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initiation factor eif2γ promotes eif2-gtp-met-trnai(met) ternary complex binding to the 40s ribosome.in contrast to prokaryotic elongation factor ef-tu, which delivers aminoacyl-trnas to the ribosomal a-site, eukaryotic initiation factor eif2 binds methionyl initiator transfer rna (met-trna(i)(met)) to the p-site of the 40s ribosomal subunit. the results of directed hydroxyl radical probing experiments to map binding of saccharomyces cerevisiae eif2 on the ribosome and on met-trna(i)(met) revealed that eif2γ primarily contacts the acceptor stem of met-trna(i)(met) and identified a key binding i ...201122002225
"stealth" melanoma cells in histology-negative sentinel lymph nodes.a proportion of patients who develop regional and distant recurrences of melanoma after a pathologically negative sentinel lymph node (sn) biopsy are reported to have enhanced signals for melanoma-associated messenger ribonucleic acid (mrna) when sensitive molecular approaches such as reverse transcriptase polymerase chain reaction (rt-pcr) are used to evaluate their sn tissue. the significance of these findings remains controversial, because the cellular source of the augmented signals cannot b ...201121997686
Biosynthesis of the Pseudomonas aeruginosa Extracellular Polysaccharides, Alginate, Pel, and Psl.Pseudomonas aeruginosa thrives in many aqueous environments and is an opportunistic pathogen that can cause both acute and chronic infections. Environmental conditions and host defenses cause differing stresses on the bacteria, and to survive in vastly different environments, P. aeruginosa must be able to adapt to its surroundings. One strategy for bacterial adaptation is to self-encapsulate with matrix material, primarily composed of secreted extracellular polysaccharides. P. aeruginosa has the ...201121991261
in vivo, in vitro, and x-ray crystallographic analyses suggest the involvement of an uncharacterized triose-phosphate isomerase (tim) barrel protein in protection against oxidative stress.accumulating genome sequences have revealed the existence of a large number of conserved hypothetical proteins. characterization of these proteins is considered essential in the elucidation of intracellular biological pathways. our previous transcriptomic analysis suggested that, in thermus thermophilus hb8, loss of an oxidized dna-repairing activity leads to the up-regulation of a function-unknown gene, tthb071, which is conserved in a wide range of bacteria. interestingly, the tthb071 gene pro ...201121984829
residual structure in unfolded proteins.the denatured state ensemble (dse) of unfolded proteins, once considered to be well-modeled by an energetically featureless random coil, is now well-known to contain flickering elements of residual structure. the position and nature of dse residual structure may provide clues toward deciphering the protein folding code. this review focuses on recent advances in our understanding of the nature of dse collapse under folding conditions, the quantification of the stability of residual structure in t ...201121978577
residual structure in unfolded proteins.the denatured state ensemble (dse) of unfolded proteins, once considered to be well-modeled by an energetically featureless random coil, is now well-known to contain flickering elements of residual structure. the position and nature of dse residual structure may provide clues toward deciphering the protein folding code. this review focuses on recent advances in our understanding of the nature of dse collapse under folding conditions, the quantification of the stability of residual structure in t ...201121978577
antimutator variants of dna polymerases.evolution balances dna replication speed and accuracy to optimize replicative fitness and genetic stability. there is no selective pressure to improve dna replication fidelity beyond the background mutation rate from other sources, such as dna damage. however, dna polymerases remain amenable to amino acid substitutions that lower intrinsic error rates. here, we review these 'antimutagenic' changes in dna polymerases and discuss what they reveal about mechanisms of replication fidelity. pioneerin ...201121977975
clustering rna structural motifs in ribosomal rnas using secondary structural alignment.rna structural motifs are the building blocks of the complex rna architecture. identification of non-coding rna structural motifs is a critical step towards understanding of their structures and functionalities. in this article, we present a clustering approach for de novo rna structural motif identification. we applied our approach on a data set containing 5s, 16s and 23s rrnas and rediscovered many known motifs including gnra tetraloop, kink-turn, c-loop, sarcin-ricin, reverse kink-turn, hook- ...201121976732
clustering rna structural motifs in ribosomal rnas using secondary structural alignment.rna structural motifs are the building blocks of the complex rna architecture. identification of non-coding rna structural motifs is a critical step towards understanding of their structures and functionalities. in this article, we present a clustering approach for de novo rna structural motif identification. we applied our approach on a data set containing 5s, 16s and 23s rrnas and rediscovered many known motifs including gnra tetraloop, kink-turn, c-loop, sarcin-ricin, reverse kink-turn, hook- ...201121976732
tagetitoxin inhibits rna polymerase through trapping of the trigger loop.tagetitoxin (tgt) inhibits multisubunit chloroplast, bacterial, and some eukaryotic rna polymerases (rnaps). a crystallographic structure of tgt bound to bacterial rnap apoenzyme shows that tgt binds near the active site but does not explain why tgt acts only at certain sites. to understand the tgt mechanism, we constructed a structural model of tgt bound to the transcription elongation complex. in this model, tgt interacts with the β' subunit trigger loop (tl), stabilizing it in an inactive con ...201121976682
massive multiplication of genome and ribosomes in dormant cells (akinetes) of aphanizomenon ovalisporum (cyanobacteria).akinetes are dormancy cells commonly found among filamentous cyanobacteria, many of which are toxic and/or nuisance, bloom-forming species. development of akinetes from vegetative cells is a process that involves morphological and biochemical modifications. here, we applied a single-cell approach to quantify genome and ribosome content of akinetes and vegetative cells in aphanizomenon ovalisporum (cyanobacteria). vegetative cells of a. ovalisporum were naturally polyploid and contained, on avera ...201121975597
massive multiplication of genome and ribosomes in dormant cells (akinetes) of aphanizomenon ovalisporum (cyanobacteria).akinetes are dormancy cells commonly found among filamentous cyanobacteria, many of which are toxic and/or nuisance, bloom-forming species. development of akinetes from vegetative cells is a process that involves morphological and biochemical modifications. here, we applied a single-cell approach to quantify genome and ribosome content of akinetes and vegetative cells in aphanizomenon ovalisporum (cyanobacteria). vegetative cells of a. ovalisporum were naturally polyploid and contained, on avera ...201121975597
thermophilic proteins: insight and perspective from in silico experiments.proteins from thermophilic and hyperthermophilic organisms are stable and function at high temperatures (50-100 °c). the importance of understanding the microscopic mechanisms underlying this thermal resistance is twofold: it is key for acquiring general clues on how proteins maintain their fold stable and for targeting those medical and industrial applications that aim at designing enzymes that can work under harsh conditions. in this tutorial review we first provide the general background of p ...201121975514
thermophilic proteins: insight and perspective from in silico experiments.proteins from thermophilic and hyperthermophilic organisms are stable and function at high temperatures (50-100 °c). the importance of understanding the microscopic mechanisms underlying this thermal resistance is twofold: it is key for acquiring general clues on how proteins maintain their fold stable and for targeting those medical and industrial applications that aim at designing enzymes that can work under harsh conditions. in this tutorial review we first provide the general background of p ...201121975514
bacterial communities of diverse drosophila species: ecological context of a host-microbe model system.drosophila melanogaster is emerging as an important model of non-pathogenic host-microbe interactions. the genetic and experimental tractability of drosophila has led to significant gains in our understanding of animal-microbial symbiosis. however, the full implications of these results cannot be appreciated without the knowledge of the microbial communities associated with natural drosophila populations. in particular, it is not clear whether laboratory cultures can serve as an accurate model o ...201121966276
protective role of catechin on d-galactosamine induced hepatotoxicity through a p53 dependent pathway.objective of this study was to obtain a better understanding of the mechanism responsible for the d-galactosamine (d-galn) induced hepatotoxicity and to study the effect of catechin against d-galn induced hepatotoxicity. catechin 50 and 100 mg/kg b.wt was administered for 1 week by oral route. liver damage was induced by intra-peritoneal administration of 400 mg/kg b.wt d-galactosamine on the last day of catechin treatment. at the end of treatment all animals were killed and liver enzyme levels ...201021966103
ucsf chimera, modeller, and imp: an integrated modeling system.structural modeling of macromolecular complexes greatly benefits from interactive visualization capabilities. here we present the integration of several modeling tools into ucsf chimera. these include comparative modeling by modeller, simultaneous fitting of multiple components into electron microscopy density maps by imp multifit, computing of small-angle x-ray scattering profiles and fitting of the corresponding experimental profile by imp foxs, and assessment of amino acid sidechain conformat ...201121963794
ucsf chimera, modeller, and imp: an integrated modeling system.structural modeling of macromolecular complexes greatly benefits from interactive visualization capabilities. here we present the integration of several modeling tools into ucsf chimera. these include comparative modeling by modeller, simultaneous fitting of multiple components into electron microscopy density maps by imp multifit, computing of small-angle x-ray scattering profiles and fitting of the corresponding experimental profile by imp foxs, and assessment of amino acid sidechain conformat ...201121963794
a mycobacterium leprae hsp65 mutant as a candidate for mitigating lupus aggravation in mice.hsp60 is an abundant and highly conserved family of intracellular molecules. increased levels of this family of proteins have been observed in the extracellular compartment in chronic inflammation. administration of m. leprae hsp65 [wt] in [nzbxnzw]f(1) mice accelerates the systemic lupus erythematosus [sle] progression whereas the point mutated k(409)a hsp65 protein delays the disease. here, the biological effects of m. leprae hsp65 leader pep and k(409)a pep synthetic peptides, which cover res ...201121961033
evidence for atp-dependent structural rearrangement of nuclease catalytic site in dna mismatch repair endonuclease mutl.dna mismatch repair (mmr) greatly contributes to genome integrity via the correction of mismatched bases that are mainly generated by replication errors. postreplicative mmr excises a relatively long tract of error-containing single-stranded dna. mutl is a widely conserved nicking endonuclease that directs the excision reaction to the error-containing strand of the duplex by specifically nicking the daughter strand. because mutl apparently exhibits nonspecific nicking endonuclease activity in vi ...201121953455
mass spectrometry-based quantification of pseudouridine in rna.direct detection of pseudouridine (ψ), an isomer of uridine, in rna is challenging. the most popular method requires chemical derivatization using n-cyclohexyl-n'-β-(4-methylmorpholinum ethyl) carbodiimide p-tosylate (cmct) followed by radiolabeled primer extension mediated by reverse transcriptase. more recently, mass spectrometry (ms)-based approaches for sequence placement of pseudouridine in rna have been developed. nearly all of these approaches, however, only yield qualitative information ...201121953190
structure and molecular evolution of cdgsh iron-sulfur domains.the recently discovered cdgsh iron-sulfur domains (cisds) are classified into seven major types with a wide distribution throughout the three domains of life. the type 1 protein mitoneet has been shown to fold into a dimer with the signature cdgsh motif binding to a [2fe-2s] cluster. however, the structures of all other types of cisds were unknown. here we report the crystal structures of type 3, 4, and 6 cisds determined at 1.5 å, 1.8 å and 1.15 å resolution, respectively. the type 3 and 4 cisd ...201121949752
Functional characterization of the RuvB homologs from Mycoplasma pneumoniae and Mycoplasma genitalium.Homologous recombination between repeated DNA elements in the genomes of Mycoplasma species has been hypothesized to be a crucial causal factor in sequence variation of antigenic proteins at the bacterial surface. To investigate this notion, studies were initiated to identify and characterize the proteins that form part of the homologous DNA recombination machinery in Mycoplasma pneumoniae as well as Mycoplasma genitalium. Among the most likely participants of this machinery are homologs of the ...201121949077
Systematic chromosomal deletion of bacterial ribosomal protein genes.Detailed studies of ribosomal proteins (RPs), essential components of the protein biosynthetic machinery, have been hampered by the lack of readily accessible chromosomal deletions of the corresponding genes. Here, we report the systematic genomic deletion of 41 individual RP genes in Escherichia coli, which are not included in the Keio collection. Chromosomal copies of these genes were replaced by an antibiotic resistance gene in the presence of an inducible, easy-to-exchange plasmid-born allel ...201121945294
enzymedetector: an integrated enzyme function prediction tool and database.the ability to accurately predict enzymatic functions is an essential prerequisite for the interpretation of cellular functions, and the reconstruction and analysis of metabolic models. several biological databases exist that provide such information. however, in many cases these databases provide partly different and inconsistent genome annotations.201121943292
Tilt-pair analysis of images from a range of different specimens in single-particle electron cryomicroscopy.The comparison of a pair of electron microscope images recorded at different specimen tilt angles provides a powerful approach for evaluating the quality of images, image-processing procedures, or three-dimensional structures. Here, we analyze tilt-pair images recorded from a range of specimens with different symmetries and molecular masses and show how the analysis can produce valuable information not easily obtained otherwise. We show that the accuracy of orientation determination of individua ...201121939668
structures of the rna-guided surveillance complex from a bacterial immune system.bacteria and archaea acquire resistance to viruses and plasmids by integrating short fragments of foreign dna into clustered regularly interspaced short palindromic repeats (crisprs). these repetitive loci maintain a genetic record of all prior encounters with foreign transgressors. crisprs are transcribed and the long primary transcript is processed into a library of short crispr-derived rnas (crrnas) that contain a unique sequence complementary to a foreign nucleic-acid challenger. in escheric ...201121938068
pseudomonas aeruginosa 4-amino-4-deoxychorismate lyase: spatial conservation of an active site tyrosine and classification of two types of enzyme.4-amino-4-deoxychorismate lyase (pabc) catalyzes the formation of 4-aminobenzoate, and release of pyruvate, during folate biosynthesis. this is an essential activity for the growth of gram-negative bacteria, including important pathogens such as pseudomonas aeruginosa. a high-resolution (1.75 å) crystal structure of pabc from p. aeruginosa has been determined, and sequence-structure comparisons with orthologous structures are reported. residues around the pyridoxal 5'-phosphate cofactor are high ...201121935381
a novel mechanism of sulfur transfer catalyzed by o-acetylhomoserine sulfhydrylase in the methionine-biosynthetic pathway of wolinella succinogenes.o-acetylhomoserine sulfhydrylase (oahs) is a pyridoxal 5'-phosphate (plp) dependent sulfide-utilizing enzyme in the l-cysteine and l-methionine biosynthetic pathways of various enteric bacteria and fungi. oahs catalyzes the conversion of o-acetylhomoserine to homocysteine using sulfide in a process known as direct sulfhydrylation. however, the source of the sulfur has not been identified and no structures of oahs have been reported in the literature. here, the crystal structure of wolinella succ ...201121931214
rdr6-mediated synthesis of complementary rna is terminated by mirna stably bound to template rna.rna-dependent rna polymerase rdr6 is involved in the biogenesis of plant trans-acting sirnas. this process is initiated by mirna-directed and argonaute (ago) protein-mediated cleavage of tas gene transcripts. one of the cleavage products is converted by rdr6 to double-stranded (ds)rna, the substrate for dicer-like 4 (dcl4). interestingly, tas3 transcript contains two target sites for mir390::ago7 complexes, 5'-non-cleavable and 3'-cleavable. here we show that rdr6-mediated synthesis of complemen ...201121930511
rdr6-mediated synthesis of complementary rna is terminated by mirna stably bound to template rna.rna-dependent rna polymerase rdr6 is involved in the biogenesis of plant trans-acting sirnas. this process is initiated by mirna-directed and argonaute (ago) protein-mediated cleavage of tas gene transcripts. one of the cleavage products is converted by rdr6 to double-stranded (ds)rna, the substrate for dicer-like 4 (dcl4). interestingly, tas3 transcript contains two target sites for mir390::ago7 complexes, 5'-non-cleavable and 3'-cleavable. here we show that rdr6-mediated synthesis of complemen ...201121930511
yeast cytochrome c oxidase: a model system to study mitochondrial forms of the haem-copper oxidase superfamily.the known subunits of yeast mitochondrial cytochrome c oxidase are reviewed. the structures of all eleven of its subunits are explored by building homology models based on the published structures of the homologous bovine subunits and similarities and differences are highlighted, particularly of the core functional subunit i. yeast genetic techniques to enable introduction of mutations into the three core mitochondrially-encoded subunits are reviewed. this article is part of a special issue enti ...201121925484
yeast cytochrome c oxidase: a model system to study mitochondrial forms of the haem-copper oxidase superfamily.the known subunits of yeast mitochondrial cytochrome c oxidase are reviewed. the structures of all eleven of its subunits are explored by building homology models based on the published structures of the homologous bovine subunits and similarities and differences are highlighted, particularly of the core functional subunit i. yeast genetic techniques to enable introduction of mutations into the three core mitochondrially-encoded subunits are reviewed. this article is part of a special issue enti ...201121925484
a novel conformation of rna polymerase sheds light on the mechanism of transcription.transcription is a complicated, multistep process requiring stringent control. its accuracy may be achieved in part by the conformational changes of rna polymerase (rnap). here, we discuss the functional relevance of the recently reported conformational changes of rnap, which may affect transcription control, rnap translocation and transcription termination.201121922057
The role of E. coli Nus-factors in transcription regulation and transcription:translation coupling: From structure to mechanism.Bacterial transcription mediated by RNA polymerase (RNAP) is a highly regulated process and RNAP action is modulated during the different phases of initiation, elongation and termination by proteins such as the Escherichia coli Nus transcription-factors. Here we discuss the structural interplay and the mechanistic role of the Nus-factors that are directly involved in the processivity of elongation, transcription:translation coupling and termination, as well as the varying effects of these protei ...201121922055
translational diffusion of macromolecular assemblies measured using transverse-relaxation-optimized pulsed field gradient nmr.in structural biology, pulsed field gradient (pfg) nmr spectroscopy for the characterization of size and hydrodynamic parameters of macromolecular solutes has the advantage over other techniques that the measurements can be recorded with identical solution conditions as used for nmr structure determination or for crystallization trials. this paper describes two transverse-relaxation-optimized (tro) (15)n-filtered pfg stimulated-echo (ste) experiments for studies of macromolecular translational d ...201121919531
Profile of Venkatraman Ramakrishnan. Interview by Prashant Nair. 201121914843
structural and mutational studies of a hyperthermophilic intein from dna polymerase ii of pyrococcus abyssi.protein splicing is a precise self-catalyzed process in which an intein excises itself from a precursor with the concomitant ligation of the flanking polypeptides (exteins). protein splicing proceeds through a four-step reaction but the catalytic mechanism is not fully understood at the atomic level. we report the solution nmr structures of the hyperthermophilic pyrococcus abyssi polii intein, which has a noncanonical c-terminal glutamine instead of an asparagine. the nmr structures were determi ...201121914805
Crystal structure of phosphopantetheine adenylyltransferase from Enterococcus faecalis in the ligand-unbound state and in complex with ATP and pantetheine.Phosphopantetheine adenylyltransferase (PPAT) catalyzes the reversible transfer of an adenylyl group from ATP to 4'-phosphopantetheine (Ppant) to form dephospho-CoA (dPCoA) and pyrophosphate in the Coenzyme A (CoA) biosynthetic pathway. Importantly, PPATs are the potential target for developing antibiotics because bacterial and mammalian PPATs share little sequence homology. Previous structural studies revealed the mechanism of the recognizing substrates and products. The binding modes of ATP, A ...201121912874
bacteriophage t4 mota activator and the β-flap tip of rna polymerase target the same set of σ70 carboxyl-terminal residues.sigma factors, the specificity subunits of rna polymerase, are involved in interactions with promoter dna, the core subunits of rna polymerase, and transcription factors. the bacteriophage t4-encoded activator, mota, is one such factor, which engages the c terminus of the escherichia coli housekeeping sigma factor, σ(70). mota functions in concert with a phage-encoded co-activator, asia, as a molecular switch. this process, termed sigma appropriation, inhibits host transcription while activating ...201121911499
Identification, tissue distribution, and molecular modeling of novel human isoforms of the key enzyme in sialic acid synthesis, UDP-GlcNAc 2-epimerase/ManNAc kinase.UDP-GlcNAc 2-epimerase/ManNAc kinase (GNE) catalyzes the first two committed steps in sialic acid synthesis. In addition to the three previously described human GNE isoforms (hGNE1-hGNE3), our database and polymerase chain reaction analysis yielded five additional human isoforms (hGNE4-hGNE8). hGNE1 is the ubiquitously expressed major isoform, while the hGNE2-hGNE8 isoforms are differentially expressed and may act as tissue-specific regulators of sialylation. hGNE2 and hGNE7 display a 31-residue ...201121910480
Manganese superoxide dismutase is a mitochondrial fidelity protein that protects Pol? against UV-induced inactivation.Manganese superoxide dismutase is a nuclear encoded primary antioxidant enzyme localized exclusively in the mitochondrial matrix. Genotoxic agents, such as ultraviolet (UV) radiation, generates oxidative stress and cause mitochondrial DNA (mtDNA) damage. The mtDNA polymerase (Pol?), a major constituent of nucleoids, is responsible for the replication and repair of the mitochondrial genome. Recent studies suggest that the mitochondria contain fidelity proteins and MnSOD constitutes an integral pa ...201121909133
Manganese superoxide dismutase is a mitochondrial fidelity protein that protects Pol? against UV-induced inactivation.Manganese superoxide dismutase is a nuclear encoded primary antioxidant enzyme localized exclusively in the mitochondrial matrix. Genotoxic agents, such as ultraviolet (UV) radiation, generates oxidative stress and cause mitochondrial DNA (mtDNA) damage. The mtDNA polymerase (Pol?), a major constituent of nucleoids, is responsible for the replication and repair of the mitochondrial genome. Recent studies suggest that the mitochondria contain fidelity proteins and MnSOD constitutes an integral pa ...201121909133
arrangement of electron transport chain components in bovine mitochondrial supercomplex i1iii2iv1.the respiratory chain in the inner mitochondrial membrane contains three large multi-enzyme complexes that together establish the proton gradient for atp synthesis, and assemble into a supercomplex. a 19-å 3d map of the 1.7-mda amphipol-solubilized supercomplex i(1)iii(2)iv(1) from bovine heart obtained by single-particle electron cryo-microscopy reveals an amphipol belt replacing the membrane lipid bilayer. a precise fit of the x-ray structures of complex i, the complex iii dimer, and monomeric ...201121909073
defense islands in bacterial and archaeal genomes and prediction of novel defense systems.the arms race between cellular life forms and viruses is a major driving force of evolution. a substantial fraction of bacterial and archaeal genomes is dedicated to antivirus defense. we analyzed the distribution of defense genes and typical mobilome components (such as viral and transposon genes) in bacterial and archaeal genomes and demonstrated statistically significant clustering of antivirus defense systems and mobile genes and elements in genomic islands. the defense islands are enriched ...201121908672
Alanyl-tRNA synthetase genes of Vanderwaltozyma polyspora arose from duplication of a dual-functional predecessor of mitochondrial origin.In eukaryotes, the cytoplasmic and mitochondrial forms of a given aminoacyl-tRNA synthetase (aaRS) are typically encoded by two orthologous nuclear genes, one of eukaryotic origin and the other of mitochondrial origin. We herein report a novel scenario of aaRS evolution in yeast. While all other yeast species studied possess a single nuclear gene encoding both forms of alanyl-tRNA synthetase (AlaRS), Vanderwaltozyma polyspora, a yeast species descended from the same whole-genome duplication even ...201221908394
Alanyl-tRNA synthetase genes of Vanderwaltozyma polyspora arose from duplication of a dual-functional predecessor of mitochondrial origin.In eukaryotes, the cytoplasmic and mitochondrial forms of a given aminoacyl-tRNA synthetase (aaRS) are typically encoded by two orthologous nuclear genes, one of eukaryotic origin and the other of mitochondrial origin. We herein report a novel scenario of aaRS evolution in yeast. While all other yeast species studied possess a single nuclear gene encoding both forms of alanyl-tRNA synthetase (AlaRS), Vanderwaltozyma polyspora, a yeast species descended from the same whole-genome duplication even ...201221908394
simultaneous polyhydroxyalkanoates and rhamnolipids production by thermus thermophilus hb8.abstract: the ability of thermus thermophilus hb8 to produce simultaneously two environmentally-friendly biodegradable products, polyhydroxyalkanoates (phas) and rhamnolipids (rls), using either sodium gluconate or glucose as sole carbon source, was demonstrated. the utilization of sodium gluconate resulted in higher levels of phas and rls production than when glucose was used as sole carbon source. the initial phosphate concentration (as po43-) influences both phas and rls productions that were ...201121906373
exploiting bacterial dna gyrase as a drug target: current state and perspectives.dna gyrase is a type ii topoisomerase that can introduce negative supercoils into dna at the expense of atp hydrolysis. it is essential in all bacteria but absent from higher eukaryotes, making it an attractive target for antibacterials. the fluoroquinolones are examples of very successful gyrase-targeted drugs, but the rise in bacterial resistance to these agents means that we not only need to seek new compounds, but also new modes of inhibition of this enzyme. we review known gyrase-specific d ...201121904817
structure of nitrilotriacetate monooxygenase component b from mycobacterium thermoresistibile.mycobacterium tuberculosis belongs to a large family of soil bacteria which can degrade a remarkably broad range of organic compounds and utilize them as carbon, nitrogen and energy sources. it has been proposed that a variety of mycobacteria can subsist on alternative carbon sources during latency within an infected human host, with the help of enzymes such as nitrilotriacetate monooxygenase (nta-mo). nta-mo is a member of a class of enzymes which consist of two components: a and b. while compo ...201121904057
Probing conformational states of glutaryl-CoA dehydrogenase by fragment screening.Glutaric acidemia type 1 is an inherited metabolic disorder which can cause macrocephaly, muscular rigidity, spastic paralysis and other progressive movement disorders in humans. The defects in glutaryl-CoA dehydrogenase (GCDH) associated with this disease are thought to increase holoenzyme instability and reduce cofactor binding. Here, the first structural analysis of a GCDH enzyme in the absence of the cofactor flavin adenine dinucleotide (FAD) is reported. The apo structure of GCDH from Burkh ...201121904051
An ensemble of structures of Burkholderia pseudomallei 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase.Burkholderia pseudomallei is a soil-dwelling bacterium endemic to Southeast Asia and Northern Australia. Burkholderia is responsible for melioidosis, a serious infection of the skin. The enzyme 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase (PGAM) catalyzes the interconversion of 3-phosphoglycerate and 2-phosphoglycerate, a key step in the glycolytic pathway. As such it is an extensively studied enzyme and X-ray crystal structures of PGAM enzymes from multiple species have been elucid ...201121904048
structures of phosphopantetheine adenylyltransferase from burkholderia pseudomallei.phosphopantetheine adenylyltransferase (ppat) catalyzes the fourth of five steps in the coenzyme a biosynthetic pathway, reversibly transferring an adenylyl group from atp onto 4'-phosphopantetheine to yield dephospho-coenzyme a and pyrophosphate. burkholderia pseudomallei is a soil- and water-borne pathogenic bacterium and the etiologic agent of melioidosis, a potentially fatal systemic disease present in southeast asia. two crystal structures are presented of the ppat from b. pseudomallei with ...201121904046
Crystal structure of the hybrid state of ribosome in complex with the guanosine triphosphatase release factor 3.Protein release factor 3 (RF3), a guanosine triphosphatase, binds to ribosome after release of the nascent peptide and promotes dissociation of the class I release factors during the termination of protein synthesis. Here we present the crystal structure of the 70S ribosome with RF3 in the presence of a nonhydrolyzable GTP analogue, guanosine 5'-ß,?-methylenetriphosphate (GDPCP), refined to 3.8 Å resolution. The structure shows that the subunits of the ribosome are rotated relative to each other ...201121903932
peptides from aminoacyl-trna synthetases can cure the defects due to mutations in mt trna genes.recent results from several laboratories have confirmed that human and yeast leucyl- and valyl-trna synthetases can rescue the respiratory defects due to mutations in mitochondrial trna genes. in this report we show that this effect cannot be ascribed to the catalytic activity per se and that isolated domains of aminoacyl-trna synthetases and even short peptides thereof have suppressing effects.201121903180
structural basis for leucine-induced allosteric activation of glutamate dehydrogenase.glutamate dehydrogenase (gdh) catalyzes reversible conversion between glutamate and 2-oxoglutarate using nad(p)(h) as a coenzyme. although mammalian gdh is regulated by gtp through the antenna domain, little is known about the mechanism of allosteric activation by leucine. an extremely thermophilic bacterium, thermus thermophilus, possesses gdh with a unique subunit configuration composed of two different subunits, gdha (regulatory subunit) and gdhb (catalytic subunit). t. thermophilus gdh is un ...201121900230
Mimivirus reveals Mre11/Rad50 fusion proteins with a sporadic distribution in eukaryotes, bacteria, viruses and plasmids.The Mre11/Rad50 complex and the homologous SbcD/SbcC complex in bacteria play crucial roles in the metabolism of DNA double-strand breaks, including DNA repair, genome replication, homologous recombination and non-homologous end-joining in cellular life forms and viruses. Here we investigated the amino acid sequence of the Mimivirus R555 gene product, originally annotated as a Rad50 homolog, and later shown to have close homologs in marine microbial metagenomes.201121899737
binding and inhibition of human spermidine synthase by decarboxylated s-adenosylhomocysteine.aminopropyltransferases are essential enzymes that form polyamines in eukaryotic and most prokaryotic cells. spermidine synthase (spds) is one of the most well-studied enzymes in this biosynthetic pathway. the enzyme uses decarboxylated s-adenosylmethionine and a short-chain polyamine (putrescine) to make a medium-chain polyamine (spermidine) and 5'-deoxy-5'-methylthioadenosine as a byproduct. here, we report a new spermidine synthase inhibitor, decarboxylated s-adenosylhomocysteine (dcsah). the ...201121898642
Is the sequence-specific binding of aminoacyl-tRNAs by EF-Tu universal among bacteria?Three base pairs in the T-stem are primarily responsible for the sequence-specific interaction of tRNA with Escherichia coli and Thermus thermophilus EF-Tu. While the amino acids on the surface of EF-Tu that contact aminoacyl-tRNA (aa-tRNA) are highly conserved among bacteria, the T-stem sequences of individual tRNA are variable, making it unclear whether or not this protein-nucleic acid interaction is also sequence specific in other bacteria. We propose and validate a thermodynamic model that p ...201121893586
ribonuclease j: how to lead a double life. 201121893280
Crystal structures of an archaeal class II DNA photolyase and its complex with UV-damaged duplex DNA.Class II photolyases ubiquitously occur in plants, animals, prokaryotes and some viruses. Like the distantly related microbial class I photolyases, these enzymes repair UV-induced cyclobutane pyrimidine dimer (CPD) lesions within duplex DNA using blue/near-UV light. Methanosarcina mazei Mm0852 is a class II photolyase of the archaeal order of Methanosarcinales, and is closely related to plant and metazoan counterparts. Mm0852 catalyses light-driven DNA repair and photoreduction, but in contrast ...201121892138
tracing the trail of protons through complex i of the mitochondrial respiratory chain.mitochondria are the structures that produce the bulk part of the cellular energy currency atp, which drives numerous energy requiring processes in the cell. this process involves a series of large enzyme complexes--the respiratory chain--that couples the transfer of electrons to the creation of a concentration gradient of protons across the inner mitochondrial membrane, which drives atp synthesis. complex i (or nadh-quinone oxidoreductase) is the largest and by far the most complicated of the r ...201121886481
the universally conserved prokaryotic gtpases.members of the large superclass of p-loop gtpases share a core domain with a conserved three-dimensional structure. in eukaryotes, these proteins are implicated in various crucial cellular processes, including translation, membrane trafficking, cell cycle progression, and membrane signaling. as targets of mutation and toxins, gtpases are involved in the pathogenesis of cancer and infectious diseases. in prokaryotes also, it is hard to overestimate the importance of gtpases in cell physiology. nu ...201121885683
identification of human fumarylacetoacetate hydrolase domain-containing protein 1 (fahd1) as a novel mitochondrial acylpyruvase.the human fumarylacetoacetate hydrolase (fah) domain-containing protein 1 (fahd1) is part of the fah protein superfamily, but its enzymatic function is unknown. in the quest for a putative enzymatic function of fahd1, we found that fahd1 exhibits acylpyruvase activity, demonstrated by the hydrolysis of acetylpyruvate and fumarylpyruvate in vitro, whereas several structurally related compounds were not hydrolyzed as efficiently. conserved amino acids asp-102 and arg-106 of fahd1 were found import ...201121878618
interaction of complexes i, iii, and iv within the bovine respirasome by single particle cryoelectron tomography.the respirasome is a multisubunit supercomplex of the respiratory chain in mitochondria. here we report the 3d reconstruction of the bovine heart respirasome, composed of dimeric complex iii and single copies of complex i and iv, at about 2.2-nm resolution, determined by cryoelectron tomography and subvolume averaging. fitting of x-ray structures of single complexes i, iii(2), and iv with high fidelity allows interpretation of the model at the level of secondary structures and shows how the indi ...201121876144
termination of protein synthesis in mammalian mitochondria.all mechanisms of protein synthesis can be considered in four stages: initiation, elongation, termination, and ribosome recycling. remarkable progress has been made in understanding how these processes are mediated in the cytosol of many species; however, details of organellar protein synthesis remain sketchy. this is an important omission, as defects in human mitochondrial translation are known to cause disease and may contribute to the aging process itself. in this minireview, we focus on the ...201121873426
enzymatic synthesis of long double-stranded dna labeled with haloderivatives of nucleobases in a precisely pre-determined sequence.restriction endonucleases are widely applied in recombinant dna technology. among them, enzymes of class iis, which cleave dna beyond recognition sites, are especially useful. we use bsai enzyme for the pinpoint introduction of halogen nucleobases into dna. this has been done for the purpose of anticancer radio- and phototherapy that is our long-term objective.201121864341
5' End-independent RNase J1 endonuclease cleavage of Bacillus subtilis model RNA.Bacillus subtilis trp leader RNA is a small (140-nucleotide) RNA that results from attenuation of trp operon transcription upon binding of the regulatory TRAP complex. Previously, endonucleolytic cleavage by ribonuclease RNase J1 in a 3'-proximal, single-stranded region was shown to be critical for initiation of trp leader RNA decay. RNase J1 is a dual-specificity enzyme, with both 5' exonucleolytic and endonucleolytic activities. Here, we provide in vivo and in vitro evidence that RNase J1 acce ...201121862575
new target for inhibition of bacterial rna polymerase: 'switch region'.a new drug target - the 'switch region' - has been identified within bacterial rna polymerase (rnap), the enzyme that mediates bacterial rna synthesis. the new target serves as the binding site for compounds that inhibit bacterial rna synthesis and kill bacteria. since the new target is present in most bacterial species, compounds that bind to the new target are active against a broad spectrum of bacterial species. since the new target is different from targets of other antibacterial agents, com ...201121862392
Properties and crystal structure of methylenetetrahydrofolate reductase from Thermus thermophilus HB8.Methylenetetrahydrofolate reductase (MTHFR) is one of the enzymes involved in homocysteine metabolism. Despite considerable genetic and clinical attention, the reaction mechanism and regulation of this enzyme are not fully understood because of difficult production and poor stability. While recombinant enzymes from thermophilic organisms are often stable and easy to prepare, properties of thermostable MTHFRs have not yet been reported.201121858212
the rela/spot homolog (rsh) superfamily: distribution and functional evolution of ppgpp synthetases and hydrolases across the tree of life.rela/spot homologue (rsh) proteins, named for their sequence similarity to the rela and spot enzymes of escherichia coli, comprise a superfamily of enzymes that synthesize and/or hydrolyze the alarmone ppgpp, activator of the "stringent" response and regulator of cellular metabolism. the classical "long" rshs rel, rela and spot with the ppgpp hydrolase, synthetase, tgs and act domain architecture have been found across diverse bacteria and plant chloroplasts, while dedicated single domain ppgpp- ...201121858139
Characterization of benzoxaborole-based antifungal resistance mutations demonstrates that editing depends on electrostatic stabilization of the leucyl-tRNA synthetase editing cap.The broad-spectrum benzoxaborole antifungal AN2690 blocks protein synthesis by inhibiting leucyl-tRNA synthetase (LeuRS) via a novel oxaborole tRNA trapping mechanism in the editing site. Herein, one set of resistance mutations is at Asp487 outside the LeuRS hydrolytic editing pocket, in a region of unknown function. It is located within a eukaryote/archaea specific insert I4, which forms part of a cap over a benzoxaborole-AMP that is bound in the LeuRS CP1 domain editing active site. Mutational ...201121856301
orphan macrodomain protein (human c6orf130) is an o-acyl-adp-ribose deacylase: solution structure and catalytic properties.post-translational modification of proteins/histones by lysine acylation has profound effects on the physiological function of modified proteins. deacylation by nad(+)-dependent sirtuin reactions yields as a product o-acyl-adp-ribose, which has been implicated as a signaling molecule in modulating cellular processes. macrodomain-containing proteins are reported to bind nad(+)-derived metabolites. here, we describe the structure and function of an orphan macrodomain protein, human c6orf130. this ...201121849506
tolerance to changes in membrane lipid composition as a selected trait of membrane proteins.membrane lipid composition can vary dramatically across the three domains of life and even within single organisms. here we review evidence that the lipid-exposed surfaces of membrane proteins have generally evolved to maintain correct structure and function in the face of major changes in lipid composition. such tolerance has allowed evolution to extensively remodel membrane lipid compositions during the emergence of new species without having to extensively remodel the associated membrane prot ...201121848311
Structural analysis of the Ras-like G protein MglA and its cognate GAP MglB and implications for bacterial polarity.The bacterium Myxococcus xanthus uses a G protein cycle to dynamically regulate the leading/lagging pole polarity axis. The G protein MglA is regulated by its GTPase-activating protein (GAP) MglB, thus resembling Ras family proteins. Here, we show structurally and biochemically that MglA undergoes a dramatic, GDP-GTP-dependent conformational change involving a screw-type forward movement of the central ß2-strand, never observed in any other G protein. This movement and complex formation with Mgl ...201121847100
insights into folate/fad-dependent trna methyltransferase mechanism: role of two highly conserved cysteines in catalysis.the flavoprotein trmfo methylates specifically the c5 carbon of the highly conserved uridine 54 in trnas. contrary to most methyltransferases, the 1-carbon unit transferred by trmfo derives from 5,10-methylenetetrahydrofolate and not from s-adenosyl-l-methionine. the enzyme also employs the fad hydroquinone as a reducing agent of the c5 methylene u54-trna intermediate in vitro. by analogy with the catalytic mechanism of thymidylate synthase thya, a conserved cysteine located near the fad isoallo ...201121846722
vma8p-gfp fusions can be functionally incorporated into v-atpase, suggesting structural flexibility at the top of v1.the vacuolar atpase (v-atpase) complex of yeast (saccharomyces cerevisiae) is comprised of two sectors, v(1) (catalytic) and v(o) (proton transfer). the hexameric (a(3)b(3)) cylinder of v(1) has a central cavity that must accommodate at least part of the rotary stalk of v-atpase, a key component of which is subunit d (vma8p). recent electron microscopy (em) data for the prokaryote v-atpase complex (thermus thermophilus) suggest that subunit d penetrates deeply into the central cavity. the functi ...201121845105
adaptation of aerobic respiration to low o2 environments.aerobic respiration in bacteria, archaea, and mitochondria is performed by oxygen reductase members of the heme-copper oxidoreductase superfamily. these enzymes are redox-driven proton pumps which conserve part of the free energy released from oxygen reduction to generate a proton motive force. the oxygen reductases can be divided into three main families based on evolutionary and structural analyses (a-, b- and c-families), with the b- and c-families evolving after the a-family. the a-family ut ...201121844375
substrate trna recognition mechanism of a multi-site-specific trna methyltransferase, aquifex aeolicus trm1, based on the x-ray crystal structure.archaeal and eukaryotic trna (n(2)-, n(2)-guanine) dimethyltransferase (trm1) produces n(2)-, n(2)-dimethylguanine at position 26 in trna. in contrast, trm1 from aquifex aeolicus, a hyper-thermophilic eubacterium, modifies g27 as well as g26. here, a gel-mobility shift assay revealed that the t-arm in trna is the binding site of a. aeolicus trm1. to address the multi-site specificity, we performed an x-ray crystal structure study. the overall structure of a. aeolicus trm1 is similar to that of a ...201121844194
the elusive middle domain of hsp104 and clpb: location and function.hsp104 in yeast and clpb in bacteria are homologous, hexameric aaa+ proteins and hsp100 chaperones, which function in the stress response as ring-translocases that drive protein disaggregation and reactivation. both hsp104 and clpb contain a distinctive coiled-coil middle domain (md) inserted in the first aaa+ domain, which distinguishes them from other aaa+ proteins and hsp100 family members. here, we focus on recent developments concerning the location and function of the md in these hexameric ...201121843558
the elusive middle domain of hsp104 and clpb: location and function.hsp104 in yeast and clpb in bacteria are homologous, hexameric aaa+ proteins and hsp100 chaperones, which function in the stress response as ring-translocases that drive protein disaggregation and reactivation. both hsp104 and clpb contain a distinctive coiled-coil middle domain (md) inserted in the first aaa+ domain, which distinguishes them from other aaa+ proteins and hsp100 family members. here, we focus on recent developments concerning the location and function of the md in these hexameric ...201121843558
mechanistic analysis of trehalose synthase from mycobacterium smegmatis.trehalose synthase (tres) catalyzes the reversible interconversion of maltose and trehalose and has been recently shown to function primarily in the mobilisation of trehalose as a glycogen precursor. consequently the mechanism of this intriguing isomerase is of both academic and potential pharmacological interest. tres catalyzes the hydrolytic cleavage of +¦-aryl glucosides, as well as +¦-glucosyl fluoride, thereby allowing facile continuous assays. reaction of tres with 5-fluoroglycosyl fluorid ...201121840994
cbpa acts as a modulator of hspr repressor dna binding activity in helicobacter pylori.the ability of pathogens to cope with disparate environmental stresses is a crucial feature for bacterial survival and for the establishment of a successful infection and colonization of the host; in this respect, chaperones and heat-shock proteins (hsps) play a fundamental role in host-pathogen interactions. in helicobacter pylori, the expression of the major hsps is tightly regulated through dedicated transcriptional repressors (named hspr and hrca), as well as via a groesl-dependent post-tran ...201121840971
structural contribution of the c-terminal segments of nuol (nd5) and nuom (nd4) subunits of complex i from e. coli.the proton-translocating nadh-quinone oxidoreductase (complex i/ndh-1) is a multi-subunit enzymatic complex. it has a characteristic l-shaped form with two domains, a hydrophilic peripheral domain and a hydrophobic membrane domain. the membrane domain contains three antiporter-like subunits (nuol, nuom and nuon, escherichia coli naming) that are considered to be involved in the proton translocation. deletion of either nuol or nuom resulted in an incomplete assembly of ndh-1 and a total loss of t ...201121835926
mechanisms and evolution of oxidative sulfur metabolism in green sulfur bacteria.green sulfur bacteria (gsb) constitute a closely related group of photoautotrophic and thiotrophic bacteria with limited phenotypic variation. they typically oxidize sulfide and thiosulfate to sulfate with sulfur globules as an intermediate. based on genome sequence information from 15 strains, the distribution and phylogeny of enzymes involved in their oxidative sulfur metabolism was investigated. at least one homolog of sulfide:quinone oxidoreductase (sqr) is present in all strains. in all sul ...201121833341
how are "atypical" sulfite dehydrogenases linked to cell metabolism? interactions between the sort sulfite dehydrogenase and small redox proteins.sulfite dehydrogenases (sdhs) are enzymes that catalyze the oxidation of the toxic and mutagenic compound sulfite to sulfate, thereby protecting cells from adverse effects associated with sulfite exposure. while some bacterial sdhs that have been characterized to date are able to use cytochrome c as an electron acceptor, the majority of these enzymes prefer ferricyanide as an electron acceptor and have therefore been termed "atypical" sdhs. identifying the natural electron acceptor of these enzy ...201121833314
engineering the respiratory complex i to an energy-converting nadph:ubiquinone oxidoreductase.the respiratory complex i couples the electron transfer from nadh to ubiquinone with a translocation of protons across the membrane. its nucleotide binding site is made up of a unique rossmann fold to accommodate the binding of the substrate nadh and of the primary electron acceptor flavin mononucleotide. binding of nadh includes interactions of the hydroxyl groups of the adenosine ribose with a conserved glutamic acid residue. structural analysis revealed that due to steric hinderance and elect ...201121832062
rnase h-dependent pcr (rhpcr): improved specificity and single nucleotide polymorphism detection using blocked cleavable primers.abstract: background: the polymerase chain reaction (pcr) is commonly used to detect the presence of nucleic acid sequences both in research and diagnostic settings. while high specificity is often achieved, biological requirements sometimes require that primers are placed in suboptimal locations which lead to problems with the formation of primer dimers and/or misamplification of homologous sequences. results: pyrococcus abyssi (p.a.) rnase h2 was used to enable pcr to be performed using blocke ...201121831278
overexpression of a cytosolic pyrophosphatase (tgppase) reveals a regulatory role of pyrophosphate in glycolysis for toxoplasma gondii.pyrophosphate (ppi) is a critical element of cellular metabolism as both an energy donor and as an allosteric regulator of several metabolic pathways. the apicomplexan parasite, toxoplasma gondii, uses ppi in place of atp as an energy donor in at least two reactions: the glycolytic ppi-dependent phosphofructokinase (pfk), and the proton translocating vacuolar pyrophosphatase (v-h+-ppase). in the present work, we report the cloning, expression, and characterization of a cytosolic pyrophosphatase ...201121831041
structural classification and properties of ketoacyl synthases.ketoacyl synthases (kss) catalyze condensing reactions combining acyl-coa or acyl-acyl carrier protein (acp) with malonyl-coa to form 3-ketoacyl-coa, or with malonyl-acp to form 3-ketoacyl-acp. in each case the resulting acyl chain is two carbon atoms longer than before, and co(2) and either coa or acp are formed. kss also join other activated molecules in the polyketide synthesis cycle. our classification of kss by their primary and tertiary structures instead of by their substrates and the rea ...201121830247
a computational study of elongation factor g (efg) duplicated genes: diverged nature underlying the innovation on the same structural template.elongation factor g (efg) is a core translational protein that catalyzes the elongation and recycling phases of translation. a more complex picture of efg's evolution and function than previously accepted is emerging from analyzes of heterogeneous efg family members. whereas the gene duplication is postulated to be a prominent factor creating functional novelty, the striking divergence between efg paralogs can be interpreted in terms of innovation in gene function.201121829651
unveiling the structural basis for translational ambiguity tolerance in a human fungal pathogen.in a restricted group of opportunistic fungal pathogens the universal leucine cug codon is translated both as serine (97%) and leucine (3%), challenging the concept that translational ambiguity has a negative impact in living organisms. to elucidate the molecular mechanisms underlying the in vivo tolerance to a nonconserved genetic code alteration, we have undertaken an extensive structural analysis of proteins containing cug-encoded residues and solved the crystal structures of the two natural ...201121825144
a single methyltransferase yefa (rlmcd) catalyses both m5u747 and m5u1939 modifications in bacillus subtilis 23s rrna.methyltransferases that use s-adenosylmethionine (adomet) as a cofactor to catalyse 5-methyl uridine (m(5)u) formation in trnas and rrnas are widespread in bacteria and eukaryota, and are also found in certain archaea. these enzymes belong to the cog2265 cluster, and the gram-negative bacterium escherichia coli possesses three paralogues. these comprise the methyltransferases trma that targets u54 in trnas, rlmc that modifies u747 in 23s rrna and rlmd that is specific for u1939 in 23s rrna. the ...201121824914
expression, purification, crystallization and preliminary crystallographic analysis of cg1458: a novel oxaloacetate decarboxylase from corynebacterium glutamicum.oxaloacetate decarboxylase catalyses the decarboxylation of oxaloacetate to pyruvate and co(2). recently, the corynebacterium glutamicum gene product cg1458 was determined to be a soluble oxaloacetate decarboxylase. to elucidate the mechanism of oxaloacetate decarboxylation by cg1458, recombinant cg1458 was purified and crystallized. the best crystal was grown from 0.2ôçàm mgcl(2), 0.1ôçàm bis-tris ph 6.0, 25%(w/v) polyethylene glycol 3350 using the hanging-drop method. the crystals belonged to ...201121821907
expression, purification and preliminary crystallographic analysis of o-acetylhomoserine sulfhydrylase from mycobacterium tuberculosis.the gene product of the open reading frame rv3340 from mycobacterium tuberculosis is annotated as encoding a probable o-acetylhomoserine (oah) sulfhydrylase (metc), an enzyme that catalyzes the last step in the biosynthesis of methionine, which is an essential amino acid in bacteria and plants. following overexpression in escherichia coli, the m. tuberculosis metc enzyme was purified and crystallized using the hanging-drop vapor-diffusion method. native diffraction data were collected from cryst ...201121821905
crystallization and preliminary x-ray analysis of a cold-active +¦-galactosidase from the psychrotrophic and halotolerant planococcus sp. l4.+¦-galactosidases catalyze the hydrolysis of a galactosyl moiety from the nonreducing termini of oligosaccharides or from glycosides. a novel gh family 42 cold-active +¦-galactosidase identified from the psychrotrophic and halotolerant planococcus sp. l4 (bgap) was crystallized and a complete data set was collected from a single frozen crystal on an in-house x-ray source. the crystal diffracted to 2.8ôçà+à resolution and belonged to space group p1, with unit-cell parameters a = 104.29, b = 118.1 ...201121821893
polyphosphate--an ancient energy source and active metabolic regulator.there are a several molecules on earth that effectively store energy within their covalent bonds, and one of these energy-rich molecules is polyphosphate. in microbial cells, polyphosphate granules are synthesised for both energy and phosphate storage and are degraded to produce nucleotide triphosphate or phosphate. energy released from these energetic carriers is used by the cell for production of all vital molecules such as amino acids, nucleobases, sugars and lipids. polyphosphate chains dire ...201121816086
noncanonical amino acids in the interrogation of cellular protein synthesis.proteins in living cells can be made receptive to bioorthogonal chemistries through metabolic labeling with appropriately designed noncanonical amino acids (ncaas). in the simplest approach to metabolic labeling, an amino acid analog replaces one of the natural amino acids specified by the protein's gene (or genes) of interest. through manipulation of experimental conditions, the extent of the replacement can be adjusted. this approach, often termed residue-specific incorporation, allows the nca ...201121815659
high resolution structure of the ba3 cytochrome c oxidase from thermus thermophilus in a lipidic environment.the fundamental chemistry underpinning aerobic life on earth involves reduction of dioxygen to water with concomitant proton translocation. this process is catalyzed by members of the heme-copper oxidase (hco) superfamily. despite the availability of crystal structures for all types of hco, the mode of action for this enzyme is not understood at the atomic level, namely how vectorial h(+) and e(-) transport are coupled. toward addressing this problem, we report wild type and a120f mutant structu ...201121814577
gln-trnagln synthesis in a dynamic transamidosome from helicobacter pylori, where glurs2 hydrolyzes excess glu-trnagln.in many bacteria and archaea, an ancestral pathway is used where asparagine and glutamine are formed from their acidic precursors while covalently linked to trna(asn) and trna(gln), respectively. stable complexes formed by the enzymes of these indirect trna aminoacylation pathways are found in several thermophilic organisms, and are called transamidosomes. we describe here a transamidosome forming gln-trna(gln) in helicobacter pylori, an ε-proteobacterium pathogenic for humans; this transamidoso ...201121813455
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