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the smpb c-terminal tail helps tmrna to recognize and enter stalled ribosomes.in bacteria, transfer-messenger rna (tmrna) and smpb comprise the most common and effective system for rescuing stalled ribosomes. ribosomes stall on mrna transcripts lacking stop codons and are rescued as the defective mrna is swapped for the tmrna template in a process known as trans-translation. the tmrna-smpb complex is recruited to the ribosome independent of a codon-anticodon interaction. given that the ribosome uses robust discriminatory mechanisms to select against non-cognate trnas duri ...201425228900
the evolutionary journey of argonaute proteins.argonaute proteins are conserved throughout all domains of life. recently characterized prokaryotic argonaute proteins (pagos) participate in host defense by dna interference, whereas eukaryotic argonaute proteins (eagos) control a wide range of processes by rna interference. here we review molecular mechanisms of guide and target binding by argonaute proteins, and describe how the conformational changes induced by target binding lead to target cleavage. on the basis of structural comparisons an ...201425192263
heteronuclear transverse and longitudinal relaxation in ax4 spin systems: application to (15)n relaxations in (15)nh4(+).the equations that describe the time-evolution of transverse and longitudinal (15)n magnetisations in tetrahedral ammonium ions, (15)nh4(+), are derived from the bloch-wangsness-redfield density operator relaxation theory. it is assumed that the relaxation of the spin-states is dominated by (1) the intra-molecular (15)n-(1)h and (1)h-(1)h dipole-dipole interactions and (2) interactions of the ammonium protons with remote spins, which also include the contribution to the relaxations that arise fr ...201425128779
acetylome analysis reveals diverse functions of lysine acetylation in mycobacterium tuberculosis.the lysine acetylation of proteins is a reversible post-translational modification that plays a critical regulatory role in both eukaryotes and prokaryotes. mycobacterium tuberculosis is a facultative intracellular pathogen and the causative agent of tuberculosis. increasing evidence shows that lysine acetylation may play an important role in the pathogenesis of m. tuberculosis. however, only a few acetylated proteins of m. tuberculosis are known, presenting a major obstacle to understanding the ...201425180227
holliday junction resolvases.four-way dna intermediates, called holliday junctions (hjs), can form during meiotic and mitotic recombination, and their removal is crucial for chromosome segregation. a group of ubiquitous and highly specialized structure-selective endonucleases catalyze the cleavage of hjs into two disconnected dna duplexes in a reaction called hj resolution. these enzymes, called hj resolvases, have been identified in bacteria and their bacteriophages, archaea, and eukaryotes. in this review, we discuss fund ...201425183833
a bridge between the aminoacylation and editing domains of leucyl-trna synthetase is crucial for its synthetic activity.leucyl-trna synthetases (leurss) catalyze the linkage of leucine with trna(leu). leurs contains a catalysis domain (aminoacylation) and a cp1 domain (editing). cp1 is inserted 35 å from the aminoacylation domain. aminoacylation and editing require cp1 to swing to the coordinated conformation. the neck between the cp1 domain and the aminoacylation domain is defined as the cp1 hairpin. the location of the cp1 hairpin suggests a crucial role in the cp1 swing and domain-domain interaction. here, the ...201425051973
evidence for a hexaheteromeric methylenetetrahydrofolate reductase in moorella thermoacetica.moorella thermoacetica can grow with h₂ and co₂, forming acetic acid from 2 co₂ via the wood-ljungdahl pathway. all enzymes involved in this pathway have been characterized to date, except for methylenetetrahydrofolate reductase (metf). we report here that the m. thermoacetica gene that putatively encodes this enzyme, metf, is part of a transcription unit also containing the genes hdrcba, mvhd, and metv. metf copurified with the other five proteins encoded in the unit in a hexaheteromeric comple ...201425002540
acetyl coenzyme a synthetase is acetylated on multiple lysine residues by a protein acetyltransferase with a single gcn5-type n-acetyltransferase (gnat) domain in saccharopolyspora erythraea.reversible lysine acetylation (rla) is used by cells of all domains of life to modulate protein function. to date, bacterial acetylation/deacetylation systems have been studied in a few bacteria (e.g., salmonella enterica, bacillus subtilis, escherichia coli, erwinia amylovora, mycobacterium tuberculosis, and geobacillus kaustophilus), but little is known about rla in antibiotic-producing actinomycetes. here, we identify the gcn5-like protein acetyltransferase acua of saccharopolyspora erythraea ...201424957627
reco protein initiates dna recombination and strand annealing through two alternative dna binding mechanisms.recombination mediator proteins (rmps) are important for genome stability in all organisms. several rmps support two alternative reactions: initiation of homologous recombination and dna annealing. we examined mechanisms of rmps in both reactions with mycobacterium smegmatis reco (msreco) and demonstrated that msreco interacts with ssdna by two distinct mechanisms. zinc stimulates msreco binding to ssdna during annealing, whereas the recombination function is zinc-independent and is regulated by ...201425170075
crystal structure of a conserved hypothetical protein mj0927 from methanocaldococcus jannaschii reveals a novel quaternary assembly in the nif3 family.a nif3 family protein of methanocaldococcus jannaschii, mj0927, is highly conserved from bacteria to humans. although several structures of bacterial nif3 proteins are known, no structure representing archaeal nif3 has yet been reported. the crystal structure of methanocaldococcus jannaschii mj0927 was determined at 2.47 å resolution to understand the structural differences between the bacterial and archaeal nif3 proteins. intriguingly, mj0927 is found to adopt an unusual assembly comprising a t ...201425243119
evolution of transfer rna and the origin of the translation system. 201425221573
escherichia coli d-malate dehydrogenase, a generalist enzyme active in the leucine biosynthesis pathway.the enzymes of the β-decarboxylating dehydrogenase superfamily catalyze the oxidative decarboxylation of d-malate-based substrates with various specificities. here, we show that, in addition to its natural function affording bacterial growth on d-malate as a carbon source, the d-malate dehydrogenase of escherichia coli (ecdmla) naturally expressed from its chromosomal gene is capable of complementing leucine auxotrophy in a leub(-) strain lacking the paralogous isopropylmalate dehydrogenase enzy ...201425160617
interplay between oxygen and fe-s cluster biogenesis: insights from the suf pathway.iron-sulfur (fe-s) cluster metalloproteins conduct essential functions in nearly all contemporary forms of life. the nearly ubiquitous presence of fe-s clusters and the fundamental requirement for fe-s clusters in both aerobic and anaerobic archaea, bacteria, and eukarya suggest that these clusters were likely integrated into central metabolic pathways early in the evolution of life prior to the widespread oxidation of earth's atmosphere. intriguingly, fe-s cluster-dependent metabolism is sensit ...201425153801
how lamina-associated polypeptide 1 (lap1) activates torsin.lamina-associated polypeptide 1 (lap1) resides at the nuclear envelope and interacts with torsins, poorly understood endoplasmic reticulum (er)-localized aaa+ atpases, through a conserved, perinuclear domain. we determined the crystal structure of the perinuclear domain of human lap1. lap1 possesses an atypical aaa+ fold. while lap1 lacks canonical nucleotide binding motifs, its strictly conserved arginine 563 is positioned exactly where the arginine finger of canonical aaa+ atpases is found. ba ...201425149450
crisprstrand: predicting repeat orientations to determine the crrna-encoding strand at crispr loci.the discovery of crispr-cas systems almost 20 years ago rapidly changed our perception of the bacterial and archaeal immune systems. crispr loci consist of several repetitive dna sequences called repeats, inter-spaced by stretches of variable length sequences called spacers. this crispr array is transcribed and processed into multiple mature rna species (crrnas). a single crrna is integrated into an interference complex, together with crispr-associated (cas) proteins, to bind and degrade invadin ...201425161238
dna binding properties of the small cascade subunit csa5.crispr-cas systems provide immunity against viral attacks in archaeal and bacterial cells. type i systems employ a cas protein complex termed cascade, which utilizes small crispr rnas to detect and degrade the exogenic dna. a small sequence motif, the pam, marks the foreign substrates. previously, a recombinant type i-a cascade complex from the archaeon thermoproteus tenax was shown to target and degrade dna in vitro, dependent on a native pam sequence. here, we present the biochemical analysis ...201425148031
comparison of histidine recognition in human and trypanosomatid histidyl-trna synthetases.as part of a project aimed at obtaining selective inhibitors and drug-like compounds targeting trna synthetases from trypanosomatids, we have elucidated the crystal structure of human cytosolic histidyl-trna synthetase (hs-chisrs) in complex with histidine in order to be able to compare human and parasite enzymes. the resultant structure of hs-chisrs•his represents the substrate-bound state (h-state) of the enzyme. it provides an interesting opportunity to compare with ligand-free and imidazole- ...201425151410
dissection of structural and functional requirements that underlie the interaction of erdj3 protein with substrates in the endoplasmic reticulum.erdj3, a mammalian endoplasmic reticulum (er) hsp40/dnaj family member, binds unfolded proteins, transfers them to bip, and concomitantly stimulates bip atpase activity. however, the requirements for erdj3 binding to and release from substrates in cells are not well understood. we found that erdj3 homodimers that cannot stimulate the atpase activity of bip (qpd mutants) bound to unfolded er proteins under steady state conditions in much greater amounts than wild-type erdj3. this was due to reduc ...201425143379
investigating the thermostability of succinate: quinone oxidoreductase enzymes by direct electrochemistry at swnts-modified electrodes and ftir spectroscopy.succinate: quinone reductases (sqrs) are the enzymes that couple the oxidation of succinate and the reduction of quinones in the respiratory chain of prokaryotes and eukaryotes. herein, we compare the temperature-dependent activity and structural stability of two sqrs, the first from the mesophilic bacterium escherichia coli and the second from the thermophilic bacterium thermus thermophilus, using a combined electrochemical and infrared spectroscopy approach. direct electron transfer was achiev ...201425139263
structure analysis of free and bound states of an rna aptamer against ribosomal protein s8 from bacillus anthracis.several protein-targeted rna aptamers have been identified for a variety of applications and although the affinities of numerous protein-aptamer complexes have been determined, the structural details of these complexes have not been widely explored. we examined the structural accommodation of an rna aptamer that binds bacterial r-protein s8. the core of the primary binding site for s8 on helix 21 of 16s rrna contains a pair of conserved base triples that mold the sugar-phosphate backbone to s8. ...201425140011
formation and structures of groel:groes2 chaperonin footballs, the protein-folding functional form.the groe chaperonins assist substrate protein (sp) folding by cycling through several conformational states. with each cycle the sp is, in turn, captured, unfolded, briefly encapsulated (t1/2 ∼ 1 s), and released by the chaperonin complex. the protein-folding functional form is the us-football-shaped groel:groes2 complex. we report structures of two such "football" complexes to ∼ 3.7-å resolution; one is empty whereas the other contains encapsulated sp in both chambers. although encapsulated sp ...201425136110
a proton wire to couple aminoacyl-trna accommodation and peptide-bond formation on the ribosome.during peptide-bond formation on the ribosome, the α-amine of an aminoacyl-trna attacks the ester carbonyl carbon of a peptidyl-trna to yield a peptide lengthened by one amino acid. although the ribosome's contribution to catalysis is predominantly entropic, the lack of high-resolution structural data for the complete active site in complex with full-length ligands has made it difficult to assess how the ribosome might influence the pathway of the reaction. here, we present crystal structures of ...201425132179
total synthesis of viridicatumtoxin b and analogues thereof: strategy evolution, structural revision, and biological evaluation.the details of the total synthesis of viridicatumtoxin b (1) are described. initial synthetic strategies toward this intriguing tetracycline antibiotic resulted in the development of key alkylation and lewis acid-mediated spirocyclization reactions to form the hindered ef spirojunction, as well as michael-dieckmann reactions to set the a and c rings. the use of an aromatic a-ring substrate, however, was found to be unsuitable for the introduction of the requisite hydroxyl groups at carbons 4a an ...201425317739
structural insights into +1 frameshifting promoted by expanded or modification-deficient anticodon stem loops.maintenance of the correct reading frame on the ribosome is essential for accurate protein synthesis. here, we report structures of the 70s ribosome bound to frameshift suppressor trna(sufa6) and n1-methylguanosine at position 37 (m(1)g37) modification-deficient anticodon stem loop(pro), both of which cause the ribosome to decode 4 rather than 3 nucleotides, resulting in a +1 reading frame. our results reveal that decoding at +1 suppressible codons causes suppressor trna(sufa6) to undergo a rear ...201425128388
structural biology. crystal structure of a crispr rna-guided surveillance complex bound to a ssdna target.in prokaryotes, rna derived from type i and type iii crispr loci direct large ribonucleoprotein complexes to destroy invading bacteriophage and plasmids. in escherichia coli, this 405-kilodalton complex is called cascade. we report the crystal structure of cascade bound to a single-stranded dna (ssdna) target at a resolution of 3.03 angstroms. the structure reveals that the crispr rna and target strands do not form a double helix but instead adopt an underwound ribbon-like structure. this noncan ...201425123481
dead-box rna helicase domains exhibit a continuum between complete functional independence and high thermodynamic coupling in nucleotide and rna duplex recognition.dead-box helicases catalyze the non-processive unwinding of double-stranded rna (dsrna) at the expense of adenosine triphosphate (atp) hydrolysis. nucleotide and rna binding and unwinding are mediated by the reca domains of the helicase core, but their cooperation in these processes remains poorly understood. we therefore investigated dsrna and nucleotide binding by the helicase cores and the isolated n- and c-terminal reca domains (reca_n, reca_c) of the dead-box proteins hera and yxin by stead ...201425123660
allosteric regulation of a protein acetyltransferase in micromonospora aurantiaca by the amino acids cysteine and arginine.act domains (amino acid-binding domains) are linked to a wide range of metabolic enzymes that are regulated by amino acid concentration. seventy proteins with act-gcn5-related n-acetyltransferase (gnat) domain organization were found in actinomycetales. in this study, we investigate the act-containing gnat acetyltransferase, micau_1670 (makat), from micromonospora aurantiaca atcc 27029. arginine and cysteine were identified as ligands by monitoring the conformational changes that occur upon amin ...201425124041
discovery of a eukaryotic pyrroloquinoline quinone-dependent oxidoreductase belonging to a new auxiliary activity family in the database of carbohydrate-active enzymes.pyrroloquinoline quinone (pqq) is a redox cofactor utilized by a number of prokaryotic dehydrogenases. not all prokaryotic organisms are capable of synthesizing pqq, even though it plays important roles in the growth and development of many organisms, including humans. the existence of pqq-dependent enzymes in eukaryotes has been suggested based on homology studies or the presence of pqq-binding motifs, but there has been no evidence that such enzymes utilize pqq as a redox cofactor. however, du ...201425121592
dna binding polarity, dimerization, and atpase ring remodeling in the cmg helicase of the eukaryotic replisome.the cdc45/mcm2-7/gins (cmg) helicase separates dna strands during replication in eukaryotes. how the cmg is assembled and engages dna substrates remains unclear. using electron microscopy, we have determined the structure of the cmg in the presence of atpγs and a dna duplex bearing a 3' single-stranded tail. the structure shows that the mcm subunits of the cmg bind preferentially to single-stranded dna, establishes the polarity by which dna enters into the mcm2-7 pore, and explains how cdc45 hel ...201425117490
dark matter in archaeal genomes: a rich source of novel mobile elements, defense systems and secretory complexes.microbial genomes encompass a sizable fraction of poorly characterized, narrowly spread fast-evolving genes. using sensitive methods for sequences comparison and protein structure prediction, we performed a detailed comparative analysis of clusters of such genes, which we denote "dark matter islands", in archaeal genomes. the dark matter islands comprise up to 20% of archaeal genomes and show remarkable heterogeneity and diversity. nevertheless, three classes of entities are common in these geno ...201425113822
comparative genomics reveals adaptations of a halotolerant thaumarchaeon in the interfaces of brine pools in the red sea.the bottom of the red sea harbors over 25 deep hypersaline anoxic basins that are geochemically distinct and characterized by vertical gradients of extreme physicochemical conditions. because of strong changes in density, particulate and microbial debris get entrapped in the brine-seawater interface (bsi), resulting in increased dissolved organic carbon, reduced dissolved oxygen toward the brines and enhanced microbial activities in the bsi. these features coupled with the deep-sea prevalence of ...201425105904
comparative genomics reveals adaptations of a halotolerant thaumarchaeon in the interfaces of brine pools in the red sea.the bottom of the red sea harbors over 25 deep hypersaline anoxic basins that are geochemically distinct and characterized by vertical gradients of extreme physicochemical conditions. because of strong changes in density, particulate and microbial debris get entrapped in the brine-seawater interface (bsi), resulting in increased dissolved organic carbon, reduced dissolved oxygen toward the brines and enhanced microbial activities in the bsi. these features coupled with the deep-sea prevalence of ...201425105904
l-lactic acid production from d-xylose with candida sonorensis expressing a heterologous lactate dehydrogenase encoding gene.bioplastics, like polylactic acid (pla), are renewable alternatives for petroleum-based plastics. lactic acid, the monomer of pla, has traditionally been produced biotechnologically with bacteria. with genetic engineering, yeast have the potential to replace bacteria in biotechnological lactic acid production, with the benefits of being acid tolerant and having simple nutritional requirements. lactate dehydrogenase genes have been introduced to various yeast to demonstrate this potential. import ...201425104116
ndho, a subunit of nadph dehydrogenase, destabilizes medium size complex of the enzyme in synechocystis sp. strain pcc 6803.two mutants that grew faster than the wild-type (wt) strain under high light conditions were isolated from synechocystis sp. strain pcc 6803 transformed with a transposon-bearing library. both mutants had a tag in ssl1690 encoding ndho. deletion of ndho increased the activity of nadph dehydrogenase (ndh-1)-dependent cyclic electron transport around photosystem i (ndh-cet), while overexpression decreased the activity. although deletion and overexpression of ndho did not have significant effects o ...201425107904
discovery and validation of novel and distinct rna regulators for ribosomal protein s15 in diverse bacterial phyla.autogenous cis-regulators of ribosomal protein synthesis play a critical role in maintaining the stoichiometry of ribosome components. structured portions within an mrna transcript typically interact with specific ribosomal proteins to prevent expression of the entire operon, thus balancing levels of ribosomal proteins across transcriptional units. three distinct rna structures from different bacterial phyla have demonstrated interactions with s15 to regulate gene expression; however, these rnas ...201425104606
comparative single-cell genomics reveals potential ecological niches for the freshwater aci actinobacteria lineage.members of the aci lineage of actinobacteria are the most abundant microorganisms in most freshwater lakes; however, our understanding of the keys to their success and their role in carbon and nutrient cycling in freshwater systems has been hampered by the lack of pure cultures and genomes. we obtained draft genome assemblies from 11 single cells representing three aci tribes (aci-a1, aci-a7, aci-b1) from four temperate lakes in the united states and europe. comparative analysis of aci sags and ...201425093637
the interface between escherichia coli elongation factor tu and aminoacyl-trna.nineteen of the highly conserved residues of escherichia coli (e. coli) elongation factor tu (ef-tu) that form the binding interface with aa-trna were mutated to alanine to better understand how modifying the thermodynamic properties of ef-tu-trna interaction can affect the decoding properties of the ribosome. comparison of δδg(o) values for binding ef-tu to aa-trna show that the majority of the interface residues stabilize the ternary complex and their thermodynamic contribution can depend on t ...201425094027
structural and functional basis of transcriptional regulation by tetr family protein cprb from s. coelicolor a3(2).antibiotic production and resistance pathways in streptomyces are dictated by the interplay of transcriptional regulatory proteins that trigger downstream responses via binding to small diffusible molecules. to decipher the mode of dna binding and the associated allosteric mechanism in the sub-class of transcription factors that are induced by γ-butyrolactones, we present the crystal structure of cprb in complex with the consensus dna element to a resolution of 3.25 å. binding of the dna results ...201425092919
afal: a web service for profiling amino acids surrounding ligands in proteins.with advancements in crystallographic technology and the increasing wealth of information populating structural databases, there is an increasing need for prediction tools based on spatial information that will support the characterization of proteins and protein-ligand interactions. herein, a new web service is presented termed amino acid frequency around ligand (afal) for determining amino acids type and frequencies surrounding ligands within proteins deposited in the protein data bank and for ...201425085083
crystal structure of the full-length atpase gspe from the vibrio vulnificus type ii secretion system in complex with the cytoplasmic domain of gspl.the type ii secretion system (t2ss) is present in many gram-negative bacteria and is responsible for secreting a large number of folded proteins, including major virulence factors, across the outer membrane. the t2ss consists of 11-15 different proteins most of which are present in multiple copies in the assembled secretion machinery. the atpase gspe, essential for the functioning of the t2ss, contains three domains (n1e, n2e and cte) of which the n1e domain is associated with the cytoplasmic do ...201425092625
bioinformatic screening of autoimmune disease genes and protein structure prediction with fams for drug discovery.autoimmune diseases are often intractable because their causes are unknown. identifying which genes contribute to these diseases may allow us to understand the pathogenesis, but it is difficult to determine which genes contribute to disease. recently, epigenetic information has been considered to activate/deactivate disease-related genes. thus, it may also be useful to study epigenetic information that differs between healthy controls and patients with autoimmune disease. among several types of ...201423855671
structural analysis of base substitutions in thermus thermophilus 16s rrna conferring streptomycin resistance.streptomycin is a bactericidal antibiotic that induces translational errors. it binds to the 30s ribosomal subunit, interacting with ribosomal protein s12 and with 16s rrna through contacts with the phosphodiester backbone. to explore the structural basis for streptomycin resistance, we determined the x-ray crystal structures of 30s ribosomal subunits from six streptomycin-resistant mutants of thermus thermophilus both in the apo form and in complex with streptomycin. base substitutions at highl ...201424820088
nature's recipe for splitting inteins.protein splicing in trans by split inteins has increasingly become a powerful protein-engineering tool for protein ligation, both in vivo and in vitro. over 100 naturally occurring and artificially engineered split inteins have been reported for protein ligation using protein trans-splicing. here, we review the current status of the reported split inteins in order to delineate an empirical or rational strategy for constructing new split inteins suitable for various applications in biotechnology ...201425096198
characterization of two homologous 2'-o-methyltransferases showing different specificities for their trna substrates.the 2'-o-methylation of the nucleoside at position 32 of trna is found in organisms belonging to the three domains of life. unrelated enzymes catalyzing this modification in bacteria (trmj) and eukarya (trm7) have already been identified, but until now, no information is available for the archaeal enzyme. in this work we have identified the methyltransferase of the archaeon sulfolobus acidocaldarius responsible for the 2'-o-methylation at position 32. this enzyme is a homolog of the bacterial tr ...201424951554
building a stable rna u-turn with a protonated cytidine.the u-turn is a classical three-dimensional rna folding motif first identified in the anticodon and t-loops of trnas. it also occurs frequently as a building block in other functional rna structures in many different sequence and structural contexts. u-turns induce sharp changes in the direction of the rna backbone and often conform to the 3-nt consensus sequence 5'-unr-3' (n = any nucleotide, r = purine). the canonical u-turn motif is stabilized by a hydrogen bond between the n3 imino group of ...201424951555
new extremophilic lipases and esterases from metagenomics.lipolytic enzymes catalyze the hydrolysis of ester bonds in the presence of water. in media with low water content or in organic solvents, they can catalyze synthetic reactions such as esterification and transesterification. lipases and esterases, in particular those from extremophilic origin, are robust enzymes, functional under the harsh conditions of industrial processes owing to their inherent thermostability and resistance towards organic solvents, which combined with their high chemo-, reg ...201424588890
antibiotic drugs targeting bacterial rnas.rnas have diverse structures that include bulges and internal loops able to form tertiary contacts or serve as ligand binding sites. the recent increase in structural and functional information related to rnas has put them in the limelight as a drug target for small molecule therapy. in addition, the recognition of the marked difference between prokaryotic and eukaryotic rrna has led to the development of antibiotics that specifically target bacterial rrna, reduce protein translation and thereby ...201426579393
synergism between a foldase and an unfoldase: reciprocal dependence between the thioredoxin-like activity of dnaj and the polypeptide-unfolding activity of dnak.the role of bacterial hsp40, dnaj, is to co-chaperone the binding of misfolded or alternatively folded proteins to bacterial hsp70, dnak, which is an atp-fuelled unfolding chaperone. in addition to its dnak targeting activity, dnaj has a weak thiol-reductase activity. in between the substrate-binding domain and the j-domain anchor to dnak, dnaj has a unique domain with four conserved cxxc motives that bind two zn(2+) and partly contribute to polypeptide binding. here, we deleted in dnaj this zn- ...201425988148
phylobiochemical characterization of class-ib aspartate/prephenate aminotransferases reveals evolution of the plant arogenate phenylalanine pathway.the aromatic amino acid phe is required for protein synthesis and serves as the precursor of abundant phenylpropanoid plant natural products. while phe is synthesized from prephenate exclusively via a phenylpyruvate intermediate in model microbes, the alternative pathway via arogenate is predominant in plant phe biosynthesis. however, the molecular and biochemical evolution of the plant arogenate pathway is currently unknown. here, we conducted phylogenetically informed biochemical characterizat ...201425070637
transcription regulation mechanisms of bacteriophages: recent advances and future prospects.phage diversity significantly contributes to ecology and evolution of new bacterial species through horizontal gene transfer. therefore, it is essential to understand the mechanisms underlying phage-host interactions. after initial infection, the phage utilizes the transcriptional machinery of the host to direct the expression of its own genes. this review presents a view on the transcriptional regulation mechanisms of bacteriophages, and its contribution to phage diversity and classification. t ...201425482231
evidence for loss of a partial flagellar glycolytic pathway during trypanosomatid evolution.classically viewed as a cytosolic pathway, glycolysis is increasingly recognized as a metabolic pathway exhibiting surprisingly wide-ranging variations in compartmentalization within eukaryotic cells. trypanosomatid parasites provide an extreme view of glycolytic enzyme compartmentalization as several glycolytic enzymes are found exclusively in peroxisomes. here, we characterize trypanosoma brucei flagellar proteins resembling glyceraldehyde-3-phosphate dehydrogenase (gapdh) and phosphoglycerate ...201425050549
an update on complex i assembly: the assembly of players.defects in complex i assembly is one of the emerging underlying causes of severe mitochondrial disorders. the assembly of complex i has been difficult to understand due to its large size, dual genetic control and the number of proteins involved. mutations in complex i subunits as well as assembly factors have been reported to hinder its assembly and give rise to a range of mitochondria disorders. in this review, we summarize the recent progress made in understanding the complex i assembly pathwa ...201425030182
the hypusine-containing translation factor eif5a.in addition to the small and large ribosomal subunits, aminoacyl-trnas, and an mrna, cellular protein synthesis is dependent on translation factors. the eukaryotic translation initiation factor 5a (eif5a) and its bacterial ortholog elongation factor p (ef-p) were initially characterized based on their ability to stimulate methionyl-puromycin (met-pmn) synthesis, a model assay for protein synthesis; however, the function of these factors in cellular protein synthesis has been difficult to resolve ...201425029904
structures of yeast 80s ribosome-trna complexes in the rotated and nonrotated conformations.the structural understanding of eukaryotic translation lags behind that of translation on bacterial ribosomes. here, we present two subnanometer resolution structures of s. cerevisiae 80s ribosome complexes formed with either one or two trnas and bound in response to an mrna fragment containing the kozak consensus sequence. the ribosomes adopt two globally different conformations that are related to each other by the rotation of the small subunit. comparison with bacterial ribosome complexes rev ...201425043550
biochemical and cellular analysis of human variants of the dyt1 dystonia protein, torsina/tor1a.early-onset dystonia is associated with the deletion of one of a pair of glutamic acid residues (c.904_906delgag/c.907_909delgag; p.glu302del/glu303del; δe 302/303) near the carboxyl-terminus of torsina, a member of the aaa(+) protein family that localizes to the endoplasmic reticulum lumen and nuclear envelope. this deletion commonly underlies early-onset dyt1 dystonia. while the role of the disease-causing mutation, torsinaδe, has been established through genetic association studies, it is muc ...201424930953
road rules for traffic on dna-systematic analysis of transcriptional roadblocking in vivo.genomic dna is bound by many proteins that could potentially impede elongation of rna polymerase (rnap), but the factors determining the magnitude of transcriptional roadblocking in vivo are poorly understood. through systematic experiments and modeling, we analyse how roadblocking by the lac repressor (laci) in escherichia coli cells is controlled by promoter firing rate, the concentration and affinity of the roadblocker protein, the transcription-coupled repair protein mfd, and promoter-roadbl ...201425034688
the cca-end of p-trna contacts both the human rpl36al and the a-site bound translation termination factor erf1 at the peptidyl transferase center of the human 80s ribosome.we have demonstrated previously that the e-site specific protein rpl36al present in human ribosomes can be crosslinked with the cca-end of a p-trna in situ. here we report the following: (i) we modeled rpl36al into the structure of the archaeal ortholog rpl44e extracted from the known x-ray structure of the 50s subunit of haloarcula marismortui. superimposing the obtained rpl36al structure with that of p/e trna observed in eukaryotic 80s ribosomes suggested that rpl36al might in addition to its ...201425191528
reverse gyrase--recent advances and current mechanistic understanding of positive dna supercoiling.reverse gyrases are topoisomerases that introduce positive supercoils into dna in an atp-dependent reaction. they consist of a helicase domain and a topoisomerase domain that closely cooperate in catalysis. the mechanism of the functional cooperation of these domains has remained elusive. recent studies have shown that the helicase domain is a nucleotide-regulated conformational switch that alternates between an open conformation with a low affinity for double-stranded dna, and a closed state wi ...201425013168
molecular characterization of the na+/h+-antiporter nhaa from salmonella typhimurium.na+/h+ antiporters are integral membrane proteins that are present in almost every cell and in every kingdom of life. they are essential for the regulation of intracellular ph-value, na+-concentration and cell volume. these secondary active transporters exchange sodium ions against protons via an alternating access mechanism, which is not understood in full detail. na+/h+ antiporters show distinct species-specific transport characteristics and regulatory properties that correlate with respective ...201425010413
phages preying on bacillus anthracis, bacillus cereus, and bacillus thuringiensis: past, present and future.many bacteriophages (phages) have been widely studied due to their major role in virulence evolution of bacterial pathogens. however, less attention has been paid to phages preying on bacteria from the bacillus cereus group and their contribution to the bacterial genetic pool has been disregarded. therefore, this review brings together the main information for the b. cereus group phages, from their discovery to their modern biotechnological applications. a special focus is given to phages infect ...201425010767
structure and function of pseudoknots involved in gene expression control.natural rna molecules can have a high degree of structural complexity but even the most complexly folded rnas are assembled from simple structural building blocks. among the simplest rna elements are double-stranded helices that participate in the formation of different folding topologies and constitute the major fraction of rna structures. one common folding motif of rna is a pseudoknot, defined as a bipartite helical structure formed by base-pairing of the apical loop in the stem-loop structur ...201425044223
structure of pneumococcal peptidoglycan hydrolase lytb reveals insights into the bacterial cell wall remodeling and pathogenesis.streptococcus pneumoniae causes a series of devastating infections in humans. previous studies have shown that the endo-β-n-acetylglucosaminidase lytb is critical for pneumococcal cell division and nasal colonization, but the biochemical mechanism of lytb action remains unknown. here we report the 1.65 å crystal structure of the catalytic domain (residues lys-375-asp-658) of lytb (termed lytbcat), excluding the choline binding domain. lytbcat consists of three structurally independent modules: s ...201425002590
ancient translation factor is essential for trna-dependent cysteine biosynthesis in methanogenic archaea.methanogenic archaea lack cysteinyl-trna synthetase; they synthesize cys-trna and cysteine in a trna-dependent manner. two enzymes are required: phosphoseryl-trna synthetase (seprs) forms phosphoseryl-trna(cys) (sep-trna(cys)), which is converted to cys-trna(cys) by sep-trna:cys-trna synthase (sepcyss). this represents the ancestral pathway of cys biosynthesis and coding in archaea. here we report a translation factor, sepcyse, essential for methanococcal cys biosynthesis; its deletion in methan ...201425002468
free-energy landscape of reverse trna translocation through the ribosome analyzed by electron microscopy density maps and molecular dynamics simulations.to understand the mechanism of reverse trna translocation in the ribosome, all-atom molecular dynamics simulations of the ribosome-trnas-mrna-efg complex were performed. the complex at the post-translocational state was directed towards the translocational and pre-translocational states by fitting the complex into cryo-em density maps. between a series of the fitting simulations, umbrella sampling simulations were performed to obtain the free-energy landscape. multistep structural changes, such ...201424999999
proteomic mapping of the human mitochondrial intermembrane space in live cells via ratiometric apex tagging.obtaining complete protein inventories for subcellular regions is a challenge that often limits our understanding of cellular function, especially for regions that are impossible to purify and are therefore inaccessible to traditional proteomic analysis. we recently developed a method to map proteomes in living cells with an engineered peroxidase (apex) that bypasses the need for organellar purification when applied to membrane-bound compartments; however, it was insufficiently specific when app ...201425002142
regulation of the mammalian elongation cycle by subunit rolling: a eukaryotic-specific ribosome rearrangement.the extent to which bacterial ribosomes and the significantly larger eukaryotic ribosomes share the same mechanisms of ribosomal elongation is unknown. here, we present subnanometer resolution cryoelectron microscopy maps of the mammalian 80s ribosome in the posttranslocational state and in complex with the eukaryotic eef1a⋅val-trna⋅gmppnp ternary complex, revealing significant differences in the elongation mechanism between bacteria and mammals. surprisingly, and in contrast to bacterial riboso ...201424995983
3'-biotin-tagged microrna-27 does not associate with argonaute proteins in cells.synthetic 3'-biotin-tagged micrornas (mirnas) have often been used to select interacting messenger rna (mrna) and noncoding rna (ncrna) targets. here, we examined the extent of association of 3'-end biotinylated mir-27 with argonaute (ago) proteins in transfected human cells using a coimmunoprecipitation assay followed by northern blot analysis. we report that biotinylated mir-27 does not efficiently associate with ago compared to unmodified mir-27. these results suggest that 3'-end biotin-modif ...201424821854
examining the gm18 and m(1)g modification positions in trna sequences.the trna structure contains conserved modifications that are responsible for its stability and are involved in the initiation and accuracy of the translation process. trna modification enzymes are prevalent in bacteria, archaea, and eukaryotes. trna gm18 methyltransferase (trmh) and trna m(1)g37 methyltransferase (trmd) are prevalent and essential enzymes in bacterial populations. trmh involves itself in methylation process at the 2'-oh group of ribose at the 18th position of guanosine (g) in tr ...201425031570
oral manifestations of hepatitis c virus infection.extrahepatic manifestations (ehms) of hepatitis c virus (hcv) infection can affect a variety of organ systems with significant morbidity and mortality. some of the most frequently reported ehm of hcv infection, involve the oral region predominantly or exclusively. oral lichen planus (olp) is a chronic inflammatory condition that is potentially malignant and represents cell-mediated reaction to a variety of extrinsic antigens, altered self-antigens, or super antigens. robust epidemiological evide ...201424976694
structural basis of transcription initiation by bacterial rna polymerase holoenzyme.the bacterial rna polymerase (rnap) holoenzyme containing σ factor initiates transcription at specific promoter sites by de novo rna priming, the first step of rna synthesis where rnap accepts two initiating ribonucleoside triphosphates (intps) and performs the first phosphodiester bond formation. we present the structure of de novo transcription initiation complex that reveals unique contacts of the intps bound at the transcription start site with the template dna and also with rnap and demonst ...201424973216
conjugation with polyamines enhances the antibacterial and anticancer activity of chloramphenicol.chloramphenicol (cam) is a broad-spectrum antibiotic, limited to occasional only use in developed countries because of its potential toxicity. to explore the influence of polyamines on the uptake and activity of cam into cells, a series of polyamine-cam conjugates were synthesized. both polyamine architecture and the position of cam-scaffold substitution were crucial in augmenting the antibacterial and anticancer potency of the synthesized conjugates. compounds 4 and 5, prepared by replacement o ...201424939899
crystal structure of clostridium acetobutylicum aspartate kinase (caak): an important allosteric enzyme for amino acids production.aspartate kinase (ak) is an enzyme which is tightly regulated through feedback control and responsible for the synthesis of 4-phospho-l-aspartate from l-aspartate. this intermediate step is at an important branch point where one path leads to the synthesis of lysine and the other to threonine, methionine and isoleucine. concerted feedback inhibition of ak is mediated by threonine and lysine and varies between the species. the crystal structure of biotechnologically important clostridium acetobut ...201425170437
macrolide antibiotics allosterically predispose the ribosome for translation arrest.translation arrest directed by nascent peptides and small cofactors controls expression of important bacterial and eukaryotic genes, including antibiotic resistance genes, activated by binding of macrolide drugs to the ribosome. previous studies suggested that specific interactions between the nascent peptide and the antibiotic in the ribosomal exit tunnel play a central role in triggering ribosome stalling. however, here we show that macrolides arrest translation of the truncated ermdl regulato ...201424961372
identification of promiscuous ene-reductase activity by mining structural databases using active site constellations.the exploitation of catalytic promiscuity and the application of de novo design have recently opened the access to novel, non-natural enzymatic activities. here we describe a structural bioinformatic method for predicting catalytic activities of enzymes based on three-dimensional constellations of functional groups in active sites ('catalophores'). as a proof-of-concept we identify two enzymes with predicted promiscuous ene-reductase activity (reduction of activated c-c double bonds) and compare ...201424954722
life at the border: adaptation of proteins to anisotropic membrane environment.this review discusses main features of transmembrane (tm) proteins which distinguish them from water-soluble proteins and allow their adaptation to the anisotropic membrane environment. we overview the structural limitations on membrane protein architecture, spatial arrangement of proteins in membranes and their intrinsic hydrophobic thickness, co-translational and post-translational folding and insertion into lipid bilayers, topogenesis, high propensity to form oligomers, and large-scale confor ...201424947665
homology modeling and comparative profiling of superoxide dismutase among extremophiles: exiguobacterium as a model organism.superoxide dismutase (sod), a well known antioxidant enzyme, is known to exert its presence across bacteria to humans. apart from their well-known antioxidant defense mechanisms, their association with various extremophiles in response to various stress conditions is poorly understood. here, we have discussed the conservation and the prevalence of sods among 21 representative extremophiles. a systematic investigation of aligned amino acid sequences of sod from all the selected extremophiles reve ...201425320445
xanthomonas campestris rpfb is a fatty acyl-coa ligase required to counteract the thioesterase activity of the rpff diffusible signal factor (dsf) synthase.in xanthomonas campestris pv. campestris (xcc), the proteins encoded by the rpf (regulator of pathogenicity factor) gene cluster produce and sense a fatty acid signal molecule called diffusible signalling factor (dsf, 2(z)-11-methyldodecenoic acid). rpfb was reported to be involved in dsf processing and was predicted to encode an acyl-coa ligase. we report that rpfb activates a wide range of fatty acids to their coa esters in vitro. moreover, rpfb can functionally replace the paradigm bacterial ...201424866092
lysine propionylation is a prevalent post-translational modification in thermus thermophilus.recent studies of protein post-translational modifications revealed that various types of lysine acylation occur in eukaryotic and bacterial proteins. lysine propionylation, a newly discovered type of acylation, occurs in several proteins, including some histones. in this study, we identified 361 propionylation sites in 183 mid-exponential phase and late stationary phase proteins from thermus thermophilus hb8, an extremely thermophilic eubacterium. functional classification of the propionylprote ...201424938286
the flavoprotein mcap0476 (rlmfo) catalyzes m5u1939 modification in mycoplasma capricolum 23s rrna.efficient protein synthesis in all organisms requires the post-transcriptional methylation of specific ribosomal ribonucleic acid (rrna) and transfer rna (trna) nucleotides. the methylation reactions are almost invariably catalyzed by enzymes that use s-adenosylmethionine (adomet) as the methyl group donor. one noteworthy exception is seen in some bacteria, where the conserved trna methylation at m5u54 is added by the enzyme trmfo using flavin adenine dinucleotide together with n5,n10-methylenet ...201424939895
the role of a novel auxiliary pocket in bacterial phenylalanyl-trna synthetase druggability.the antimicrobial activity of phenyl-thiazolylurea-sulfonamides against staphylococcus aureus phers are dependent upon phenylalanine levels in the extracellular fluids. inhibitor efficacy in animal models of infection is substantially diminished by dietary phenylalanine intake, thereby reducing the perceived clinical utility of this inhibitor class. the search for novel antibacterial compounds against gram-negative pathogens led to a re-evaluation of this phenomenon, which is shown here to be un ...201424936059
genomic and phenotypic attributes of novel salinivibrios from stromatolites, sediment and water from a high altitude lake.salinivibrios are moderately halophilic bacteria found in salted meats, brines and hypersaline environments. we obtained three novel conspecific salinivibrio strains closely related to s. costicola, from socompa lake, a high altitude hypersaline andean lake (approx. 3,570 meters above the sea level).201424927949
unravelling the structural and mechanistic basis of crispr-cas systems.bacteria and archaea have evolved sophisticated adaptive immune systems, known as crispr-cas (clustered regularly interspaced short palindromic repeats-crispr-associated proteins) systems, which target and inactivate invading viruses and plasmids. immunity is acquired by integrating short fragments of foreign dna into crispr loci, and following transcription and processing of these loci, the crispr rnas (crrnas) guide the cas proteins to complementary invading nucleic acid, which results in targ ...201424909109
the structural basis of transfer rna mimicry and conformational plasticity by a viral rna.rna is arguably the most functionally diverse biological macromolecule. in some cases a single discrete rna sequence performs multiple roles, and this can be conferred by a complex three-dimensional structure. such multifunctionality can also be driven or enhanced by the ability of a given rna to assume different conformational (and therefore functional) states. despite its biological importance, a detailed structural understanding of the paradigm of rna structure-driven multifunctionality is la ...201424909993
determination of kinetics and the crystal structure of a novel type 2 isopentenyl diphosphate: dimethylallyl diphosphate isomerase from streptococcus pneumoniae.isopentenyl diphosphate isomerase (idi) is a key enzyme in the isoprenoid biosynthetic pathway and is required for all organisms that synthesize isoprenoid metabolites from mevalonate. type 1 idi (idi-1) is a metalloprotein that is found in eukaryotes, whereas the type 2 isoform (idi-2) is a flavoenzyme found in bacteria that is completely absent from human. idi-2 from the pathogenic bacterium streptococcus pneumoniae was recombinantly expressed in escherichia coli. steady-state kinetic studies ...201424910111
archaeal tuc1/ncs6 homolog required for wobble uridine trna thiolation is associated with ubiquitin-proteasome, translation, and rna processing system homologs.while cytoplasmic trna 2-thiolation protein 1 (tuc1/ncs6) and ubiquitin-related modifier-1 (urm1) are important in the 2-thiolation of 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2u) at wobble uridines of trnas in eukaryotes, the biocatalytic roles and properties of ncs6/tuc1 and its homologs are poorly understood. here we present the first report of an ncs6 homolog of archaea (ncsa of haloferax volcanii) that is essential for maintaining cellular pools of thiolated trna(lys)uuu and for growth a ...201424906001
evidence for a catalytically and kinetically competent enzyme-substrate cross-linked intermediate in catalysis by lipoyl synthase.lipoyl synthase (ls) catalyzes the final step in lipoyl cofactor biosynthesis: the insertion of two sulfur atoms at c6 and c8 of an (n(6)-octanoyl)-lysyl residue on a lipoyl carrier protein (lcp). ls is a member of the radical sam superfamily, enzymes that use a [4fe-4s] cluster to effect the reductive cleavage of s-adenosyl-l-methionine (sam) to l-methionine and a 5'-deoxyadenosyl 5'-radical (5'-da(•)). in the ls reaction, two equivalents of 5'-da(•) are generated sequentially to abstract hydro ...201424901788
crystal structure of trna m(1)a58 methyltransferase trmi from aquifex aeolicus in complex with s-adenosyl-l-methionine.the n (1)-methyladenosine residue at position 58 of trna is found in the three domains of life, and contributes to the stability of the three-dimensional l-shaped trna structure. in thermophilic bacteria, this modification is important for thermal adaptation, and is catalyzed by the trna m(1)a58 methyltransferase trmi, using s-adenosyl-l-methionine (adomet) as the methyl donor. we present the 2.2 å crystal structure of trmi from the extremely thermophilic bacterium aquifex aeolicus, in complex w ...201424894648
condensation and localization of the partitioning protein parb on the bacterial chromosome.the parabs system mediates chromosome segregation and plasmid partitioning in many bacteria. as part of the partitioning mechanism, parb proteins form a nucleoprotein complex at pars sites. the biophysical basis underlying parb-dna complex formation and localization remains elusive. specifically, it is unclear whether parb spreads in 1d along dna or assembles into a 3d protein-dna complex. we show that a combination of 1d spreading bonds and a single 3d bridging bond between parb proteins consti ...201424927534
oxidation of cellular amino acid pools leads to cytotoxic mistranslation of the genetic code.aminoacyl-trna synthetases use a variety of mechanisms to ensure fidelity of the genetic code and ultimately select the correct amino acids to be used in protein synthesis. the physiological necessity of these quality control mechanisms in different environments remains unclear, as the cost vs benefit of accurate protein synthesis is difficult to predict. we show that in escherichia coli, a non-coded amino acid produced through oxidative damage is a significant threat to the accuracy of protein ...201424891238
nd3, nd1 and 39kda subunits are more exposed in the de-active form of bovine mitochondrial complex i.an intriguing feature of mitochondrial complex i from several species is the so-called a/d transition, whereby the idle enzyme spontaneously converts from the active (a) form to the de-active (d) form. the a/d transition plays an important role in tissue response to the lack of oxygen and hypoxic deactivation of the enzyme is one of the key regulatory events that occur in mitochondria during ischaemia. we demonstrate for the first time that the a/d conformational change of complex i does not aff ...201424560811
life-history constraints on the mechanisms that control the rate of ros production.the quest to understand why and how we age has led to numerous lines of investigation that have gradually converged to consider mitochondrial metabolism as a major player. during mitochondrial respiration a small and variable amount of the consumed oxygen is converted to reactive species of oxygen (ros). for many years, these ros have been perceived as harmful by-products of respiration. however, evidence from recent years indicates that ros fulfill important roles as cellular messengers. result ...201424955029
parb spreading requires dna bridging.the parabs system is a widely employed mechanism for plasmid partitioning and chromosome segregation in bacteria. parb binds to pars sites on plasmids and chromosomes and associates with broad regions of adjacent dna, a phenomenon known as spreading. although essential for parb function, the mechanism of spreading remains poorly understood. using single-molecule approaches, we discovered that bacillus subtilis parb (spo0j) is able to trap dna loops. point mutants in spo0j that disrupt dna bridgi ...201424829297
randomly selected suppressor mutations in genes for nadh : quinone oxidoreductase-1, which rescue motility of a salmonella ubiquinone-biosynthesis mutant strain.the primary mobile electron-carrier in the aerobic respiratory chain of salmonella is ubiquinone. demethylmenaquinone and menaquinone are alternative electron-carriers involved in anaerobic respiration. ubiquinone biosynthesis was disrupted in strains bearing deletions of the ubia or ubie genes. in soft tryptone agar both mutant strains swam poorly. however, the ubia deletion mutant strain produced suppressor mutant strains with somewhat rescued motility and growth. six independent suppressor mu ...201424692644
chemical synthesis of the 5-taurinomethyl(-2-thio)uridine modified anticodon arm of the human mitochondrial trna(leu(uur)) and trna(lys).5-taurinomethyluridine (τm(5)u) and 5-taurinomethyl-2-thiouridine (τm(5)s(2)u) are located at the wobble position of human mitochondrial (hmt) trna(leu(uur)) and trna(lys), respectively. both hypermodified units restrict decoding of the third codon letter to a and g. pathogenic mutations in the genes encoding hmt-trna(leu(uur)) and hmt-trna(lys) are responsible for the loss of the discussed modifications and, as a consequence, for the occurrence of severe mitochondrial dysfunctions (melas, merrf ...201424757169
non-nearest-neighbor dependence of stability for group iii rna single nucleotide bulge loops.thirty-five rna duplexes containing single nucleotide bulge loops were optically melted and the thermodynamic parameters for each duplex determined. the bulge loops were of the group iii variety, where the bulged nucleotide is either a ag/u or cu/g, leading to ambiguity to the exact position and identity of the bulge. all possible group iii bulge loops with watson-crick nearest-neighbors were examined. the data were used to develop a model to predict the free energy of an rna duplex containing a ...201424742935
amino acid-dependent stability of the acyl linkage in aminoacyl-trna.aminoacyl-trnas are the biologically active substrates for peptide bond formation in protein synthesis. the stability of the acyl linkage in each aminoacyl-trna, formed through an ester bond that connects the amino acid carboxyl group with the trna terminal 3'-oh group, is thus important. while the ester linkage is the same for all aminoacyl-trnas, the stability of each is not well characterized, thus limiting insight into the fundamental process of peptide bond formation. here, we show, by anal ...201424751649
structural and bioinformatic characterization of an acinetobacter baumannii type ii carrier protein.microorganisms produce a variety of natural products via secondary metabolic biosynthetic pathways. two of these types of synthetic systems, the nonribosomal peptide synthetases (nrpss) and polyketide synthases (pkss), use large modular enzymes containing multiple catalytic domains in a single protein. these multidomain enzymes use an integrated carrier protein domain to transport the growing, covalently bound natural product to the neighboring catalytic domains for each step in the synthesis. i ...201424914982
prokaryotic ubiquitin-like protein modification.prokaryotes form ubiquitin (ub)-like isopeptide bonds on the lysine residues of proteins by at least two distinct pathways that are reversible and regulated. in mycobacteria, the c-terminal gln of pup (prokaryotic ubiquitin-like protein) is deamidated and isopeptide linked to proteins by a mechanism distinct from ubiquitylation in enzymology yet analogous to ubiquitylation in targeting proteins for destruction by proteasomes. ub-fold proteins of archaea (samps, small archaeal modifier proteins) ...201424995873
loss of quaternary structure is associated with rapid sequence divergence in the osbs family.the rate of protein evolution is determined by a combination of selective pressure on protein function and biophysical constraints on protein folding and structure. determining the relative contributions of these properties is an unsolved problem in molecular evolution with broad implications for protein engineering and function prediction. as a case study, we examined the structural divergence of the rapidly evolving o-succinylbenzoate synthase (osbs) family, which catalyzes a step in menaquino ...201424872444
crystallization and preliminary x-ray crystallographic analysis of a bacterial asn-transamidosome.most canonical aminoacyl-trnas are synthesized directly by their cognate aminoacyl-trna synthetases (aarss), but glutaminyl-trna(gln) and asparaginyl-trna(asn) are synthesized indirectly by two-step processes. these processes are catalyzed by the transamidosome, a large ribonucleoprotein particle composed of gata, gatb, gatc, aars and trna. in this study, the asn-transamidosome from pseudomonas aeruginosa was reconstructed and crystallized by mixing purified gatcab complex, asprs and trna(asn). ...201424915095
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