranslationally trimethylated at multiple amino acid positions by the l11 methyltransferase prma, the product of the prma gene. the role of l11 methylation in ribosome function or assembly has yet to be determined, although the deletion of escherichia coli prma has no apparent phenotype. we have constructed a mutant of the extreme thermophile thermus thermophilus in which the prma gene has been disrupted with the htk gene encoding a heat-stable kanamy ... | 2004 | 15317787 |
| substrate-induced asymmetry and channel closure revealed by the apoenzyme structure of mycobacterium tuberculosis phosphopantetheine adenylyltransferase. | phosphopantetheine adenylyltransferase (ppat) catalyzes the penultimate step in prokaryotic coenzyme a (coa) biosynthesis, directing the transfer of an adenylyl group from atp to 4'-phosphopantetheine (ppant) to yield dephospho-coa (dpcoa). the crystal structures of escherichia coli ppat bound to its substrates, product, and inhibitor revealed an allosteric hexameric enzyme with half-of-sites reactivity, and established an in-line displacement catalytic mechanism. to provide insight into the mec ... | 2004 | 15322293 |
| incidence of antibiotic resistance in campylobacter jejuni isolated in alberta, canada, from 1999 to 2002, with special reference to tet(o)-mediated tetracycline resistance. | of 203 human clinical isolates of campylobacter jejuni from alberta, canada (1999 to 2002), 101 isolates (50%) were resistant to at least 64 microg of tetracycline/ml, with four isolates exhibiting higher levels of tetracycline resistance (512 microg/ml). in total, the mics for 37% of tetracycline-resistant isolates (256 to 512 microg/ml) were higher than those previously reported in c. jejuni (64 to 128 microg/ml). in the tetracycline-resistant clinical isolates, 67% contained plasmids and all ... | 2004 | 15328109 |
| dna ligases ensure fidelity by interrogating minor groove contacts. | dna ligases, found in both prokaryotes and eukaryotes, covalently link the 3'-hydroxyl and 5'-phosphate ends of duplex dna segments. this reaction represents a completion step for dna replication, repair and recombination. it is well established that ligases are sensitive to mispairs present on the 3' side of the ligase junction, but tolerant of mispairs on the 5' side. while such discrimination would increase the overall accuracy of dna replication and repair, the mechanisms by which this fidel ... | 2004 | 15328364 |
| photolabile anticodon stem-loop analogs of trnaphe as probes of ribosomal structure and structural fluctuation at the decoding center. | with the recent availability of high-resolution structures of bacterial ribosomes, studies of ribosome-catalyzed protein biosynthesis are now focusing on the nature of conformational changes that occur as the ribosome exerts its complex catalytic function. photocrosslinking can be relevant for this purpose by providing clues to ribosomal structural fluctuations and dynamics. here we describe crosslinking experiments on 70s ribosomes using two photolabile anticodon stem-loop derivatives of escher ... | 2004 | 15337844 |
| crystal structure of ribosomal protein l27 from thermus thermophilus hb8. | ribosomal protein l27 is located near the peptidyltransferase center at the interface of ribosomal subunits, and is important for ribosomal assembly and function. we report the crystal structure of ribosomal protein l27 from thermus thermophilus hb8, which was determined by the multiwavelength anomalous dispersion method and refined to an r-factor of 19.7% (r(free) = 23.6%) at 2.8 a resolution. the overall fold is an all beta-sheet hybrid. it consists of two sets of four-stranded beta-sheets for ... | 2004 | 15340170 |
| reverse transcriptase activity innate to dna polymerase i and dna topoisomerase i proteins of streptomyces telomere complex. | replication of streptomyces linear chromosomes and plasmids proceeds bidirectionally from a central origin, leaving recessed 5' termini that are extended by a telomere binding complex. this complex contains both a telomere-protecting terminal protein (tpg) and a telomere-associated protein that interacts with tpg and the dna ends of linear streptomyces replicons. by using histidine-tagged telomere-associated protein (tap) as a scaffold, we identified dna polymerase (pola) and topoisomerase i (to ... | 2004 | 15353591 |
| biochemical characterization of cdc6/orc1 binding to the replication origin of the euryarchaeon methanothermobacter thermoautotrophicus. | archaeal cell division cycle protein 6 (cdc6)/origin replication complex subunit 1 (orc1) proteins share sequence homology with eukaryotic dna replication initiation factors but are also structurally similar to the bacterial initiator dnaa. to better understand whether cdc6/orc1 functions in an eukaryotic or bacterial-like manner, we have characterized the interaction of two cdc6/orc1 paralogs (mthcdc6-1 and mthcdc6-2) with the replication origin from methanothermobacter thermoautotrophicus. we ... | 2004 | 15358831 |
| reaction cycle of the yeast isw2 chromatin remodeling complex. | members of the iswi family of chromatin remodeling factors hydrolyze atp to reposition nucleosomes along dna. here we show that the yeast isw2 complex interacts with dna in a nucleotide-dependent manner at physiological ionic strength. isw2 efficiently binds dna in the absence of nucleotides and in the presence of a nonhydrolyzable atp analog. conversely, adp promotes the dissociation of isw2 from dna. in contrast, isw2 remains bound to mononucleosomes through multiple cycles of atp hydrolysis. ... | 2004 | 15359274 |
| structure and dna-binding properties of the cytolysin regulator cylr2 from enterococcus faecalis. | enterococcus faecalis is one of the major causes for hospital-acquired antibiotic-resistant infections. it produces an exotoxin, called cytolysin, which is lethal for a wide range of gram-positive bacteria and is toxic to higher organisms. recently, the regulation of the cytolysin operon was connected to autoinduction by a quorum-sensing mechanism involving the cylr1/cylr2 two-component regulatory system. we report here the crystal structure of cylr2 and its properties in solution as determined ... | 2004 | 15359276 |
| theoretical identification of proton channels in the quinol oxidase aa3 from acidianus ambivalens. | heme-copper oxidases are membrane proteins found in the respiratory chain of aerobic organisms. they are the terminal electron acceptors coupling the translocation of protons across the membrane with the reduction of oxygen to water. because the catalytic process occurs in the heme cofactors positioned well inside the protein matrix, proton channels must exist. however, due to the high structural divergence among this kind of proteins, the proton channels previously described are not necessarily ... | 2004 | 15377522 |
| liver portal fibrosis in dioxin receptor-null mice that overexpress the latent transforming growth factor-beta-binding protein-1. | mice lacking aryl hydrocarbon (dioxin) receptor (ahr) had variable degree of hepatic fibrosis and altered liver architecture. transforming growth factor-beta (tgf-beta), a major profibrogenic molecule in the liver, is localized to the extracellular matrix by its association to the latent tgf-beta-binding protein-1 (ltbp-1). very recently, ltbp-1 has been shown to be negatively regulated by the ahr. embryonic fibroblasts from ahr-null (ahr(-/-)) mice overexpress ltbp-1 and secrete four times more ... | 2004 | 15379962 |
| efficient stimulation of site-specific ribosome frameshifting by antisense oligonucleotides. | evidence is presented that morpholino, 2'-o-methyl, phosphorothioate, and rna antisense oligonucleotides can direct site-specific -1 translational frameshifting when annealed to mrna downstream from sequences where the p- and a-site trnas are both capable of repairing with -1 frame codons. the efficiency of ribosomes shifting into the new frame can be as high as 40%, determined by the sequence of the frameshift site, as well as the location, sequence composition, and modification of the antisens ... | 2004 | 15383681 |
| selectivity and specificity of substrate binding in methionyl-trna synthetase. | the accuracy of in vivo incorporation of amino acids during protein biosynthesis is controlled to a significant extent by aminoacyl-trna synthetases (aars). this paper describes the application of the hierdock computational method to study the molecular basis of amino acid binding to the escherichia coli methionyl trna synthetase (metrs). starting with the protein structure from the metrs cocrystal, the hierdock calculations predict the binding site of methionine in metrs to a root mean square d ... | 2004 | 15388861 |
| atomic model of the thermus thermophilus 70s ribosome developed in silico. | the ribosome is a large molecular complex that consists of at least three ribonucleic acid molecules and a large number of proteins. it translates genetic information from messenger ribonucleic acid and makes protein accordingly. to better understand ribosomal function and provide information for designing biochemical experiments require knowledge of the complete structure of the ribosome. for expanding the structural information of the ribosome, we took on the challenge of developing a detailed ... | 2004 | 15454463 |
| crystal structures of possible lysine decarboxylases from thermus thermophilus hb8. | tt1887 and tt1465 from thermus thermophilus hb8 are conserved hypothetical proteins, and are annotated as possible lysine decarboxylases in the pfam database. here we report the crystal structures of tt1887 and tt1465 at 1.8 a and 2.2 a resolutions, respectively, as determined by the multiwavelength anomalous dispersion (mad) method. tt1887 is a homotetramer, while tt1465 is a homohexamer in the crystal and in solution. the structures of the tt1887 and tt1465 monomers contain single domains with ... | 2004 | 15459330 |
| complete genome sequence of the genetically tractable hydrogenotrophic methanogen methanococcus maripaludis. | the genome sequence of the genetically tractable, mesophilic, hydrogenotrophic methanogen methanococcus maripaludis contains 1,722 protein-coding genes in a single circular chromosome of 1,661,137 bp. of the protein-coding genes (open reading frames [orfs]), 44% were assigned a function, 48% were conserved but had unknown or uncertain functions, and 7.5% (129 orfs) were unique to m. maripaludis. of the unique orfs, 27 were confirmed to encode proteins by the mass spectrometric identification of ... | 2004 | 15466049 |
| transcriptional analysis of biofilm formation processes in the anaerobic, hyperthermophilic bacterium thermotoga maritima. | thermotoga maritima, a fermentative, anaerobic, hyperthermophilic bacterium, was found to attach to bioreactor glass walls, nylon mesh, and polycarbonate filters during chemostat cultivation on maltose-based media at 80 degrees c. a whole-genome cdna microarray was used to examine differential expression patterns between biofilm and planktonic populations. mixed-model statistical analysis revealed differential expression (twofold or more) of 114 open reading frames in sessile cells (6% of the ge ... | 2004 | 15466556 |
| comparative genomics of the ftsk-hera superfamily of pumping atpases: implications for the origins of chromosome segregation, cell division and viral capsid packaging. | recently, it has been shown that a predicted p-loop atpase (the hera or mlaa protein), which is highly conserved in archaea and also present in many bacteria but absent in eukaryotes, has a bidirectional helicase activity and forms hexameric rings similar to those described for the trwb atpase. in this study, the ftsk-hera superfamily of p-loop atpases, in which the hera clade comprises one of the major branches, is analyzed in detail. we show that, in addition to the ftsk and hera clades, this ... | 2004 | 15466593 |
| interaction of mitochondrial initiation factor 2 with mitochondrial fmet-trna. | the mammalian mitochondrial genome contains a single trna(met) gene that gives rise to the initiator and elongator trna(met). it is generally believed that mitochondrial protein synthesis begins with formylmethionyl-trna, which indicates that the formylation of mitochondrial met-trna specifies its participation in initiation through its interaction with initiation factor 2 (if-2). however, recent studies in yeast mitochondria, suggest that formylation is not required for protein synthesis. in ad ... | 2004 | 15477394 |
| the a2453-c2499 wobble base pair in escherichia coli 23s ribosomal rna is responsible for ph sensitivity of the peptidyltransferase active site conformation. | peptide bond formation, catalyzed by the ribosomal peptidyltransferase, has long been known to be sensitive to monovalent cation concentrations and ph. more recently, we and others have shown that residue a2451 in the peptidyltransferase center of the escherichia coli 50s ribosomal subunit changes conformation in response to alterations in ph, depending on ionic conditions and temperature. two wobble pairs, a2453-c2499 and a2450-c2063, have been proposed as potential candidates to convey ph-depe ... | 2004 | 15479786 |
| human mitochondrial peptide deformylase, a new anticancer target of actinonin-based antibiotics. | peptide deformylase activity was thought to be limited to ribosomal protein synthesis in prokaryotes, where new peptides are initiated with an n-formylated methionine. we describe here a new human peptide deformylase (homo sapiens pdf, or hspdf) that is localized to the mitochondria. hspdf is capable of removing formyl groups from n-terminal methionines of newly synthesized mitochondrial proteins, an activity previously not thought to be necessary in mammalian cells. we show that actinonin, a pe ... | 2004 | 15489958 |
| the prc-barrel domain of the ribosome maturation protein rimm mediates binding to ribosomal protein s19 in the 30s ribosomal subunits. | the rimm protein in escherichia coli is associated with free 30s ribosomal subunits but not with 70s ribosomes. a deltarimm mutant is defective in 30s maturation and accumulates 17s rrna. to study the interaction of rimm with the 30s and its involvement in 30s maturation, rimm amino acid substitution mutants were constructed. a mutant rimm (rimm-yy-->aa), containing alanine substitutions for two adjacent tyrosines within the prc beta-barrel domain, showed a reduced binding to 30s and an accumula ... | 2004 | 15496525 |
| maturation of the unusual single-cysteine (xxxch) mitochondrial c-type cytochromes found in trypanosomatids must occur through a novel biogenesis pathway. | the c-type cytochromes are characterized by the covalent attachment of haem to the polypeptide via thioether bonds formed from haem vinyl groups and, normally, the thiols of two cysteines in a cxxch motif. intriguingly, the mitochondrial cytochromes c and c1 from two euglenids and the trypanosomatidae contain only a single cysteine within the haem-binding motif (xxxch). there are three known distinct pathways by which c-type cytochromes are matured post-translationally in different organisms. th ... | 2004 | 15500440 |
| fluoroquinolone resistance in penicillin-resistant streptococcus pneumoniae clones, spain. | among 2,882 streptococcus pneumoniae sent to the spanish reference laboratory during 2002, 75 (2.6%) were ciprofloxacin-resistant. resistance was associated with older patients (3.9% in adults and 7.2% in patients > or =65 years of age), with isolation from noninvasive sites (4.3% vs. 1.0%), and with penicillin and macrolide resistance. among 14 low-level resistant (mic 4-8 microg/ml) strains, 1 had a fluoroquinolone efflux phenotype, and 13 showed single parc changes. the 61 high-level ciproflo ... | 2004 | 15504260 |
| nosocomial acquisition of dengue. | recent transmission of dengue viruses has increased in tropical and subtropical areas and in industrialized countries because of international travel. we describe a case of nosocomial transmission of dengue virus in germany by a needlestick injury. diagnosis was made by taqman reverse transcription-polymerase chain reaction when serologic studies were negative. | 2004 | 15504282 |
| analysis of mupirocin resistance and fitness in staphylococcus aureus by molecular genetic and structural modeling techniques. | chromosomal resistance to mupirocin in clinical isolates of staphylococcus aureus arises from v(588)f or v(631)f mutations in isoleucyl-trna synthetase (irs). whether these are the only irs mutations that confer mupirocin resistance or simply those that survive in the clinic is unknown. mupirocin-resistant mutants of s. aureus 8325-4 were therefore generated to examine their iles genotypes and the in vitro and in vivo fitness costs associated with them before and after compensatory evolution. mo ... | 2004 | 15504866 |
| guanosine tetra- and pentaphosphate synthase activity in chloroplasts of a higher plant: association with 70s ribosomes and inhibition by tetracycline. | chloroplasts possess bacterial-type systems for transcription and translation. on the basis of the identification of a chlamydomonas reinhardtii gene encoding a rela-spot homolog (rsh) that catalyzes the synthesis of guanosine tetra- or pentaphosphate [(p)ppgpp], we have previously suggested the operation of stringent control in the chloroplast genetic system. although rsh genes have also been identified in several higher plants, the activities of the encoded enzymes and their mode of action in ... | 2004 | 15507686 |
| multiple defects in translation associated with altered ribosomal protein l4. | the ribosomal proteins l4 and l22 form part of the peptide exit tunnel in the large ribosomal subunit. in escherichia coli, alterations in either of these proteins can confer resistance to the macrolide antibiotic, erythromycin. the structures of the 30s as well as the 50s subunits from each antibiotic resistant mutant differ from wild type in distinct ways and l4 mutant ribosomes have decreased peptide bond-forming activity. our analyses of the decoding properties of both mutants show that ribo ... | 2004 | 15509870 |
| a novel archaeal alanine dehydrogenase homologous to ornithine cyclodeaminase and mu-crystallin. | a novel alanine dehydrogenase (aladh) showing no significant amino acid sequence homology with previously known bacterial aladhs was purified to homogeneity from the soluble fraction of the hyperthermophilic archaeon archaeoglobus fulgidus. aladh catalyzed the reversible, nad+-dependent deamination of l-alanine to pyruvate and nh4+. nadp(h) did not serve as a coenzyme. the enzyme is a homodimer of 35 kda per subunit. the km values for l-alanine, nad+, pyruvate, nadh, and nh4+ were estimated at 0 ... | 2004 | 15516582 |
| identification and functional verification of archaeal-type phosphoenolpyruvate carboxylase, a missing link in archaeal central carbohydrate metabolism. | despite the fact that phosphoenolpyruvate carboxylase (pepc) activity has been measured and in some cases even purified from some archaea, the gene responsible for this activity has not been elucidated. using sensitive sequence comparison methods, we detected a highly conserved, uncharacterized archaeal gene family that is distantly related to the catalytic core of the canonical pepc. to verify the predicted function of this archaeal gene family, we cloned a representative from the hyperthermoph ... | 2004 | 15516590 |
| use of an antisense rna strategy to investigate the functional significance of mn-catalase in the extreme thermophile thermus thermophilus. | the expression of an antisense rna revealed that an mn-catalase was required in thermus thermophilus for aerobic but not for anaerobic growth. the antisense system is based on the constitutive expression of a "bicistronic" transcript consisting of the kanamycin resistance gene mrna followed by the antisense rna against the selected target. | 2004 | 15516595 |
| cold sensitivity of thermophilic and mesophilic rna polymerases. | rna polymerase from mesophilic deinococcus radiodurans displays the same cold sensitivity of promoter opening as rna polymerase from the closely related thermophilic thermus aquaticus. this suggests that, contrary to the accepted view, cold sensitivity of promoter opening by thermophilic rna polymerases may not be a consequence of their thermostability. | 2004 | 15516599 |
| cleavage of double-stranded rna by rnase hi from a thermoacidophilic archaeon, sulfolobus tokodaii 7. | st0753, the orthologous gene of type 1 rnase h found in a thermoacidophilic archaeon, sulfolobus tokodaii, was analyzed. the recombinant st0753 protein exhibited rnase h activity in both in vivo and in vitro assays. the protein expressed in an rnase h-deficient mutant escherichia coli strain functioned to suppress the temperature-sensitive phenotype associated with the lack of rnase h. the in vitro characteristics of the gene's rnase h activity were similar to those of halobacterium rnase hi, th ... | 2004 | 15520465 |
| assembly of the 30s ribosomal subunit: positioning ribosomal protein s13 in the s7 assembly branch. | studies of escherichia coli 30s ribosomal subunit assembly have revealed a hierarchical and cooperative association of ribosomal proteins with 16s ribosomal rna; these results have been used to compile an in vitro 30s subunit assembly map. in single protein addition and omission studies, ribosomal protein s13 was shown to be dependent on the prior association of ribosomal protein s20 for binding to the ribonucleoprotein particle. while the overwhelming majority of interactions revealed in the as ... | 2004 | 15525707 |
| post-transfer editing in vitro and in vivo by the beta subunit of phenylalanyl-trna synthetase. | translation of the genetic code requires attachment of trnas to their cognate amino acids. errors during amino-acid activation and trna esterification are corrected by aminoacyl-trna synthetase-catalyzed editing reactions, as extensively described for aliphatic amino acids. the contribution of editing to aromatic amino-acid discrimination is less well understood. we show that phenylalanyl-trna synthetase misactivates tyrosine and that it subsequently corrects such errors through hydrolysis of ty ... | 2004 | 15526031 |
| testing the conservation of the translational machinery over evolution in diverse environments: assaying thermus thermophilus ribosomes and initiation factors in a coupled transcription-translation system from escherichia coli. | ribosomes from the extreme thermophile thermus thermophilus are capable of translation in a coupled transcription-translation system derived from escherichia coli. at 45 degrees c, t.thermophilus ribosomes translate at approximately 25-30% of the maximal rate of e.coli ribosomes, and synthesize full-length protein. t.thermophilus and e.coli subunits can be combined to effect translation, with the spectrum of proteins produced depending upon the source of the 30s subunit. in this system, t.thermo ... | 2004 | 15534366 |
| the bag or the spindle: the cell factory at the time of systems' biology. | genome programs changed our view of bacteria as cell factories, by making them amenable to systematic rational improvement. as a first step, isolated genes (including those of the metagenome), or small gene clusters are improved and expressed in a variety of hosts. new techniques derived from functional genomics (transcriptome, proteome and metabolome studies) now allow users to shift from this single-gene approach to a more integrated view of the cell, where it is more and more considered as a ... | 2004 | 15537427 |
| interaction of the n-terminal domain of escherichia coli heat-shock protein clpb and protein aggregates during chaperone activity. | the escherichia coli heat-shock protein clpb reactivates protein aggregates in cooperation with the dnak chaperone system. the clpb n-terminal domain plays an important role in the chaperone activity, but its mechanism remains unknown. in this study, we investigated the effect of the clpb n-terminal domain on malate dehydrogenase (mdh) refolding. clpb reduced the yield of mdh refolding by a strong interaction with the intermediate. however, the refolding kinetics was not affected by deletion of ... | 2004 | 15537752 |
| crystal structure of yeast v-atpase subunit c reveals its stator function. | vacuolar h(+)-atpase (v-atpase) has a crucial role in the vacuolar system of eukaryotic cells. it provides most of the energy required for transport systems that utilize the proton-motive force that is generated by atp hydrolysis. some, but not all, of the v-atpase subunits are homologous to those of f-atpase and the nonhomologous subunits determine the unique features of v-atpase. we determined the crystal structure of v-atpase subunit c (vma5p), which does not show any homology with f-atpase s ... | 2004 | 15540116 |
| artificial tertiary motifs stabilize trans-cleaving hammerhead ribozymes under conditions of submillimolar divalent ions and high temperatures. | tertiary stabilizing motifs (tsms) between terminal loops or internal bulges facilitate folding of natural hammerhead ribozymes (hrz) under physiological conditions. however, both substrate and enzyme strands contribute nucleotides to the tsms of trans-cleaving hrz, complicating the design of hrz that exploit tsms to target specific mrna. to overcome this limitation, we used selex to identify new, artificial tsms that are less sensitive to sequence context. nucleotides in loop ii or in a bulge w ... | 2004 | 15547137 |
| two outer membrane proteins are required for maximal type i secretion of the caulobacter crescentus s-layer protein. | transport of rsaa, the crystalline s-layer subunit protein of caulobacter crescentus, is mediated by a type i secretion mechanism. two proteins have been identified that play the role of the outer membrane protein (omp) component in the rsaa secretion machinery. the genes rsaf(a) and rsaf(b) were identified by similarity to the escherichia coli hemolysin secretion omp tolc by using the c. crescentus genome sequence. the rsaf(a) gene is located several kilobases downstream of the other transporte ... | 2004 | 15547272 |
| divergent anticodon recognition in contrasting glutamyl-trna synthetases. | the pathogenic bacterium helicobacter pylori utilizes two essential glutamyl-trna synthetases (glurs1 and glurs2). these two enzymes are closely related in evolution and yet they aminoacylate contrasting trnas. glurs1 is a canonical discriminating glurs (d-glurs) that biosynthesizes glu-trna(glu) and cannot make glu-trna(gln). in contrast, glurs2 is non-canonical as it is only essential for the production of misacylated glu-trna(gln). the co-existence and evident divergence of these two enzymes ... | 2004 | 15561136 |
| cloning of cvipii nicking and modification system from chlorella virus nys-1 and application of nt.cvipii in random dna amplification. | the cloning and expression of the cvipii dna nicking and modification system encoded by chlorella virus nys-1 is described. the system consists of a co-linear mtase encoding gene (cvipiim) and a nicking endonuclease encoding gene (cvipiint) separated by 12 nt. m.cvipii possesses eight conserved amino acid motifs (i to viii) typical of c5 mtases, but, like another chlorella virus mtase m.cviji, lacks conserved motifs ix and x. in addition to modification of the first cytosine in ccd (d = a, g or ... | 2004 | 15570069 |
| mechanistic approach to the problem of hybridization efficiency in fluorescent in situ hybridization. | in fluorescent in situ hybridization (fish), the efficiency of hybridization between the dna probe and the rrna has been related to the accessibility of the rrna when ribosome content and cell permeability are not limiting. published rrna accessibility maps show that probe brightness is sensitive to the organism being hybridized and the exact location of the target site and, hence, it is highly unpredictable based on accessibility only. in this study, a model of fish based on the thermodynamics ... | 2004 | 15574909 |
| characterization of a novel amylolytic enzyme encoded by a gene from a soil-derived metagenomic library. | it has been estimated that less than 1% of the microorganisms in nature can be cultivated by conventional techniques. thus, the classical approach of isolating enzymes from pure cultures allows the analysis of only a subset of the total naturally occurring microbiota in environmental samples enriched in microorganisms. to isolate useful microbial enzymes from uncultured soil microorganisms, a metagenome was isolated from soil samples, and a metagenomic library was constructed by using the puc19 ... | 2004 | 15574921 |