| molecular mechanisms of crispr-mediated microbial immunity. | bacteriophages (phages) infect bacteria in order to replicate and burst out of the host, killing the cell, when reproduction is completed. thus, from a bacterial perspective, phages pose a persistent lethal threat to bacterial populations. not surprisingly, bacteria evolved multiple defense barriers to interfere with nearly every step of phage life cycles. phages respond to this selection pressure by counter-evolving their genomes to evade bacterial resistance. the antagonistic interaction betwe ... | 2013 | 23959171 |
| an archaeal crispr type iii-b system exhibiting distinctive rna targeting features and mediating dual rna and dna interference. | crispr-cas systems provide a small rna-based mechanism to defend against invasive genetic elements in archaea and bacteria. to investigate the in vivo mechanism of rna interference by two type iii-b systems (cmr-α and cmr-β) in sulfolobus islandicus, a genetic assay was developed using plasmids carrying an artificial mini-crispr (ac) locus with a single spacer. after pac plasmids were introduced into different strains, northern analyses confirmed that mature crrnas were produced from the plasmid ... | 2014 | 25505143 |
| an archaeal crispr type iii-b system exhibiting distinctive rna targeting features and mediating dual rna and dna interference. | crispr-cas systems provide a small rna-based mechanism to defend against invasive genetic elements in archaea and bacteria. to investigate the in vivo mechanism of rna interference by two type iii-b systems (cmr-α and cmr-β) in sulfolobus islandicus, a genetic assay was developed using plasmids carrying an artificial mini-crispr (ac) locus with a single spacer. after pac plasmids were introduced into different strains, northern analyses confirmed that mature crrnas were produced from the plasmid ... | 2014 | 25505143 |
| crystal structure of cas9 in complex with guide rna and target dna. | the crispr-associated endonuclease cas9 can be targeted to specific genomic loci by single guide rnas (sgrnas). here, we report the crystal structure of streptococcus pyogenes cas9 in complex with sgrna and its target dna at 2.5 å resolution. the structure revealed a bilobed architecture composed of target recognition and nuclease lobes, accommodating the sgrna:dna heteroduplex in a positively charged groove at their interface. whereas the recognition lobe is essential for binding sgrna and dna, ... | 2014 | 24529477 |
| structures of cas9 endonucleases reveal rna-mediated conformational activation. | type ii crispr (clustered regularly interspaced short palindromic repeats)-cas (crispr-associated) systems use an rna-guided dna endonuclease, cas9, to generate double-strand breaks in invasive dna during an adaptive bacterial immune response. cas9 has been harnessed as a powerful tool for genome editing and gene regulation in many eukaryotic organisms. we report 2.6 and 2.2 angstrom resolution crystal structures of two major cas9 enzyme subtypes, revealing the structural core shared by all cas9 ... | 2014 | 24505130 |
| essentiality of threonylcarbamoyladenosine (t(6)a), a universal trna modification, in bacteria. | threonylcarbamoyladenosine (t(6)a) is a modified nucleoside universally conserved in trnas in all three kingdoms of life. the recently discovered genes for t(6)a synthesis, including tsac and tsad, are essential in model prokaryotes but not essential in yeast. these genes had been identified as antibacterial targets even before their functions were known. however, the molecular basis for this prokaryotic-specific essentiality has remained a mystery. here, we show that t(6)a is a strong positive ... | 2015 | 26337258 |
| structure principles of crispr-cas surveillance and effector complexes. | the pathway of crispr-cas immunity redefines the roles of rna in the flow of genetic information and ignites excitement for next-generation gene therapy tools. crispr-cas machineries offer a fascinating set of new enzyme assemblies from which one can learn principles of molecular interactions and chemical activities. the interference step of the crispr-cas immunity pathway congregates proteins, rna, and dna into a single molecular entity that selectively destroys invading nucleic acids. although ... | 2015 | 26048003 |
| applications of cas9 as an rna-programmed rna-binding protein. | the streptococcus pyogenes crispr-cas system has gained widespread application as a genome editing and gene regulation tool as simultaneous cellular delivery of the cas9 protein and guide rnas enables recognition of specific dna sequences. the recent discovery that cas9 can also bind and cleave rna in an rna-programmable manner indicates the potential utility of this system as a universal nucleic acid-recognition technology. rna-targeted cas9 (rcas9) could allow identification and manipulation o ... | 2015 | 25880497 |
| prokaryotic nitrate reduction: molecular properties and functional distinction among bacterial nitrate reductases. | | 1999 | 10542156 |
| bioenergetics of the archaea. | in the late 1970s, on the basis of rrna phylogeny, archaea (archaebacteria) was identified as a distinct domain of life besides bacteria (eubacteria) and eucarya. though forming a separate domain, archaea display an enormous diversity of lifestyles and metabolic capabilities. many archaeal species are adapted to extreme environments with respect to salinity, temperatures around the boiling point of water, and/or extremely alkaline or acidic ph. this has posed the challenge of studying the molecu ... | 1999 | 10477309 |
| membrane protein crystallization in amphiphile phases: practical and theoretical considerations. | integral membrane proteins are amphiphilic molecules. in order to enable chromatographic purification and crystallization, a complementary amphiphilic microenvironment must be created and maintained. various types of amphiphilic phases have been employed in crystallizations and intricate amphiphilic microenvironmental structures have resulted from these and are found inside membrane protein crystals. in this review the process of crystallization is put into the context of amphiphile phase transi ... | 2004 | 15652249 |
| membrane protein crystallization in amphiphile phases: practical and theoretical considerations. | integral membrane proteins are amphiphilic molecules. in order to enable chromatographic purification and crystallization, a complementary amphiphilic microenvironment must be created and maintained. various types of amphiphilic phases have been employed in crystallizations and intricate amphiphilic microenvironmental structures have resulted from these and are found inside membrane protein crystals. in this review the process of crystallization is put into the context of amphiphile phase transi ... | 2004 | 15652249 |
| catalytic reduction of no to n2o by a designed heme copper center in myoglobin: implications for the role of metal ions. | the effects of metal ions on the reduction of nitric oxide (no) with a designed heme copper center in myoglobin (f43h/l29h sperm whale mb, cubmb) were investigated under reducing anaerobic conditions using uv-vis and epr spectroscopic techniques as well as gc/ms. in the presence of cu(i), catalytic reduction of no to n2o by cubmb was observed with turnover number of 2 mol no.mol cubmb-1.min-1, close to 3 mol no.mol enzyme-1.min-1 reported for the ba3 oxidases from t. thermophilus. formation of a ... | 2006 | 16719438 |
| energy transduction: proton transfer through the respiratory complexes. | a series of metalloprotein complexes embedded in a mitochondrial or bacterial membrane utilize electron transfer reactions to pump protons across the membrane and create an electrochemical potential (deltamuh+). current understanding of the principles of electron-driven proton transfer is discussed, mainly with respect to the wealth of knowledge available from studies of cytochrome c oxidase. structural, experimental, and theoretical evidence supports the model of long-distance proton transfer v ... | 2006 | 16756489 |
| conserved lipid-binding sites in membrane proteins: a focus on cytochrome c oxidase. | specific interactions between lipids and membrane proteins have been observed in recent high-resolution crystal structures of membrane proteins. a number of cytochrome oxidase structures were analyzed, along with many amino acid sequences of membrane-spanning regions aligned according to their location in the membrane. the results reveal conservation of lipid-binding sites and of the residues that form them. these studies imply that bound lipids have important roles that are crucial to the assem ... | 2007 | 17719219 |
| spectroscopic characterization of heme iron-nitrosyl species and their role in no reductase mechanisms in diiron proteins. | | 2007 | 17534533 |
| cytochrome c oxidase: exciting progress and remaining mysteries. | cytochrome c oxidase generates a proton motive force by two separate mechanisms. the first mechanism is similar to that postulated by peter mitchell, and is based on electrons and protons used to generate water coming from opposite sides of the membrane. the second mechanism was not initially anticipated, but is now firmly established as a proton pump. a brief review of the current state of our understanding of the proton pump of cytochrome oxidase is presented. we have come a long way since the ... | 2008 | 18975062 |
| the chemistry and biochemistry of heme c: functional bases for covalent attachment. | a discussion of the literature concerning the synthesis, function, and activity of heme c-containing proteins is presented. comparison of the properties of heme c, which is covalently bound to protein, is made to heme b, which is bound noncovalently. a question of interest is why nature uses biochemically expensive heme c in many proteins when its properties are expected to be similar to heme b. considering the effects of covalent heme attachment on heme conformation and on the proximal histidin ... | 2008 | 19030605 |
| one heme, diverse functions: using biosynthetic myoglobin models to gain insights into heme-copper oxidases and nitric oxide reductases. | | 2008 | 18729107 |
| the q-cycle reviewed: how well does a monomeric mechanism of the bc(1) complex account for the function of a dimeric complex? | recent progress in understanding the q-cycle mechanism of the bc(1) complex is reviewed. the data strongly support a mechanism in which the q(o)-site operates through a reaction in which the first electron transfer from ubiquinol to the oxidized iron-sulfur protein is the rate-determining step for the overall process. the reaction involves a proton-coupled electron transfer down a hydrogen bond between the ubiquinol and a histidine ligand of the [2fe-2s] cluster, in which the unfavorable protoni ... | 2008 | 18501698 |
| nadh/nad+ interaction with nadh: ubiquinone oxidoreductase (complex i). | the quantitative data on the binding affinity of nadh, nad(+), and their analogues for complex i as emerged from the steady-state kinetics data and from more direct studies under equilibrium conditions are summarized and discussed. the redox-dependency of the nucleotide binding and the reductant-induced change of fmn affinity to its tight non-covalent binding site indicate that binding (dissociation) of the substrate (product) may energetically contribute to the proton-translocating activity of ... | 2008 | 18471432 |
| electron flow through proteins. | electron transfers in photosynthesis and respiration commonly occur between metal-containing cofactors that are separated by large molecular distances. employing laser flash-quench triggering methods, we have shown that 20-å, coupling-limited fe(ii) to ru(iii) and cu(i) to ru(iii) electron tunneling in ru-modified cytochromes and blue copper proteins can occur on the microsecond timescale both in solutions and crystals. redox equivalents can be transferred even longer distances by multistep tunn ... | 2009 | 20161522 |
| a stable hyponitrite-bridged iron porphyrin complex. | the coupling of two nitric oxide (no) molecules in heme active sites is an important contributor to the conversion of no to nitrous oxide (n(2)o) by heme-containing enzymes. several formulations for the presumed heme-fe{n(2)o(2)}(n-) intermediates have been proposed previously, however, no crystal structures of heme-fe{n(2)o(2)}(n-) systems have been reported to date. we report the first isolation and characterization of a stable bimetallic hyponitrite iron porphyrin, [(oep)fe](2)(mu-n(2)o(2)), ... | 2009 | 19191487 |
| architecture of complex i and its implications for electron transfer and proton pumping. | proton pumping nadh:ubiquinone oxidoreductase (complex i) is the largest and remains by far the least understood enzyme complex of the respiratory chain. it consists of a peripheral arm harbouring all known redox active prosthetic groups and a membrane arm with a yet unknown number of proton translocation sites. the ubiquinone reduction site close to iron-sulfur cluster n2 at the interface of the 49-kda and psst subunits has been mapped by extensive site directed mutagenesis. independent lines o ... | 2009 | 19366614 |
| structures of membrane proteins. | in reviewing the structures of membrane proteins determined up to the end of 2009, we present in words and pictures the most informative examples from each family. we group the structures together according to their function and architecture to provide an overview of the major principles and variations on the most common themes. the first structures, determined 20 years ago, were those of naturally abundant proteins with limited conformational variability, and each membrane protein structure det ... | 2010 | 20667175 |
| seleno-detergent mad phasing of leukotriene c4 synthase in complex with dodecyl-β-d-selenomaltoside. | dodecyl-β-d-selenomaltoside (seddm) is a seleno-detergent with a β-glycosidic seleno-ether in place of the ether moiety in dodecyl-β-d-maltoside. seleno-detergents are candidates for heavy-atom agents in experimental phasing of membrane proteins in protein crystallography. crystals of a nuclear membrane-embedded enzyme, leukotriene c(4) synthase (ltc(4)s), in complex with seddm were prepared and a multiwavelength anomalous diffraction (mad) experiment was performed. the seddm in the ltc(4)s crys ... | 2011 | 22139193 |
| 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 ... | 2011 | 21963794 |
| 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 ... | 2011 | 21963794 |
| a bioinformatics classifier and database for heme-copper oxygen reductases. | heme-copper oxygen reductases (hcos) are the last enzymatic complexes of most aerobic respiratory chains, reducing dioxygen to water and translocating up to four protons across the inner mitochondrial membrane (eukaryotes) or cytoplasmatic membrane (prokaryotes). the number of completely sequenced genomes is expanding exponentially, and concomitantly, the number and taxonomic distribution of hco sequences. these enzymes were initially classified into three different types being this classificati ... | 2011 | 21559461 |
| spectral identification of intermediates generated during the reaction of dioxygen with the wild-type and eq(i-286) mutant of rhodobacter sphaeroides cytochrome c oxidase. | cytochrome c oxidase from rhodobacter sphaeroides is frequently used to model the more complex mitochondrial enzyme. the o(2) reduction in both enzymes is generally described by a unidirectional mechanism involving the sequential formation of the ferrous-oxy complex (compound a), the p(r) state, the oxyferryl f form, and the oxidized state. in this study we investigated the reaction of dioxygen with the wild-type reduced r. sphaeroides cytochrome oxidase and the eq(i-286) mutant using the co flo ... | 2012 | 23057757 |
| roles of subunit nuok (nd4l) in the energy-transducing mechanism of escherichia coli ndh-1 (nadh:quinone oxidoreductase). | the bacterial h(+)-translocating nadh:quinone oxidoreductase (ndh-1) catalyzes electron transfer from nadh to quinone coupled with proton pumping across the cytoplasmic membrane. the nuok subunit (counterpart of the mitochondrial nd4l subunit) is one of the seven hydrophobic subunits in the membrane domain and bears three transmembrane segments (tm1-3). two glutamic residues located in the adjacent transmembrane helices of nuok are important for the energy coupled activity of ndh-1. in particula ... | 2012 | 23105119 |
| cell-free protein synthesis: the state of the art. | cell-free protein synthesis harnesses the synthetic power of biology, programming the ribosomal translational machinery of the cell to create macromolecular products. like pcr, which uses cellular replication machinery to create a dna amplifier, cell-free protein synthesis is emerging as a transformative technology with broad applications in protein engineering, biopharmaceutical development, and post-genomic research. by breaking free from the constraints of cell-based systems, it takes the nex ... | 2012 | 23086573 |
| cell-free protein synthesis: the state of the art. | cell-free protein synthesis harnesses the synthetic power of biology, programming the ribosomal translational machinery of the cell to create macromolecular products. like pcr, which uses cellular replication machinery to create a dna amplifier, cell-free protein synthesis is emerging as a transformative technology with broad applications in protein engineering, biopharmaceutical development, and post-genomic research. by breaking free from the constraints of cell-based systems, it takes the nex ... | 2012 | 23086573 |
| copper starvation-inducible protein for cytochrome oxidase biogenesis in bradyrhizobium japonicum. | microarray analysis of bradyrhizobium japonicum grown under copper limitation uncovered five genes named pcuabcde, which are co-transcribed and co-regulated as an operon. the predicted gene products are periplasmic proteins (pcua, pcuc, and pcud), a tonb-dependent outer membrane receptor (pcub), and a cytoplasmic membrane-integral protein (pcue). homologs of pcuc and pcue had been discovered in other bacteria, namely pcu(a)c and ycnj, where they play a role in cytochrome oxidase biogenesis and c ... | 2012 | 23012364 |
| spectroscopic and kinetic investigation of the fully reduced and mixed valence states of ba3-cytochrome c oxidase from thermus thermophilus: a fourier transform infrared (ftir) and time-resolved step-scan ftir study. | the complete understanding of a molecular mechanism of action requires the thermodynamic and kinetic characterization of different states and intermediates. cytochrome c oxidase reduces o(2) to h(2)o, a reaction coupled to proton translocation across the membrane. therefore, it is necessary to undertake a thorough characterization of the reduced form of the enzyme and the determination of the electron transfer processes and pathways between the redox-active centers. in this study fourier transfo ... | 2012 | 22927441 |
| a three-dimensional topology of complex i inferred from evolutionary correlations. | the quaternary structure of eukaryotic nadh:ubiquinone oxidoreductase (complex i), the largest complex of the oxidative phosphorylation, is still mostly unresolved. furthermore, it is unknown where transiently bound assembly factors interact with complex i. we therefore asked whether the evolution of complex i contains information about its 3d topology and the binding positions of its assembly factors. we approached these questions by correlating the evolutionary rates of eukaryotic complex i su ... | 2012 | 22857522 |
| structural determinants of the β-selectivity of a bacterial aminotransferase. | chiral β-amino acids occur as constituents of various natural and synthetic compounds with potentially useful bioactivities. the pyridoxal 5'-phosphate (plp)-dependent s-selective transaminase from mesorhizobium sp. strain luk (mesat) is a fold type i aminotransferase that can be used for the preparation of enantiopure β-phe and derivatives thereof. using x-ray crystallography, we solved structures of mesat in complex with (s)-β-phe, (r)-3-amino-5-methylhexanoic acid, 2-oxoglutarate, and the inh ... | 2012 | 22745123 |
| from static structure to living protein: computational analysis of cytochrome c oxidase main-chain flexibility. | crystallographic structure and deuterium accessibility comparisons of cco in different redox states have suggested conformational changes of mechanistic significance. to predict the intrinsic flexibility and low energy motions in cco, this work has analyzed available high-resolution crystallographic structures with proflex and elnémo computational methods. the results identify flexible regions and potential conformational changes in cco that correlate well with published structural and biochemic ... | 2012 | 22824280 |
| structure, function, and assembly of heme centers in mitochondrial respiratory complexes. | the sequential flow of electrons in the respiratory chain, from a low reduction potential substrate to o(2), is mediated by protein-bound redox cofactors. in mitochondria, hemes-together with flavin, iron-sulfur, and copper cofactors-mediate this multi-electron transfer. hemes, in three different forms, are used as a protein-bound prosthetic group in succinate dehydrogenase (complex ii), in bc(1) complex (complex iii) and in cytochrome c oxidase (complex iv). the exact function of heme b in comp ... | 2012 | 22554985 |
| mechanistic stoichiometry of proton translocation by cytochrome cbb3. | cytochrome cbb(3) belongs to the superfamily of respiratory heme-copper oxidases that couple the reduction of molecular oxygen to proton translocation across the bacterial or mitochondrial membrane. the cbb(3)-type enzymes are found only in bacteria, and are both structurally and functionally the most distant from their mitochondrial counterparts. the mechanistic h(+)/e(-) stoichiometry of proton translocation in these cbb(3)-type cytochrome c oxidases has remained controversial. a stoichiometri ... | 2012 | 22529361 |
| product-controlled steady-state kinetics between cytochrome aa(3) from rhodobacter sphaeroides and equine ferrocytochrome c analyzed by a novel spectrophotometric approach. | cytochrome c oxidase (cco) catalyzes the reduction of molecular oxygen to water using ferrocytochrome c (cyt c(2+)) as the electron donor. in this study, the oxidation of horse cyt c(2+) by cco from rhodobacter sphaeroides, was monitored using stopped-flow spectrophotometry. a novel analytic procedure was applied in which the spectra were deconvoluted into the reduced and oxidized forms of cyt c by a least-squares fitting method, yielding the reaction rates at various concentrations of cyt c(2+) ... | 2012 | 22516686 |
| electron transfer in subunit nuoi (tyky) of escherichia coli nadh:quinone oxidoreductase (ndh-1). | bacterial proton-translocating nadh:quinone oxidoreductase (ndh-1) consists of a peripheral and a membrane domain. the peripheral domain catalyzes the electron transfer from nadh to quinone through a chain of seven iron-sulfur (fe/s) clusters. subunit nuoi in the peripheral domain contains two [4fe-4s] clusters (n6a and n6b) and plays a role in bridging the electron transfer from cluster n5 to the terminal cluster n2. we constructed mutants for eight individual cys-coordinating fe/s clusters. wi ... | 2012 | 22474289 |
| exploring the proton pump and exit pathway for pumped protons in cytochrome ba3 from thermus thermophilus. | the heme-copper oxygen reductases are redox-driven proton pumps. in the current work, the effects of mutations in a proposed exit pathway for pumped protons are examined in the ba(3)-type oxygen reductase from thermus thermophilus, leading from the propionates of heme a(3) to the interface between subunits i and ii. recent studies have proposed important roles for his376 and asp372, both of which are hydrogen-bonded to propionate-a of heme a(3), and for glu126(ii) (subunit ii), which is hydrogen ... | 2012 | 22431640 |
| density functional study for the bridged dinuclear center based on a high-resolution x-ray crystal structure of ba3 cytochrome c oxidase from thermus thermophilus. | strong electron density for a peroxide type dioxygen species bridging the fea3 and cub dinuclear center (dnc) was observed in the high-resolution (1.8 å) x-ray crystal structures (pdb entries 3s8g and 3s8f) of ba3 cytochrome c oxidase (cco) from thermus thermophilus. the crystals represent the as-isolated x-ray photoreduced cco structures. the bridging peroxide was proposed to arise from the recombination of two radiation-produced ho(•) radicals formed either very near to or even in the space be ... | 2013 | 24262070 |
| investigating the function of [2fe-2s] cluster n1a, the off-pathway cluster in complex i, by manipulating its reduction potential. | nadh:quinone oxidoreductase (complex i) couples nadh oxidation and quinone reduction to proton translocation across an energy-transducing membrane. all complexes i contain a flavin to oxidize nadh, seven iron-sulfur clusters to transfer electrons from the flavin to quinone and an eighth cluster (n1a) on the opposite side of the flavin. the role of cluster n1a is unknown, but escherichia coli complex i has an unusually high-potential cluster n1a and its reduced flavin produces h2o2, not superoxid ... | 2013 | 23980528 |
| molecular mechanism and physiological role of active-deactive transition of mitochondrial complex i. | the unique feature of mitochondrial complex i is the so-called a/d transition (active-deactive transition). the a-form catalyses rapid oxidation of nadh by ubiquinone (k ~104 min-1) and spontaneously converts into the d-form if the enzyme is idle at physiological temperatures. such deactivation occurs in vitro in the absence of substrates or in vivo during ischaemia, when the ubiquinone pool is reduced. the d-form can undergo reactivation given both nadh and ubiquinone availability during slow ( ... | 2013 | 24059527 |
| the nitric-oxide reductase from paracoccus denitrificans uses a single specific proton pathway. | the no reductase from paracoccus denitrificans reduces no to n2o (2no + 2h(+) + 2e(-) → n2o + h2o) with electrons donated by periplasmic cytochrome c (cytochrome c-dependent no reductase; cnor). cnors are members of the heme-copper oxidase superfamily of integral membrane proteins, comprising the o2-reducing, proton-pumping respiratory enzymes. in contrast, although no reduction is as exergonic as o2 reduction, there are no protons pumped in cnor, and in addition, protons needed for no reduction ... | 2013 | 24014024 |
| axial interactions in the mixed-valent cua active site and role of the axial methionine in electron transfer. | within cu-containing electron transfer active sites, the role of the axial ligand in type 1 sites is well defined, yet its role in the binuclear mixed-valent cua sites is less clear. recently, the mutation of the axial met to leu in a cua site engineered into azurin (cua az) was found to have a limited effect on e(0) relative to this mutation in blue copper (bc). detailed low-temperature absorption and magnetic circular dichroism, resonance raman, and electron paramagnetic resonance studies on c ... | 2013 | 23964128 |
| comparative genomics in acid mine drainage biofilm communities reveals metabolic and structural differentiation of co-occurring archaea. | metal sulfide mineral dissolution during bioleaching and acid mine drainage (amd) formation creates an environment that is inhospitable to most life. despite dominance by a small number of bacteria, amd microbial biofilm communities contain a notable variety of coexisting and closely related euryarchaea, most of which have defied cultivation efforts. for this reason, we used metagenomics to analyze variation in gene content that may contribute to niche differentiation among co-occurring amd arch ... | 2013 | 23865623 |
| characterization of the nitric oxide reductase from thermus thermophilus. | nitrous oxide (n2o) is a powerful greenhouse gas implicated in climate change. the dominant source of atmospheric n2o is incomplete biological dentrification, and the enzymes responsible for the release of n2o are no reductases. it was recently reported that ambient emissions of n2o from the great boiling spring in the united states great basin are high, and attributed to incomplete denitrification by thermus thermophilus and related bacterial species [hedlund bp, et al. (2011) geobiology 9(6)47 ... | 2013 | 23858452 |
| post-translational modifications near the quinone binding site of mammalian complex i. | complex i (nadh:ubiquinone oxidoreductase) in mammalian mitochondria is an l-shaped assembly of 44 protein subunits with one arm buried in the inner membrane of the mitochondrion and the orthogonal arm protruding about 100 å into the matrix. the protruding arm contains the binding sites for nadh, the primary acceptor of electrons flavin mononucleotide (fmn), and a chain of seven iron-sulfur clusters that carries the electrons one at a time from fmn to a coenzyme q molecule bound in the vicinity ... | 2013 | 23836892 |
| combined effect of loss of the caa3 oxidase and crp regulation drives shewanella to thrive in redox-stratified environments. | shewanella species are a group of facultative gram-negative microorganisms with remarkable respiration abilities that allow the use of a diverse array of terminal electron acceptors (ea). like most bacteria, s. oneidensis possesses multiple terminal oxidases, including two heme-copper oxidases (caa3- and cbb3-type) and a bd-type quinol oxidase. as aerobic respiration is energetically favored, mechanisms underlying the fact that these microorganisms thrive in redox-stratified environments remain ... | 2013 | 23575370 |
| semiquinone and cluster n6 signals in his-tagged proton-translocating nadh:ubiquinone oxidoreductase (complex i) from escherichia coli. | nadh:ubiquinone oxidoreductase (complex i) pumps protons across the membrane using downhill redox energy. the escherichia coli complex i consists of 13 different subunits named nuoa-n coded by the nuo operon. due to the low abundance of the protein and some difficulty with the genetic manipulation of its large ~15-kb operon, purification of e. coli complex i has been technically challenging. here, we generated a new strain in which a polyhistidine sequence was inserted upstream of nuoe in the op ... | 2013 | 23543743 |
| the mechanism of ubihydroquinone oxidation at the qo-site of the cytochrome bc1 complex. | 1. recent results suggest that the major flux is carried by a monomeric function, not by an intermonomer electron flow. 2. the bifurcated reaction at the qo-site involves sequential partial processes, - a rate limiting first electron transfer generating a semiquinone (sq) intermediate, and a rapid second electron transfer in which the sq is oxidized by the low potential chain. 3. the rate constant for the first step in a strongly endergonic, proton-first-then-electron mechanism, is given by a ma ... | 2013 | 23396004 |
| etmb-rbf: discrimination of metal-binding sites in electron transporters based on rbf networks with pssm profiles and significant amino acid pairs. | cellular respiration is the process by which cells obtain energy from glucose and is a very important biological process in living cell. as cells do cellular respiration, they need a pathway to store and transport electrons, the electron transport chain. the function of the electron transport chain is to produce a trans-membrane proton electrochemical gradient as a result of oxidation-reduction reactions. in these oxidation-reduction reactions in electron transport chains, metal ions play very i ... | 2013 | 23405059 |
| ligand access to the active site in thermus thermophilus ba(3) and bovine heart aa(3) cytochrome oxidases. | knowledge of the structure and dynamics of the ligand channel(s) in heme-copper oxidases is critical for understanding how the protein environment modulates the functions of these enzymes. using photolabile no and o(2) carriers, we recently found that no and o(2) binding in thermus thermophilus (tt) ba(3) is ~10 times faster than in the bovine enzyme, indicating that inherent structural differences affect ligand access in these enzymes. using x-ray crystallography, time-resolved optical absorpti ... | 2013 | 23282175 |
| atypical features of thermus thermophilus succinate:quinone reductase. | the thermus thermophilus succinate:quinone reductase (sqr), serving as the respiratory complex ii, has been homologously produced under the control of a constitutive promoter and subsequently purified. the detailed biochemical characterization of the resulting wild type (wt-rcii) and his-tagged (rcii-his(8)-sdhb and rcii-sdhb-his(6)) complex ii variants showed the same properties as the native enzyme with respect to the subunit composition, redox cofactor content and sensitivity to the inhibitor ... | 2013 | 23308253 |
| conserved amino acid residues of the nuod segment important for structure and function of escherichia coli ndh-1 (complex i). | the nuod segment (homologue of mitochondrial 49 kda subunit) of the proton-translocating nadh:quinone oxidoreductase (complex i/ndh-1) from escherichia coli is in the hydrophilic domain and bears many highly conserved amino acid residues. the three-dimensional structural model of ndh-1 suggests that the nuod segment, together with the neighboring subunits, constitutes a putative quinone binding cavity. we used the homologous dna recombination technique to clarify the role of selected key amino a ... | 2014 | 25545070 |
| conserved amino acid residues of the nuod segment important for structure and function of escherichia coli ndh-1 (complex i). | the nuod segment (homologue of mitochondrial 49 kda subunit) of the proton-translocating nadh:quinone oxidoreductase (complex i/ndh-1) from escherichia coli is in the hydrophilic domain and bears many highly conserved amino acid residues. the three-dimensional structural model of ndh-1 suggests that the nuod segment, together with the neighboring subunits, constitutes a putative quinone binding cavity. we used the homologous dna recombination technique to clarify the role of selected key amino a ... | 2014 | 25545070 |
| re-evaluation of the near infrared spectra of mitochondrial cytochrome c oxidase: implications for non invasive in vivo monitoring of tissues. | we re-determined the near infrared (nir) spectral signatures (650-980nm) of the different cytochrome c oxidase redox centres, in the process separating them into their component species. we confirm that the primary contributor to the oxidase nir spectrum between 700 and 980nm is cupric cua, which in the beef heart enzyme has a maximum at 835nm. the 655nm band characterises the fully oxidised haem a3/cub binuclear centre; it is bleached either when one or more electrons are added to the binuclear ... | 2014 | 25175349 |
| how periplasmic thioredoxin tlpa reduces bacterial copper chaperone scoi and cytochrome oxidase subunit ii (coxb) prior to metallation. | two critical cysteine residues in the copper-a site (cu(a)) on subunit ii (coxb) of bacterial cytochrome c oxidase lie on the periplasmic side of the cytoplasmic membrane. as the periplasm is an oxidizing environment as compared with the reducing cytoplasm, the prediction was that a disulfide bond formed between these cysteines must be eliminated by reduction prior to copper insertion. we show here that a periplasmic thioredoxin (tlpa) acts as a specific reductant not only for the cu(2+) transfe ... | 2014 | 25274631 |
| a third subunit in ancestral cytochrome c-dependent nitric oxide reductases. | reduction of no to n2o by denitrifiying bacteria is catalyzed either by a monomeric quinol-nitric oxide reductase (qnor) or by a heterodimeric cytochrome c-dependent nitric oxide reductase (cnor). in ancient thermophilic bacteria belonging to the thermales and aquificales phylogenetic groups, the cluster encoding the cnor includes a small third gene (norh), in addition to those encoding homologues to the subunits of a typical cnor (norc and norb). we show in thermus thermophilus that the three g ... | 2014 | 24907324 |
| evidence for distinct electron transfer processes in terminal oxidases from different origin by means of protein film voltammetry. | cytochrome aa3 from paracoccus denitrificans and cytochrome ba3 from thermus thermophilus, two distinct members of the heme-copper oxidase superfamily, were immobilized on electrodes modified with gold nanoparticles. this procedure allowed us to achieve direct electron transfer between the enzyme and the gold nanoparticles and to obtain evidence for different electrocatalytic properties of the two enzymes. the ph dependence and thermostability reveal that the enzymes are highly adapted to their ... | 2014 | 25054669 |
| the pathway of o₂to the active site in heme-copper oxidases. | the route of o₂to and from the high-spin heme in heme-copper oxidases has generally been believed to emulate that of carbon monoxide (co). time-resolved and stationary infrared experiments in our laboratories of the fully reduced co-bound enzymes, as well as transient optical absorption saturation kinetics studies as a function of co pressure, have provided strong support for co binding to cub⁺ on the pathway to and from the high-spin heme. the presence of co on cub⁺ suggests that o₂binding may ... | 2014 | 24998308 |
| the pathway of o₂to the active site in heme-copper oxidases. | the route of o₂to and from the high-spin heme in heme-copper oxidases has generally been believed to emulate that of carbon monoxide (co). time-resolved and stationary infrared experiments in our laboratories of the fully reduced co-bound enzymes, as well as transient optical absorption saturation kinetics studies as a function of co pressure, have provided strong support for co binding to cub⁺ on the pathway to and from the high-spin heme. the presence of co on cub⁺ suggests that o₂binding may ... | 2014 | 24998308 |
| characterisation of the active/de-active transition of mitochondrial complex i. | oxidation of nadh in the mitochondrial matrix of aerobic cells is catalysed by mitochondrial complex i. the regulation of this mitochondrial enzyme is not completely understood. an interesting characteristic of complex i from some organisms is the ability to adopt two distinct states: the so-called catalytically active (a) and the de-active, dormant state (d). the a-form in situ can undergo de-activation when the activity of the respiratory chain is limited (i.e. in the absence of oxygen). the m ... | 2014 | 24569053 |
| linking chemical electron-proton transfer to proton pumping in cytochrome c oxidase: broken-symmetry dft exploration of intermediates along the catalytic reaction pathway of the iron-copper dinuclear complex. | after a summary of the problem of coupling electron and proton transfer to proton pumping in cytochrome c oxidase, we present the results of our earlier and recent density functional theory calculations for the dinuclear fe-a3-cub reaction center in this enzyme. a specific catalytic reaction wheel diagram is constructed from the calculations, based on the structures and relative energies of the intermediate states of the reaction cycle. a larger family of tautomers/protonation states is generate ... | 2014 | 24960612 |
| conserved glycine 232 in the ligand channel of ba3 cytochrome oxidase from thermus thermophilus. | knowing how the protein environment modulates ligand pathways and redox centers in the respiratory heme-copper oxidases is fundamental for understanding the relationship between the structure and function of these enzymes. in this study, we investigated the reactions of o2 and no with the fully reduced g232v mutant of ba3 cytochrome c oxidase from thermus thermophilus (tt ba3) in which a conserved glycine residue in the o2 channel of the enzyme was replaced with a bulkier valine residue. previou ... | 2014 | 24937405 |
| molecular characterization of novel pyridoxal-5'-phosphate-dependent enzymes from the human microbiome. | pyridoxal-5'-phosphate or plp, the active form of vitamin b6, is a highly versatile cofactor that participates in a large number of mechanistically diverse enzymatic reactions in basic metabolism. plp-dependent enzymes account for ∼1.5% of most prokaryotic genomes and are estimated to be involved in ∼4% of all catalytic reactions, making this an important class of enzymes. here, we structurally and functionally characterize three novel plp-dependent enzymes from bacteria in the human microbiome: ... | 2014 | 24888348 |
| parallel pathways for nitrite reduction during anaerobic growth in thermus thermophilus. | respiratory reduction of nitrate and nitrite is encoded in thermus thermophilus by the respective transferable gene clusters. nitrate is reduced by a heterotetrameric nitrate reductase (nar) encoded along transporters and regulatory signal transduction systems within the nitrate respiration conjugative element (nce). the nitrite respiration cluster (nic) encodes homologues of nitrite reductase (nir) and nitric oxide reductase (nor). the expression and role of the nirsjm genes in nitrite respirat ... | 2014 | 24443532 |
| a sco protein among the hypothetical proteins of bacillus lehensis g1: its 3d macromolecular structure and association with cytochrome c oxidase. | at least a quarter of any complete genome encodes for hypothetical proteins (hps) which are largely non-similar to other known, well-characterized proteins. predicting and solving their structures and functions is imperative to aid understanding of any given organism as a complete biological system. the present study highlights the primary effort to classify and cluster 1202 hps of bacillus lehensis g1 alkaliphile to serve as a platform to mine and select specific hp(s) to be studied further in ... | 2014 | 24641837 |
| crystal structure of an (r)-selective ω-transaminase from aspergillus terreus. | chiral amines are important building blocks for the synthesis of pharmaceutical products, fine chemicals, and agrochemicals. ω-transaminases are able to directly synthesize enantiopure chiral amines by catalysing the transfer of an amino group from a primary amino donor to a carbonyl acceptor with pyridoxal 5'-phosphate (plp) as cofactor. in nature, (s)-selective amine transaminases are more abundant than the (r)-selective enzymes, and therefore more information concerning their structures is av ... | 2014 | 24498081 |
| the mononuclear molybdenum enzymes. | | 2014 | 24467397 |
| a biochemical approach to study the role of the terminal oxidases in aerobic respiration in shewanella oneidensis mr-1. | the genome of the facultative anaerobic γ-proteobacterium shewanella oneidensis mr-1 encodes for three terminal oxidases: a bd-type quinol oxidase and two heme-copper oxidases, a a-type cytochrome c oxidase and a cbb 3-type oxidase. in this study, we used a biochemical approach and directly measured oxidase activities coupled to mass-spectrometry analysis to investigate the physiological role of the three terminal oxidases under aerobic and microaerobic conditions. our data revealed that the cbb ... | 2014 | 24466040 |
| transferable denitrification capability of thermus thermophilus. | laboratory-adapted strains of thermus spp. have been shown to require oxygen for growth, including the model strains t. thermophilus hb27 and hb8. in contrast, many isolates of this species that have not been intensively grown under laboratory conditions keep the capability to grow anaerobically with one or more electron acceptors. the use of nitrogen oxides, especially nitrate, as electron acceptors is one of the most widespread capabilities among these facultative strains. in this process, nit ... | 2014 | 24141123 |
| fold and flexibility: what can proteins' mechanical properties tell us about their folding nucleus? | the determination of a protein's folding nucleus, i.e. a set of native contacts playing an important role during its folding process, remains an elusive yet essential problem in biochemistry. in this work, we investigate the mechanical properties of 70 protein structures belonging to 14 protein families presenting various folds using coarse-grain brownian dynamics simulations. the resulting rigidity profiles combined with multiple sequence alignments show that a limited set of rigid residues, wh ... | 2015 | 26577596 |
| binuclear cu(a) formation in biosynthetic models of cu(a) in azurin proceeds via a novel cu(cys)2his mononuclear copper intermediate. | cu(a) is a binuclear electron transfer (et) center found in cytochrome c oxidases (ccos), nitrous oxide reductases (n₂ors), and nitric oxide reductase (nor). in these proteins, the cu(a) centers facilitate efficient et (ket > 10⁴s⁻¹) under low thermodynamic driving forces (10-90 mv). while the structure and functional properties of cu(a) are well understood, a detailed mechanism of the incorporation of copper into the protein and the identity of the intermediates formed during the cu(a) maturati ... | 2015 | 26352296 |
| characterization of clinically identified mutations in ndufv1, the flavin-binding subunit of respiratory complex i, using a yeast model system. | dysfunctions in mitochondrial complex i (nadh:ubiquinone oxidoreductase) are both genetically and clinically highly diverse and a major cause of human mitochondrial diseases. the genetic determinants of individual clinical cases are increasingly being described, but how these genetic defects affect complex i on the molecular and cellular level, and have different clinical consequences in different individuals, is little understood. furthermore, without molecular-level information innocent geneti ... | 2015 | 26345448 |
| broken symmetry dft calculations/analysis for oxidized and reduced dinuclear center in cytochrome c oxidase: relating structures, protonation states, energies, and mössbauer properties in ba3 thermus thermophilus. | the fea3(3+)···cub(2+) dinuclear center (dnc) structure of the as-isolated oxidized ba3 cytochrome c oxidase (cco) from thermus thermophilus (tt) is still not fully understood. when the proteins are initially crystallized in the oxidized state, they typically become radiolyticly reduced through x-ray irradiation. several x-ray crystal structures of reduced ba3 cco from tt are available. however, depending on whether the crystals were prepared in a lipidic cubic phase environment or in detergent ... | 2015 | 26192749 |
| during cytochrome c maturation ccmi chaperones the class i apocytochromes until the formation of their b-type cytochrome intermediates. | the c-type cytochromes are electron transfer proteins involved in energy transduction. they have heme-binding (cxxch) sites that covalently ligate heme b via thioether bonds and are classified into different classes based on their protein folds and the locations and properties of their cofactors. rhodobacter capsulatus produces various c-type cytochromes using the cytochrome c maturation (ccm) system i, formed from the ccmabcdefghi proteins. ccmi, a component of the heme ligation complex ccmfhi, ... | 2015 | 25979338 |
| reassessment of the transhydrogenase/malate shunt pathway in clostridium thermocellum atcc 27405 through kinetic characterization of malic enzyme and malate dehydrogenase. | clostridium thermocellum produces ethanol as one of its major end products from direct fermentation of cellulosic biomass. therefore, it is viewed as an attractive model for the production of biofuels via consolidated bioprocessing. however, a better understanding of the metabolic pathways, along with their putative regulation, could lead to improved strategies for increasing the production of ethanol. in the absence of an annotated pyruvate kinase in the genome, alternate means of generating py ... | 2015 | 25616802 |
| mutation of a single residue in the ba3 oxidase specifically impairs protonation of the pump site. | the ba3-type cytochrome c oxidase from thermus thermophilus is a membrane-bound protein complex that couples electron transfer to o2 to proton translocation across the membrane. to elucidate the mechanism of the redox-driven proton pumping, we investigated the kinetics of electron and proton transfer in a structural variant of the ba3 oxidase where a putative "pump site" was modified by replacement of asp372 by ile. in this structural variant, proton pumping was uncoupled from internal electron ... | 2015 | 25733886 |
| cryo-em structures of the autoinhibited e. coli atp synthase in three rotational states. | a molecular model that provides a framework for interpreting the wealth of functional information obtained on the e. coli f-atp synthase has been generated using cryo-electron microscopy. three different states that relate to rotation of the enzyme were observed, with the central stalk's ε subunit in an extended autoinhibitory conformation in all three states. the fo motor comprises of seven transmembrane helices and a decameric c-ring and invaginations on either side of the membrane indicate th ... | 2016 | 28001127 |
| energy conversion, redox catalysis and generation of reactive oxygen species by respiratory complex i. | complex i (nadh:ubiquinone oxidoreductase) is critical for respiration in mammalian mitochondria. it oxidizes nadh produced by the krebs' tricarboxylic acid cycle and β-oxidation of fatty acids, reduces ubiquinone, and transports protons to contribute to the proton-motive force across the inner membrane. complex i is also a significant contributor to cellular oxidative stress. in complex i, nadh oxidation by a flavin mononucleotide, followed by intramolecular electron transfer along a chain of i ... | 2016 | 26721206 |
| characterization of the nqo5 subunit of bacterial complex i in the isolated state. | the subunits that comprise bacterial complex i (nadh:ubiquinone oxidoreductase) are also found in more complicated mitochondrial enzymes in eukaryotic organisms. although the nqo5 subunit is one of these conserved components and important for the formation of complex, it has been little studied. here, we report structure analyses of isolated nqo5 from thermus thermophilus. biochemical studies indicated that the c-terminal region following the 30-kd subunit motif is disordered in the isolated sta ... | 2016 | 27398308 |
| evidence for fast electron transfer between the high-spin haems in cytochrome bd-i from escherichia coli. | cytochrome bd-i is one of the three proton motive force-generating quinol oxidases in the o2-dependent respiratory chain of escherichia coli. it contains one low-spin haem (b558) and the two high-spin haems (b595 and d) as the redox-active cofactors. in order to examine the flash-induced intraprotein reverse electron transfer (the so-called ''electron backflow''), co was photolyzed from the ferrous haem d in one-electron reduced (b5583+b5953+d2+-co) cytochrome bd-i, and the fully reduced (b5582+ ... | 2016 | 27152644 |
| different functions of phylogenetically distinct bacterial complex i isozymes. | nadh:quinone oxidoreductase (complex i) is a bioenergetic enzyme that transfers electrons from nadh to quinone, conserving the energy of this reaction by contributing to the proton motive force. while the importance of nadh oxidation to mitochondrial aerobic respiration is well documented, the contribution of complex i to bacterial electron transport chains has been tested in only a few species. here, we analyze the function of two phylogenetically distinct complex i isozymes in rhodobacter spha ... | 2016 | 26833419 |
| investigating the proton donor in the no reductase from paracoccus denitrificans. | variant nomenclature: the variants were made in the norb subunit if not indicated by the superscript c, which are variants in the norc subunit (e.g. e122a = exchange of glu-122 in norb for an ala, e71cd; exchange of glu-71 in norc for an asp). bacterial no reductases (nors) are integral membrane proteins from the heme-copper oxidase superfamily. most heme-copper oxidases are proton-pumping enzymes that reduce o2 as the last step in the respiratory chain. with electrons from cytochrome c, no redu ... | 2016 | 27030968 |
| models for the a subunits of the thermus thermophilus v/a-atpase and saccharomyces cerevisiae v-atpase enzymes by cryo-em and evolutionary covariance. | rotary atpases couple atp synthesis or hydrolysis to proton translocation across a membrane. however, understanding proton translocation has been hampered by a lack of structural information for the membrane-embedded a subunit. the v/a-atpase from the eubacterium thermus thermophilus is similar in structure to the eukaryotic v-atpase but has a simpler subunit composition and functions in vivo to synthesize atp rather than pump protons. we determined the t. thermophilus v/a-atpase structure by cr ... | 2016 | 26951669 |
| cysteine biosynthesis in lactobacillus casei: identification and characterization of a serine acetyltransferase. | in bacteria, cysteine can be synthesized from serine by two steps involving an l-serine o-acetyltransferase (sat) and a cysteine synthase (cysk). while cysk is found in the publicly available annotated genome from lactobacillus casei atcc 334, a gene encoding sat (cyse) is missing. in this study, we found that various strains of l. casei grew in a chemically defined medium containing sulfide as the sole sulfur source, indicating the presence of a serine o-acetyltransferase. the gene lying upstre ... | 2016 | 26790714 |
| improved dna sequencing accuracy and detection of heterozygous alleles using manganese citrate and different fluorescent dye terminators. | the use of dideoxynucleotide triphosphates labeled with different fluorescent dyes (dye terminators) is the most versatile method for automated dna sequencing. however, variation in peak heights reduces base-calling accuracy and limits heterozygous allele detection, favoring use of dye-labeled primers for this purpose. we have discovered that the addition of a manganese salt to the pe applied biosystems dye-terminator sequencing kits overcomes these limitations for the older rhodamine dyes as we ... | 1999 | 10400927 |
| demystified... molecular pathology in oncology. | in the past 10 years, molecular biology has found major applications in pathology, particularly in oncology. this has been a field of enormous expansion, where pure science has found a place in clinical practice and is now of everyday use in any academic unit. this demystified review will discuss the techniques used in molecular pathology and then provide examples of how these can be used in oncology. | 2002 | 12456768 |
| perspectives on biotechnological applications of archaea. | many archaea colonize extreme environments. they include hyperthermophiles, sulfur-metabolizing thermophiles, extreme halophiles and methanogens. because extremophilic microorganisms have unusual properties, they are a potentially valuable resource in the development of novel biotechnological processes. despite extensive research, however, there are few existing industrial applications of either archaeal biomass or archaeal enzymes. this review summarizes current knowledge about the biotechnolog ... | 2002 | 15803645 |
| nucleic acid recognition by ob-fold proteins. | the ob-fold domain is a compact structural motif frequently used for nucleic acid recognition. structural comparison of all ob-fold/nucleic acid complexes solved to date confirms the low degree of sequence similarity among members of this family while highlighting several structural sequence determinants common to most of these ob-folds. loops connecting the secondary structural elements in the ob-fold core are extremely variable in length and in functional detail. however, certain features of l ... | 2003 | 12598368 |
| reverse transcriptase at bacterial telomeres. | | 2004 | 15454610 |
| antibacterial peptide microcin j25 inhibits transcription by binding within and obstructing the rna polymerase secondary channel. | the antibacterial peptide microcin j25 (mccj25) inhibits transcription by bacterial rna polymerase (rnap). biochemical results indicate that inhibition of transcription occurs at the level of ntp uptake or ntp binding by rnap. genetic results indicate that inhibition of transcription requires an extensive determinant, comprising more than 50 amino acid residues, within the rnap secondary channel (also known as the "ntp-uptake channel" or "pore"). biophysical results indicate that inhibition of t ... | 2004 | 15200952 |
| a low-cost open-source snp genotyping platform for association mapping applications. | association mapping aimed at identifying dna polymorphisms that contribute to variation in complex traits entails genotyping a large number of single-nucleotide polymorphisms (snps) in a very large panel of individuals. few technologies, however, provide inexpensive high-throughput genotyping. here, we present an efficient approach developed specifically for genotyping large fixed panels of diploid individuals. the cost-effective, open-source nature of our methodology may make it particularly at ... | 2005 | 16356268 |
| a unique trna recognition mechanism of caenorhabditis elegans mitochondrial ef-tu2. | nematode mitochondria expresses two types of extremely truncated trnas that are specifically recognized by two distinct elongation factor tu (ef-tu) species named ef-tu1 and ef-tu2. this is unlike the canonical ef-tu molecule that participates in the standard protein biosynthesis systems, which basically recognizes all elongator trnas. ef-tu2 specifically recognizes ser-trna(ser) that lacks a d arm but has a short t arm. our previous study led us to speculate the lack of the d arm may be essenti ... | 2005 | 16113240 |
| molecular mimicry: quantitative methods to study structural similarity between protein and rna. | with rapidly increasing availability of three-dimensional structures, one major challenge for the post-genome era is to infer the functions of biological molecules based on their structural similarity. while quantitative studies of structural similarity between the same type of biological molecules (e.g., protein vs. protein) have been carried out intensively, the comparable study of structural similarity between different types of biological molecules (e.g., protein vs. rna) remains unexplored. ... | 2005 | 16043503 |
| correcting errors in synthetic dna through consensus shuffling. | although efficient methods exist to assemble synthetic oligonucleotides into genes and genomes, these suffer from the presence of 1-3 random errors/kb of dna. here, we introduce a new method termed consensus shuffling and demonstrate its use to significantly reduce random errors in synthetic dna. in this method, errors are revealed as mismatches by re-hybridization of the population. the dna is fragmented, and mismatched fragments are removed upon binding to an immobilized mismatch binding prote ... | 2005 | 15800206 |