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lack of discrimination against non-proteinogenic amino acid norvaline by elongation factor tu from escherichia coli.the gtp-bound form of elongation factor tu (ef-tu) brings aminoacylated trnas (aa-trna) to the a-site of the ribosome. ef-tu binds all cognate elongator aa-trnas with highly similar affinities, and its weaker or tighter binding of misacylated trnas may discourage their participation in translation. norvaline (nva) is a non-proteinogenic amino acid that is activated and transferred to trna(leu) by leucyl-trna synthetase (leurs). no notable accumulation of nva-trna(leu) has been observed in vitro, ...201323750044
rotary atpases: models, machine elements and technical specifications.rotary atpases are molecular rotary motors involved in biological energy conversion. they either synthesize or hydrolyze the universal biological energy carrier adenosine triphosphate. recent work has elucidated the general architecture and subunit compositions of all three sub-types of rotary atpases. composite models of the intact f-, v- and a-type atpases have been constructed by fitting high-resolution x-ray structures of individual subunits or sub-complexes into low-resolution electron dens ...201323369889
morphological development and cytochrome c oxidase activity in streptomyces lividans are dependent on the action of a copper bound sco protein.copper has an important role in the life cycle of many streptomycetes, stimulating the developmental switch between vegetative mycelium and aerial hyphae concomitant with the production of antibiotics. in streptomycetes, a gene encoding for a putative sco-like protein has been identified and is part of an operon that contains two other genes predicted to handle cellular copper. we report on the sco-like protein from streptomyces lividans (sco(sl)) and present a series of experiments that firmly ...201323345541
dna repair by reversal of dna damage.endogenous and exogenous factors constantly challenge cellular dna, generating cytotoxic and/or mutagenic dna adducts. as a result, organisms have evolved different mechanisms to defend against the deleterious effects of dna damage. among these diverse repair pathways, direct dna-repair systems provide cells with simple yet efficient solutions to reverse covalent dna adducts. in this review, we focus on recent advances in the field of direct dna repair, namely, photolyase-, alkyltransferase-, an ...201323284047
a structural basis for streptomycin-induced misreading of the genetic code.during protein synthesis, the ribosome selects aminoacyl-transfer rnas with anticodons matching the messenger rna codon present in the a site of the small ribosomal subunit. the aminoglycoside antibiotic streptomycin disrupts decoding by binding close to the site of codon recognition. here we use x-ray crystallography to define the impact of streptomycin on the decoding site of the thermus thermophilus 30s ribosomal subunit in complexes with cognate or near-cognate anticodon stem-loop analogues ...201323322043
a mutation of the rna polymerase β' subunit (rpoc) confers cephalosporin resistance in bacillus subtilis.in bacteria, mutations affecting the major catalytic subunits of rna polymerase (encoded by rpob and rpoc) emerge in response to a variety of selective pressures. here we isolated a bacillus subtilis strain with high-level resistance to cefuroxime (cef). whole-genome resequencing revealed only one missense mutation affecting an invariant residue in close proximity to the c-terminal dna-binding domain of rpoc (g1122d). genetic reconstruction experiments demonstrate that this substitution is suffi ...201323070162
characterization of recombinant fluoroquinolone-resistant pneumococcus-like isolates.fourteen fluoroquinolone-resistant streptococcal isolates with recombinant dna topoisomerase genes, preliminarily identified as pneumococci, were further characterized using phenotypic and genotypic approaches. phenotypic tests classified them as atypical pneumococci. phylogenetic relationships were analyzed by using the sequences of seven housekeeping alleles from these isolates and from isolates of streptococcus pneumoniae, streptococcus mitis, streptococcus oralis, and streptococcus pseudopne ...201323114769
thermodynamic properties distinguish human mitochondrial aspartyl-trna synthetase from bacterial homolog with same 3d architecture.in the mammalian mitochondrial translation apparatus, the proteins and their partner rnas are coded by two genomes. the proteins are nuclear-encoded and resemble their homologs, whereas the rnas coming from the rapidly evolving mitochondrial genome have lost critical structural information. this raises the question of molecular adaptation of these proteins to their peculiar partner rnas. the crystal structure of the homodimeric bacterial-type human mitochondrial aspartyl-trna synthetase (drs) co ...201223275545
thermodynamic properties distinguish human mitochondrial aspartyl-trna synthetase from bacterial homolog with same 3d architecture.in the mammalian mitochondrial translation apparatus, the proteins and their partner rnas are coded by two genomes. the proteins are nuclear-encoded and resemble their homologs, whereas the rnas coming from the rapidly evolving mitochondrial genome have lost critical structural information. this raises the question of molecular adaptation of these proteins to their peculiar partner rnas. the crystal structure of the homodimeric bacterial-type human mitochondrial aspartyl-trna synthetase (drs) co ...201223275545
crystallization and preliminary x-ray diffraction analysis of the nif3-family protein mj0927 from methanocaldococcus jannaschii.mj0927 is a member of the nif3 family and is widely distributed across living organisms. although several crystal structures of nif3 proteins have been reported, structural information on archaeal nif3 is still limited. to understand the structural differences between bacterial and archaeal nif3 proteins, mj0927 from methanocaldococcus jannaschii was purified and crystallized using the sitting-drop vapour-diffusion method. the crystals diffracted to a resolution of 2.47 å and belonged to the ort ...201223295494
crystallization and preliminary x-ray diffraction analysis of the nif3-family protein mj0927 from methanocaldococcus jannaschii.mj0927 is a member of the nif3 family and is widely distributed across living organisms. although several crystal structures of nif3 proteins have been reported, structural information on archaeal nif3 is still limited. to understand the structural differences between bacterial and archaeal nif3 proteins, mj0927 from methanocaldococcus jannaschii was purified and crystallized using the sitting-drop vapour-diffusion method. the crystals diffracted to a resolution of 2.47 å and belonged to the ort ...201223295494
effects of select histidine to cysteine mutations on transcriptional regulation by escherichia coli rcnr.the rcnr metalloregulator represses the transcription of the co(ii) and ni(ii) exporter, rcnab. previous studies have shown that co(ii) and ni(ii) bind to rcnr in six-coordinate sites, resulting in derepression. here, the roles of his60, his64, and his67 in specific metal recognition are examined. his60 and his64 correspond to ligands that are important for cu(i) binding in the homologous cu(i)-responsive metalloregulator, csor. these residues are known to be functionally important in rcnr trans ...201223215580
effects of select histidine to cysteine mutations on transcriptional regulation by escherichia coli rcnr.the rcnr metalloregulator represses the transcription of the co(ii) and ni(ii) exporter, rcnab. previous studies have shown that co(ii) and ni(ii) bind to rcnr in six-coordinate sites, resulting in derepression. here, the roles of his60, his64, and his67 in specific metal recognition are examined. his60 and his64 correspond to ligands that are important for cu(i) binding in the homologous cu(i)-responsive metalloregulator, csor. these residues are known to be functionally important in rcnr trans ...201223215580
crucial role of the c-terminal domain of mycobacterium tuberculosis leucyl-trna synthetase in aminoacylation and editing.the c-terminal extension of prokaryotic leucyl-trna synthetase (leurs) has been shown to make contacts with the tertiary structure base pairs of trna(leu) as well as its long variable arm. however, the precise role of the flexibly linked leurs c-terminal domain (ctd) in aminoacylation and editing processes has not been clarified. in this study, we carried out aspartic acid scanning within the ctd of mycobacterium tuberculosis leurs (mtbleurs) and studied the effects on trna(leu)-binding capacity ...201223268443
crucial role of the c-terminal domain of mycobacterium tuberculosis leucyl-trna synthetase in aminoacylation and editing.the c-terminal extension of prokaryotic leucyl-trna synthetase (leurs) has been shown to make contacts with the tertiary structure base pairs of trna(leu) as well as its long variable arm. however, the precise role of the flexibly linked leurs c-terminal domain (ctd) in aminoacylation and editing processes has not been clarified. in this study, we carried out aspartic acid scanning within the ctd of mycobacterium tuberculosis leurs (mtbleurs) and studied the effects on trna(leu)-binding capacity ...201223268443
genome-guided analysis of physiological and morphological traits of the fermentative acetate oxidizer thermacetogenium phaeum.thermacetogenium phaeum is a thermophilic strictly anaerobic bacterium oxidizing acetate to co(2) in syntrophic association with a methanogenic partner. it can also grow in pure culture, e.g., by fermentation of methanol to acetate. the key enzymes of homoacetate fermentation (wood-ljungdahl pathway) are used both in acetate oxidation and acetate formation. the obvious reversibility of this pathway in this organism is of specific interest since syntrophic acetate oxidation operates close to the ...201223259483
post-translation modification in archaea: lessons from haloferax volcanii and other haloarchaea.as an ever-growing number of genome sequences appear, it is becoming increasingly clear that factors other than genome sequence impart complexity to the proteome. of the various sources of proteomic variability, post-translational modifications (ptms) most greatly serve to expand the variety of proteins found in the cell. likewise, modulating the rates at which different proteins are degraded also results in a constantly changing cellular protein profile. while both strategies for generating pro ...201223167813
post-translation modification in archaea: lessons from haloferax volcanii and other haloarchaea.as an ever-growing number of genome sequences appear, it is becoming increasingly clear that factors other than genome sequence impart complexity to the proteome. of the various sources of proteomic variability, post-translational modifications (ptms) most greatly serve to expand the variety of proteins found in the cell. likewise, modulating the rates at which different proteins are degraded also results in a constantly changing cellular protein profile. while both strategies for generating pro ...201223167813
a trna-independent mechanism for transamidosome assembly promotes aminoacyl-trna transamidation.many bacteria lack genes encoding asparaginyl- and/or glutaminyl-trna synthetase and consequently rely on an indirect path for the synthesis of both asn-trna(asn) and gln-trna(gln). in some bacteria such as thermus thermophilus, efficient delivery of misacylated trna to the downstream amidotransferase (adt) is ensured by formation of a stable, trna-dependent macromolecular complex called the asn-transamidosome. this complex enables direct delivery of asp-trna(asn) from the non-discriminating asp ...201223258533
a trna-independent mechanism for transamidosome assembly promotes aminoacyl-trna transamidation.many bacteria lack genes encoding asparaginyl- and/or glutaminyl-trna synthetase and consequently rely on an indirect path for the synthesis of both asn-trna(asn) and gln-trna(gln). in some bacteria such as thermus thermophilus, efficient delivery of misacylated trna to the downstream amidotransferase (adt) is ensured by formation of a stable, trna-dependent macromolecular complex called the asn-transamidosome. this complex enables direct delivery of asp-trna(asn) from the non-discriminating asp ...201223258533
structural and functional insights into (s)-ureidoglycolate dehydrogenase, a metabolic branch point enzyme in nitrogen utilization.nitrogen metabolism is one of essential processes in living organisms. the catabolic pathways of nitrogenous compounds play a pivotal role in the storage and recovery of nitrogen. in escherichia coli, two different, interconnecting metabolic routes drive nitrogen utilization through purine degradation metabolites. the enzyme (s)-ureidoglycolate dehydrogenase (alld), which is a member of l-sulfolactate dehydrogenase-like family, converts (s)-ureidoglycolate, a key intermediate in the purine degra ...201223284870
structural analysis of protein-protein interactions in type i polyketide synthases.polyketide synthases (pkss) are responsible for synthesizing a myriad of natural products with agricultural, medicinal relevance. the pkss consist of multiple functional domains of which each can catalyze a specified chemical reaction leading to the synthesis of polyketides. biochemical studies showed that protein-substrate and protein-protein interactions play crucial roles in these complex regio-/stereo-selective biochemical processes. recent developments on x-ray crystallography and protein n ...201223249187
structural analysis of protein-protein interactions in type i polyketide synthases.polyketide synthases (pkss) are responsible for synthesizing a myriad of natural products with agricultural, medicinal relevance. the pkss consist of multiple functional domains of which each can catalyze a specified chemical reaction leading to the synthesis of polyketides. biochemical studies showed that protein-substrate and protein-protein interactions play crucial roles in these complex regio-/stereo-selective biochemical processes. recent developments on x-ray crystallography and protein n ...201223249187
anticodon g recognition by trna synthetases mimics the trna core.ancient mechanisms for nucleotide base recognition in the rna world are candidates for mimicking by early proteins like trna synthetases. in the core of the trna, conserved g22 interacts with two internal bases in a complex further stabilized by stacking interactions. this particular trna format for g recognition is shown here to be adapted by nine different and even nonhomologous anticodon binding domains (abds) of trna synthetases, in which amino acid side chains mimic all of the trna g22 base ...201223266103
anticodon g recognition by trna synthetases mimics the trna core.ancient mechanisms for nucleotide base recognition in the rna world are candidates for mimicking by early proteins like trna synthetases. in the core of the trna, conserved g22 interacts with two internal bases in a complex further stabilized by stacking interactions. this particular trna format for g recognition is shown here to be adapted by nine different and even nonhomologous anticodon binding domains (abds) of trna synthetases, in which amino acid side chains mimic all of the trna g22 base ...201223266103
role of nonspecific interactions in molecular chaperones through model-based bioinformatics.molecular chaperones are large proteins or protein complexes from which many proteins require assistance in order to fold. one unique property of molecular chaperones is the cavity they provide in which proteins fold. the interior surface residues which make up the cavities of molecular chaperone complexes from different organisms has recently been identified, including the well-studied groel-groes chaperonin complex found in escherichia coli. it was found that the interior of these protein comp ...201223260050
hiv-1 frameshift efficiency is primarily determined by the stability of base pairs positioned at the mrna entrance channel of the ribosome.the human immunodeficiency virus (hiv) requires a programmed -1 ribosomal frameshift for pol gene expression. the hiv frameshift site consists of a heptanucleotide slippery sequence (uuuuuua) followed by a spacer region and a downstream rna stem-loop structure. here we investigate the role of the rna structure in promoting the -1 frameshift. the stem-loop was systematically altered to decouple the contributions of local and overall thermodynamic stability towards frameshift efficiency. no correl ...201223248007
hiv-1 frameshift efficiency is primarily determined by the stability of base pairs positioned at the mrna entrance channel of the ribosome.the human immunodeficiency virus (hiv) requires a programmed -1 ribosomal frameshift for pol gene expression. the hiv frameshift site consists of a heptanucleotide slippery sequence (uuuuuua) followed by a spacer region and a downstream rna stem-loop structure. here we investigate the role of the rna structure in promoting the -1 frameshift. the stem-loop was systematically altered to decouple the contributions of local and overall thermodynamic stability towards frameshift efficiency. no correl ...201223248007
sediment microbial communities in great boiling spring are controlled by temperature and distinct from water communities.great boiling spring is a large, circumneutral, geothermal spring in the us great basin. twelve samples were collected from water and four different sediment sites on four different dates. microbial community composition and diversity were assessed by pcr amplification of a portion of the small subunit rrna gene using a universal primer set followed by pyrosequencing of the v8 region. analysis of 164 178 quality-filtered pyrotags clearly distinguished sediment and water microbial communities. wa ...201223235293
sediment microbial communities in great boiling spring are controlled by temperature and distinct from water communities.great boiling spring is a large, circumneutral, geothermal spring in the us great basin. twelve samples were collected from water and four different sediment sites on four different dates. microbial community composition and diversity were assessed by pcr amplification of a portion of the small subunit rrna gene using a universal primer set followed by pyrosequencing of the v8 region. analysis of 164 178 quality-filtered pyrotags clearly distinguished sediment and water microbial communities. wa ...201223235293
disruption of ionic interactions between the nucleotide binding domain 1 (nbd1) and middle (m) domain in hsp100 disaggregase unleashes toxic hyperactivity and partial independence from hsp70.hsp100 chaperones cooperate with the hsp70 chaperone system to disaggregate and reactivate heat-denatured aggregated proteins to promote cell survival after heat stress. the homology models of hsp100 disaggregases suggest the presence of a conserved network of ionic interactions between the first nucleotide binding domain (nbd1) and the coiled-coil middle subdomain, the signature domain of disaggregating chaperones. mutations intended to disrupt the putative ionic interactions in yeast hsp104 an ...201223233670
disruption of ionic interactions between the nucleotide binding domain 1 (nbd1) and middle (m) domain in hsp100 disaggregase unleashes toxic hyperactivity and partial independence from hsp70.hsp100 chaperones cooperate with the hsp70 chaperone system to disaggregate and reactivate heat-denatured aggregated proteins to promote cell survival after heat stress. the homology models of hsp100 disaggregases suggest the presence of a conserved network of ionic interactions between the first nucleotide binding domain (nbd1) and the coiled-coil middle subdomain, the signature domain of disaggregating chaperones. mutations intended to disrupt the putative ionic interactions in yeast hsp104 an ...201223233670
uv damage endonuclease employs a novel dual-dinucleotide flipping mechanism to recognize different dna lesions.repairing damaged dna is essential for an organism's survival. uv damage endonuclease (uvde) is a dna-repair enzyme that can recognize and incise different types of damaged dna. we present the structure of sulfolobus acidocaldarius uvde on its own and in a pre-catalytic complex with uv-damaged dna containing a 6-4 photoproduct showing a novel 'dual dinucleotide flip' mechanism for recognition of damaged dipyrimidines: the two purines opposite to the damaged pyrimidine bases are flipped into a di ...201223221644
uv damage endonuclease employs a novel dual-dinucleotide flipping mechanism to recognize different dna lesions.repairing damaged dna is essential for an organism's survival. uv damage endonuclease (uvde) is a dna-repair enzyme that can recognize and incise different types of damaged dna. we present the structure of sulfolobus acidocaldarius uvde on its own and in a pre-catalytic complex with uv-damaged dna containing a 6-4 photoproduct showing a novel 'dual dinucleotide flip' mechanism for recognition of damaged dipyrimidines: the two purines opposite to the damaged pyrimidine bases are flipped into a di ...201223221644
ebec 2012--an energetic time in freiburg.the seventeenth european bioenergetics conference (ebec) took place in september 2012 at the albert-ludwigs-university in freiburg, germany, and was hosted by thorsten friedrich. the conference is a biannual event that brings together researchers from across the globe to discuss progress in this diverse and challenging field.201223229583
ebec 2012--an energetic time in freiburg.the seventeenth european bioenergetics conference (ebec) took place in september 2012 at the albert-ludwigs-university in freiburg, germany, and was hosted by thorsten friedrich. the conference is a biannual event that brings together researchers from across the globe to discuss progress in this diverse and challenging field.201223229583
a gating motif in the translocation channel sets the hydrophobicity threshold for signal sequence function.a critical event in protein translocation across the endoplasmic reticulum is the structural transition between the closed and open conformations of sec61, the eukaryotic translocation channel. channel opening allows signal sequence insertion into a gap between the n- and c-terminal halves of sec61. we have identified a gating motif that regulates the transition between the closed and open channel conformations. polar amino acid substitutions in the gating motif cause a gain-of-function phenotyp ...201223229898
molecular mechanisms of survival strategies in extreme conditions.today, one of the major challenges in biophysics is to disclose the molecular mechanisms underlying biological processes. in such a frame, the understanding of the survival strategies in extreme conditions received a lot of attention both from the scientific and applicative points of view. since nature provides precious suggestions to be applied for improving the quality of life, extremophiles are considered as useful model-systems. the main goal of this review is to present an overview of some ...201225371270
space-related pharma-motifs for fast search of protein binding motifs and polypharmacological targets.to discover a compound inhibiting multiple proteins (i.e. polypharmacological targets) is a new paradigm for the complex diseases (e.g. cancers and diabetes). in general, the polypharmacological proteins often share similar local binding environments and motifs. as the exponential growth of the number of protein structures, to find the similar structural binding motifs (pharma-motifs) is an emergency task for drug discovery (e.g. side effects and new uses for old drugs) and protein functions.201223281852
functional relevance of dynamic properties of dimeric nadp-dependent isocitrate dehydrogenases.isocitrate dehydrogenases (idhs) are important enzymes present in all living cells. three subfamilies of functionally dimeric idhs (subfamilies i, ii, iii) are known. subfamily i are well-studied bacterial idhs, like that of escherischia coli. subfamily ii has predominantly eukaryotic members, but it also has several bacterial members, many being pathogens or endosymbionts. subfamily iii idhs are nad-dependent. the eukaryotic-like subfamily ii idh from pathogenic bacteria such as mycobacterium t ...201223281650
give it ago: the search for mirna-argonaute sorting signals in arabidopsis thaliana indicates a relevance of sequence positions other than the 5'-position alone.the specific recognition of mirnas by argonaute (ago) proteins, the effector proteins of the rna-induced silencing complex, constitutes the final step of the biogenesis of mirnas and is crucial for their target interaction. in the genome of arabidopsis thaliana (ath), 10 different ago proteins are encoded and the sorting decision, which mirna associates with which ago protein, was reported to depend exclusively on the identity of the 5'-sequence position of mature mirnas. hence, with only four d ...201223233858
promiscuous behaviour of archaeal ribosomal proteins: implications for eukaryotic ribosome evolution.in all living cells, protein synthesis occurs on ribonucleoprotein particles called ribosomes. molecular models have been reported for complete bacterial 70s and eukaryotic 80s ribosomes; however, only molecular models of large 50s subunits have been reported for archaea. here, we present a complete molecular model for the pyrococcus furiosus 70s ribosome based on a 6.6 å cryo-electron microscopy map. moreover, we have determined cryo-electron microscopy reconstructions of the euryarchaeota meth ...201223222135
promiscuous behaviour of archaeal ribosomal proteins: implications for eukaryotic ribosome evolution.in all living cells, protein synthesis occurs on ribonucleoprotein particles called ribosomes. molecular models have been reported for complete bacterial 70s and eukaryotic 80s ribosomes; however, only molecular models of large 50s subunits have been reported for archaea. here, we present a complete molecular model for the pyrococcus furiosus 70s ribosome based on a 6.6 å cryo-electron microscopy map. moreover, we have determined cryo-electron microscopy reconstructions of the euryarchaeota meth ...201223222135
selection of trna charging quality control mechanisms that increase mistranslation of the genetic code.mistranslation can follow two events during protein synthesis: production of non-cognate amino acid:transfer rna (trna) pairs by aminoacyl-trna synthetases (aarss) and inaccurate selection of aminoacyl-trnas by the ribosome. many aarss actively edit non-cognate amino acids, but editing mechanisms are not evolutionarily conserved, and their physiological significance remains unclear. to address the connection between aarss and mistranslation, the evolutionary divergence of tyrosine editing by phe ...201223222133
selection of trna charging quality control mechanisms that increase mistranslation of the genetic code.mistranslation can follow two events during protein synthesis: production of non-cognate amino acid:transfer rna (trna) pairs by aminoacyl-trna synthetases (aarss) and inaccurate selection of aminoacyl-trnas by the ribosome. many aarss actively edit non-cognate amino acids, but editing mechanisms are not evolutionarily conserved, and their physiological significance remains unclear. to address the connection between aarss and mistranslation, the evolutionary divergence of tyrosine editing by phe ...201223222133
archaeal β-casp ribonucleases of the acpsf1 family are orthologs of the eukaryal cpsf-73 factor.bacterial rnase j and eukaryal cleavage and polyadenylation specificity factor (cpsf-73) are members of the β-casp family of ribonucleases involved in mrna processing and degradation. here we report an in-depth phylogenomic analysis that delineates arnase j and archaeal cpsf (acpsf) as distinct orthologous groups and establishes their repartition in 110 archaeal genomes. the acpsf1 subgroup, which has been inherited vertically and is strictly conserved, is characterized by an n-terminal extensio ...201223222134
archaeal β-casp ribonucleases of the acpsf1 family are orthologs of the eukaryal cpsf-73 factor.bacterial rnase j and eukaryal cleavage and polyadenylation specificity factor (cpsf-73) are members of the β-casp family of ribonucleases involved in mrna processing and degradation. here we report an in-depth phylogenomic analysis that delineates arnase j and archaeal cpsf (acpsf) as distinct orthologous groups and establishes their repartition in 110 archaeal genomes. the acpsf1 subgroup, which has been inherited vertically and is strictly conserved, is characterized by an n-terminal extensio ...201223222134
isolation and characterization of rna polymerase rpob mutations that alter transcription slippage during elongation in escherichia coli.transcription fidelity is critical for maintaining the accurate flow of genetic information. the study of transcription fidelity has been limited because the intrinsic error rate of transcription is obscured by the higher error rate of translation, making identification of phenotypes associated with transcription infidelity challenging. slippage of elongating rna polymerase (rnap) on homopolymeric a/t tracts in dna represents a special type of transcription error leading to disruption of open re ...201223223236
isolation and characterization of rna polymerase rpob mutations that alter transcription slippage during elongation in escherichia coli.transcription fidelity is critical for maintaining the accurate flow of genetic information. the study of transcription fidelity has been limited because the intrinsic error rate of transcription is obscured by the higher error rate of translation, making identification of phenotypes associated with transcription infidelity challenging. slippage of elongating rna polymerase (rnap) on homopolymeric a/t tracts in dna represents a special type of transcription error leading to disruption of open re ...201223223236
crystal structures and kinetics of monofunctional proline dehydrogenase provide insight into substrate recognition and conformational changes associated with flavin reduction and product release.proline dehydrogenase (prodh) catalyzes the fad-dependent oxidation of proline to δ(1)-pyrroline-5-carboxylate, which is the first step of proline catabolism. here, we report the structures of proline dehydrogenase from deinococcus radiodurans in the oxidized state complexed with the proline analogue l-tetrahydrofuroic acid and in the reduced state with the proline site vacant. the analogue binds against the si face of the fad isoalloxazine and is protected from bulk solvent by helix α8 and the ...201223151026
in vivo protein interactions and complex formation in the pectobacterium atrosepticum subtype i-f crispr/cas system.clustered regularly interspaced short palindromic repeats (crispr) and their associated proteins (cas; crispr associated) are a bacterial defense mechanism against extra-chromosomal elements. crispr/cas systems are distinct from other known defense mechanisms insofar as they provide acquired and heritable immunity. resistance is accomplished in multiple stages in which the cas proteins provide the enzymatic machinery. importantly, subtype-specific proteins have been shown to form complexes in co ...201223226499
functional diversification of rok-family transcriptional regulators of sugar catabolism in the thermotogae phylum.large and functionally heterogeneous families of transcription factors have complex evolutionary histories. what shapes specificities toward effectors and dna sites in paralogous regulators is a fundamental question in biology. bacteria from the deep-branching lineage thermotogae possess multiple paralogs of the repressor, open reading frame, kinase (rok) family regulators that are characterized by carbohydrate-sensing domains shared with sugar kinases. we applied an integrated genomic approach ...201223209028
functional diversification of rok-family transcriptional regulators of sugar catabolism in the thermotogae phylum.large and functionally heterogeneous families of transcription factors have complex evolutionary histories. what shapes specificities toward effectors and dna sites in paralogous regulators is a fundamental question in biology. bacteria from the deep-branching lineage thermotogae possess multiple paralogs of the repressor, open reading frame, kinase (rok) family regulators that are characterized by carbohydrate-sensing domains shared with sugar kinases. we applied an integrated genomic approach ...201223209028
microbe-host interactions: structure and role of gram-negative bacterial porins.gram negative bacteria have evolved many mechanisms of attaching to and invading host epithelial and immune cells. in particular, many outer membrane proteins (omps) are involved in this initial interaction between the pathogen and their host. the outer membrane (om) of gram-negative bacteria performs the crucial role of providing an extra layer of protection to the organism without compromising the exchange of material required for sustaining life. the om, therefore, represents a sophisticated ...201223305369
direct interactions between the coiled-coil tip of dksa and the trigger loop of rna polymerase mediate transcriptional regulation.escherichia coli dksa is a transcription factor that binds to rna polymerase (rnap) without binding to dna, destabilizing rnap-promoter interactions, sensitizing rnap to the global regulator ppgpp, and regulating transcription of several hundred target genes, including those encoding rrna. previously, we described promoter sequences and kinetic properties that account for dksa's promoter specificity, but how dksa exerts its effects on rnap has remained unclear. to better understand dksa's mechan ...201223207918
cellular and molecular mechanisms of mitochondrial function.mitochondria are membrane bound organelles present in almost all eukaryotic cells. responsible for orchestrating cellular energy production, they are central to the maintenance of life and the gatekeepers of cell death. thought to have originated from symbiotic ancestors, they carry a residual genome as mtdna encoding 13 proteins essential for respiratory chain function. mitochondria comprise an inner and outer membrane that separate and maintain the aqueous regions, the intermembrane space and ...201223168274
identification and characterization of a highly conserved crenarchaeal protein lysine methyltransferase with broad substrate specificity.protein lysine methylation occurs extensively in the crenarchaeota, a major kingdom in the archaea. however, the enzymes responsible for this type of posttranslational modification have not been found. here we report the identification and characterization of the first crenarchaeal protein lysine methyltransferase, designated akmt, from the hyperthermophilic crenarchaeon sulfolobus islandicus. the enzyme was capable of transferring methyl groups to selected lysine residues in a substrate protein ...201223086207
twist-joints and double twist-joints in rna structure.analysis of available rna crystal structures has allowed us to identify a new family of rna arrangements that we call double twist-joints, or dtjs. each dtj is composed of a double helix that contains two bulges incorporated into different strands and separated from each other by 2 or 3 bp. at each bulge, the double helix is over-twisted, while the unpaired nucleotides of both bulges form a complex network of stacking and hydrogen-bonding with nucleotides of helical regions. in total, we identif ...201223060425
trmt61b is a methyltransferase responsible for 1-methyladenosine at position 58 of human mitochondrial trnas.in human mitochondria, 1-methyladenosine (m¹a) occurs at position 58 of trna(leu(uur)). in addition, partial m¹a58 modifications have been found in human mitochondrial trna(lys) and trna(ser(ucn)). we identified human trmt61b, which encodes a mitochondria-specific trna methyltransferase responsible for m¹a58 in these three trnas. trmt61b is dominantly localized to the mitochondria. m¹a58 formation in human mitochondrial trna(leu(uur)) could be reconstituted in vitro using recombinant trmt61b in ...201223097428
the rna polymerase bridge helix yfi motif in catalysis, fidelity and translocation.the bridge α-helix in the β' subunit of rna polymerase (rnap) borders the active site and may have roles in catalysis and translocation. in escherichia coli rnap, a bulky hydrophobic segment near the n-terminal end of the bridge helix is identified (β' 772-yfi-774; the yfi motif). yfi is located at a distance from the active center and adjacent to a glycine hinge (β' 778-garkg-782) involved in dynamic bending of the bridge helix. remarkably, amino acid substitutions in yfi significantly alter in ...201223202476
the rna polymerase bridge helix yfi motif in catalysis, fidelity and translocation.the bridge α-helix in the β' subunit of rna polymerase (rnap) borders the active site and may have roles in catalysis and translocation. in escherichia coli rnap, a bulky hydrophobic segment near the n-terminal end of the bridge helix is identified (β' 772-yfi-774; the yfi motif). yfi is located at a distance from the active center and adjacent to a glycine hinge (β' 778-garkg-782) involved in dynamic bending of the bridge helix. remarkably, amino acid substitutions in yfi significantly alter in ...201223202476
application of molecular diagnostic techniques for viral testing.nucleic acid amplification techniques are commonly used currently to diagnose viral diseases and manage patients with this kind of illnesses. these techniques have had a rapid but unconventional route of development during the last 30 years, with the discovery and introduction of several assays in clinical diagnosis. the increase in the number of commercially available methods has facilitated the use of this technology in the majority of laboratories worldwide. this technology has reduced the us ...201223248732
malolactic enzyme from oenococcus oeni: heterologous expression in escherichia coli and biochemical characterization.malolactic enzymes (mle) are known to directly convert l-malic acid into l-lactic acid with a catalytical requirement of nicotinamide adenine dinucleotide (nad (+) ) and mn ( 2+) ; however, the reaction mechanism is still unclear. to study a mle, the structural gene from oenococcus oeni strain dsm 20255 was heterologously expressed in escherichia coli, yielding 22.9 ku l (-1) fermentation broth. after affinity chromatography and removal of apparently inactive protein by precipitation, purified r ...201223196745
malolactic enzyme from oenococcus oeni: heterologous expression in escherichia coli and biochemical characterization.malolactic enzymes (mle) are known to directly convert l-malic acid into l-lactic acid with a catalytical requirement of nicotinamide adenine dinucleotide (nad (+) ) and mn ( 2+) ; however, the reaction mechanism is still unclear. to study a mle, the structural gene from oenococcus oeni strain dsm 20255 was heterologously expressed in escherichia coli, yielding 22.9 ku l (-1) fermentation broth. after affinity chromatography and removal of apparently inactive protein by precipitation, purified r ...201223196745
catalysis of dioxygen reduction by thermus thermophilus strain hb27 laccase on ketjen black electrodes.we present electrochemical analyses of the catalysis of dioxygen reduction by thermus thermophilus strain hb27 laccase on ketjen black substrates. our cathodes reliably produce 0.56 ma cm(-2) at 0.0 v vs ag|agcl reference at 30 °c in air-saturated buffer, under conditions of nonlimiting o(2) flux. we report the electrochemical activity of this laccase as a function of temperature, ph, time, and the efficiency of its conversion of dioxygen to water. we have measured the surface concentration of e ...201223163614
catalysis of dioxygen reduction by thermus thermophilus strain hb27 laccase on ketjen black electrodes.we present electrochemical analyses of the catalysis of dioxygen reduction by thermus thermophilus strain hb27 laccase on ketjen black substrates. our cathodes reliably produce 0.56 ma cm(-2) at 0.0 v vs ag|agcl reference at 30 °c in air-saturated buffer, under conditions of nonlimiting o(2) flux. we report the electrochemical activity of this laccase as a function of temperature, ph, time, and the efficiency of its conversion of dioxygen to water. we have measured the surface concentration of e ...201223163614
plasmodium falciparum uvrd helicase translocates in 3' to 5' direction, colocalizes with mlh and modulates its activity through physical interaction.malaria is a global disease and a major health problem. the control of malaria is a daunting task due to the increasing drug resistance. therefore, there is an urgent need to identify and characterize novel parasite specific drug targets. in the present study we report the biochemical characterization of parasite specific uvrd helicase from plasmodium falciparum. the n-terminal fragment (pfudn) containing uvrd helicase domain, which consists of helicase motifs q, ia-id, ii, iii and most of motif ...201223185322
crystallization and preliminary x-ray analysis of the open form of human ecto-5'-nucleotidase (cd73).eukaryotic ecto-5'-nucleotidase (e5nt) catalyses the hydrolysis of extracellular amp to adenosine and plays a pivotal role in switching on adenosine signalling via the p1 receptors of the purinergic signalling pathway. with such an important regulatory role, e5nt has become an appealing new drug target, with potential applications in the treatment of inflammation, chronic pain, hypoxia and cancer. in order to gain insight into the structure and function of the eukaryotic e5nt enzymes and to assi ...201223192044
mechanical modulation of atp-binding affinity of v1-atpase.v(1)-atpase is a rotary motor protein that rotates the central shaft in a counterclockwise direction hydrolyzing atp. although the atp-binding process is suggested to be the most critical reaction step for torque generation in f(1)-atpase (the closest relative of v(1)-atpase evolutionarily), the role of atp binding for v(1)-atpase in torque generation has remained unclear. in the present study, we performed single-molecule manipulation experiments on v(1)-atpase from thermus thermophilus to inve ...201223155048
mechanical modulation of atp-binding affinity of v1-atpase.v(1)-atpase is a rotary motor protein that rotates the central shaft in a counterclockwise direction hydrolyzing atp. although the atp-binding process is suggested to be the most critical reaction step for torque generation in f(1)-atpase (the closest relative of v(1)-atpase evolutionarily), the role of atp binding for v(1)-atpase in torque generation has remained unclear. in the present study, we performed single-molecule manipulation experiments on v(1)-atpase from thermus thermophilus to inve ...201223155048
crystallization and preliminary x-ray crystallographic analysis of human apaf-1-interacting protein.apaf-1-interacting protein (apip) is known to inhibit two different types of cell death: caspase-1-dependent pyroptosis and caspase-9-dependent apoptosis. apip is also involved in the methionine-salvage pathway, where it is called 5-methylthioribulose-1-phosphate dehydratase (mtnb). the enzyme activity seems to be essential for inhibition of pyroptosis by apip, but not for inhibition of apoptosis. in this study, human apip was overproduced in escherichia coli, purified and crystallized. an x-ray ...201223192037
crystallization and preliminary x-ray crystallographic analysis of quinolinate phosphoribosyltransferase from porcine kidney in complex with nicotinate mononucleotide.quinolinate phosphoribosyltransferase (qaprtase) is a key enzyme in nad biosynthesis; it catalyzes the formation of nicotinate mononucleotide (namn) from quinolinate and 5-phosphoribosyl-1-pyrophosphate. in order to elucidate the mechanism of namn biosynthesis, crystals of sus scrofa qaprtase (ss-qaprtase) purified from porcine kidney in complex with namn were obtained and diffraction data were collected and processed to 2.1 å resolution. the ss-qaprtase-namn cocrystals belonged to space group p ...201223192029
inorganic pyrophosphatase crystals from thermococcus thioreducens for x-ray and neutron diffraction.inorganic pyrophosphatase (ippase) from the archaeon thermococcus thioreducens was cloned, overexpressed in escherichia coli, purified and crystallized in restricted geometry, resulting in large crystal volumes exceeding 5 mm3. ippase is thermally stable and is able to resist denaturation at temperatures above 348 k. owing to the high temperature tolerance of the enzyme, the protein was amenable to room-temperature manipulation at the level of protein preparation, crystallization and x-ray and n ...201223192028
structural variation and uniformity among tetraloop-receptor interactions and other loop-helix interactions in rna crystal structures.tetraloop-receptor interactions are prevalent structural units in rnas, and include the gaaa/11-nt and gnra-minor groove interactions. in this study, we have compiled a set of 78 nonredundant loop-helix interactions from x-ray crystal structures, and examined them for the extent of their sequence and structural variation. of the 78 interactions in the set, only four were classical gaaa/11-nt motifs, while over half (48) were gnra-minor groove interactions. the gnra-minor groove interactions were ...201223152878
unraveling the dynamics of ribosome translocation.translocation is one of the key events in translation, requiring large-scale conformational changes in the ribosome, movements of two transfer rnas (trnas) across a distance of more than 20å, and the coupled movement of the messenger rna (mrna) by one codon, completing one cycle of peptide-chain elongation. translocation is catalyzed by elongation factor g (ef-g in bacteria), which hydrolyzes gtp in the process. however, how the conformational rearrangements of the ribosome actually drive the mo ...201223142574
ubiquitin-like proteins and their roles in archaea.this review highlights the finding that ubiquitin-like (ubl) proteins of archaea (termed samps) function not only as sulfur carriers but also as protein modifiers. ubaa (an e1 ubiquitin-activating enzyme homolog of archaea) is required for the samps to be covalently attached to proteins. the samps and ubaa are also needed to form sulfur-containing biomolecules (e.g., thiolated trna and molybdenum cofactor). these findings provide a new perspective on how ubl proteins can serve as both sulfur car ...201223140889
ubiquitin-like proteins and their roles in archaea.this review highlights the finding that ubiquitin-like (ubl) proteins of archaea (termed samps) function not only as sulfur carriers but also as protein modifiers. ubaa (an e1 ubiquitin-activating enzyme homolog of archaea) is required for the samps to be covalently attached to proteins. the samps and ubaa are also needed to form sulfur-containing biomolecules (e.g., thiolated trna and molybdenum cofactor). these findings provide a new perspective on how ubl proteins can serve as both sulfur car ...201223140889
the fungal α-aminoadipate pathway for lysine biosynthesis requires two enzymes of the aconitase family for the isomerization of homocitrate to homoisocitrate.fungi produce α-aminoadipate, a precursor for penicillin and lysine via the α-aminoadipate pathway. despite the biotechnological importance of this pathway, the essential isomerization of homocitrate via homoaconitate to homoisocitrate has hardly been studied. therefore, we analysed the role of homoaconitases and aconitases in this isomerization. although we confirmed an essential contribution of homoaconitases from saccharomyces cerevisiae and aspergillus fumigatus, these enzymes only catalysed ...201223106124
role of the -pewy-glutamate in catalysis at the q(o)-site of the cyt bc(1) complex.we re-examine the ph dependence of partial processes of ubihydroquinone (qh(2)) turnover in glu-295 mutants in rhodobacter sphaeroides to clarify the mechanistic role. in more crippled mutants, the bell-shaped ph profile of wildtype was replaced by dependence on a single pk at ~8.5 favoring electron transfer. loss of the pk at 6.5 reflects a change in the rate-limiting step from the first to the second electron transfer. over the range of ph 6-8, no major ph dependence of formation of the initia ...201223123515
role of the -pewy-glutamate in catalysis at the q(o)-site of the cyt bc(1) complex.we re-examine the ph dependence of partial processes of ubihydroquinone (qh(2)) turnover in glu-295 mutants in rhodobacter sphaeroides to clarify the mechanistic role. in more crippled mutants, the bell-shaped ph profile of wildtype was replaced by dependence on a single pk at ~8.5 favoring electron transfer. loss of the pk at 6.5 reflects a change in the rate-limiting step from the first to the second electron transfer. over the range of ph 6-8, no major ph dependence of formation of the initia ...201223123515
inhibition of protein synthesis on the ribosome by tildipirosin compared with other veterinary macrolides.tildipirosin is a 16-membered-ring macrolide developed to treat bacterial pathogens, including mannheimia haemolytica and pasteurella multocida, that cause respiratory tract infections in cattle and swine. here we evaluated the efficacy of tildipirosin at inhibiting protein synthesis on the ribosome (50% inhibitory concentration [ic(50)], 0.23 ± 0.01 μm) and compared it with the established veterinary macrolides tylosin, tilmicosin, and tulathromycin. mutation and methylation at key rrna nucleot ...201222926570
inactivation of ribosomal protein genes in bacillus subtilis reveals importance of each ribosomal protein for cell proliferation and cell differentiation.among the 57 genes that encode ribosomal proteins in the genome of bacillus subtilis, a gram-positive bacterium, 50 genes were targeted by systematic inactivation. individual deletion mutants of 16 ribosomal proteins (l1, l9, l15, l22, l23, l28, l29, l32, l33.1, l33.2, l34, l35, l36, s6, s20, and s21) were obtained successfully. in conjunction with previous reports, 22 ribosomal proteins have been shown to be nonessential in b. subtilis, at least for cell proliferation. although several mutants ...201223002217
insights into glycogen metabolism in chemolithoautotrophic bacteria from distinctive kinetic and regulatory properties of adp-glucose pyrophosphorylase from nitrosomonas europaea.nitrosomonas europaea is a chemolithoautotroph that obtains energy by oxidizing ammonia in the presence of oxygen and fixes co(2) via the benson-calvin cycle. despite its environmental and evolutionary importance, very little is known about the regulation and metabolism of glycogen, a source of carbon and energy storage. here, we cloned and heterologously expressed the genes coding for two major putative enzymes of the glycogen synthetic pathway in n. europaea, adp-glucose pyrophosphorylase and ...201222961847
crystal structures of complexes of the branched-chain aminotransferase from deinococcus radiodurans with α-ketoisocaproate and l-glutamate suggest the radiation resistance of this enzyme for catalysis.branched-chain aminotransferases (bcat), which utilize pyridoxal 5'-phosphate (plp) as a cofactor, reversibly catalyze the transfer of the α-amino groups of three of the most hydrophobic branched-chain amino acids (bcaa), leucine, isoleucine, and valine, to α-ketoglutarate to form the respective branched-chain α-keto acids and glutamate. the bcat from deinococcus radiodurans (drbcat), an extremophile, was cloned and expressed in escherichia coli for structure and functional studies. the crystal ...201222984263
functional characterization of the role of the chromosome i partitioning system in genome segregation in deinococcus radiodurans.deinococcus radiodurans, a radiation-resistant bacterium, harbors a multipartite genome. chromosome i contains three putative centromeres (segs1, segs2, and segs3), and para (para1) and parb (parb1) homologues. the parb1 interaction with segs was sequence specific, and para1 was shown to be a dna binding atpase. the atpase activity of para1 was stimulated when segs elements were coincubated with parb1, but the greatest increase was observed with segs3. para1 incubated with the segs-parb1 complex ...201222843847
cas5d processes pre-crrna and is a member of a larger family of crispr rna endonucleases.small rnas derived from clustered, regularly interspaced, short palindromic repeat (crispr) loci in bacteria and archaea are involved in an adaptable and heritable gene-silencing pathway. resistance to invasive genetic material is conferred by the incorporation of short dna sequences derived from this material into the genome as crispr spacer elements separated by short repeat sequences. processing of long primary transcripts (pre-crrnas) containing these repeats by a crispr-associated (cas) rna ...201223006625
structural asymmetry in the thermus thermophilus ruvc dimer suggests a basis for sequential strand cleavages during holliday junction resolution.holliday junction (hj) resolvases are structure-specific endonucleases that cleave four-way dna junctions (hjs) generated during dna recombination and repair. bacterial ruvc, a prototypical hj resolvase, functions as homodimer and nicks dna strands precisely across the junction point. to gain insights into the mechanisms underlying symmetrical strand cleavages by ruvc, we performed crystallographic and biochemical analyses of ruvc from thermus thermophilus (t.th. ruvc). the crystal structure of ...201223118486
structural asymmetry in the thermus thermophilus ruvc dimer suggests a basis for sequential strand cleavages during holliday junction resolution.holliday junction (hj) resolvases are structure-specific endonucleases that cleave four-way dna junctions (hjs) generated during dna recombination and repair. bacterial ruvc, a prototypical hj resolvase, functions as homodimer and nicks dna strands precisely across the junction point. to gain insights into the mechanisms underlying symmetrical strand cleavages by ruvc, we performed crystallographic and biochemical analyses of ruvc from thermus thermophilus (t.th. ruvc). the crystal structure of ...201223118486
alkyltransferase-like protein (atl1) distinguishes alkylated guanines for dna repair using cation-π interactions.alkyltransferase-like (atl) proteins in schizosaccharomyces pombe (atl1) and thermus thermophilus (ttha1564) protect against the adverse effects of dna alkylation damage by flagging o(6)-alkylguanine lesions for nucleotide excision repair (ner). we show that both atl proteins bind with high affinity to oligodeoxyribonucleotides containing o(6)-alkylguanines differing in size, polarity, and charge of the alkyl group. however, atl1 shows a greater ability than ttha1564 to distinguish between o(6)- ...201223112169
directed evolution: selection of the host organism.directed evolution has become a well-established tool for improving proteins and biological systems. a critical aspect of directed evolution is the selection of a suitable host organism for achieving functional expression of the target gene. to date, most directed evolution studies have used either escherichia coli or saccharomyces cerevisiae as a host; however, other bacterial and yeast species, as well as mammalian and insect cell lines, have also been successfully used. recent advances in syn ...201224688653
proteomic properties reveal phyloecological clusters of archaea.in this study, we propose a novel way to describe the variety of environmental adaptations of archaea. we have clustered 57 archaea by using a non-redundant set of proteomic features, and verified that the clusters correspond to environmental adaptations to the archaeal habitats. the first cluster consists dominantly of hyperthermophiles and hyperthermoacidophilic aerobes. the second cluster joins together halophilic and extremely halophilic archaea, while the third cluster contains mesophilic ( ...201223133575
the trna recognition mechanism of folate/fad-dependent trna methyltransferase (trmfo).the conserved u54 in trna is often modified to 5-methyluridine (m(5)u) and forms a reverse hoogsteen base pair with a58 that stabilizes the l-shaped trna structure. in gram-positive and some gram-negative eubacteria, m(5)u54 is produced by folate/fad-dependent trna (m(5)u54) methyltransferase (trmfo). trmfo utilizes n(5),n(10)-methylenetetrahydrofolate (ch(2)thf) as a methyl donor. we previously reported an in vitro trmfo assay system, in which unstable [(14)c]ch(2)thf was supplied from [(14)c]s ...201223095745
structural basis of transcription initiation.during transcription initiation, rna polymerase (rnap) binds and unwinds promoter dna to form an rnap-promoter open complex. we have determined crystal structures at 2.9 and 3.0 å resolution of functional transcription initiation complexes comprising thermus thermophilus rna polymerase, σ(a), and a promoter dna fragment corresponding to the transcription bubble and downstream double-stranded dna of the rnap-promoter open complex. the structures show that σ recognizes the -10 element and discrimi ...201223086998
leishmania donovani argininosuccinate synthase is an active enzyme associated with parasite pathogenesis.gene expression analysis in leishmania donovani (ld) identified an orthologue of the urea cycle enzyme, argininosuccinate synthase (ldass), that was more abundantly expressed in amastigotes than in promastigotes. in order to characterize in detail this newly identified protein in leishmania, we determined its enzymatic activity, subcellular localization in the parasite and affect on virulence in vivo.201223094117
nucleic acid binding surface and dimer interface revealed by crispr-associated casb protein structures.the crispr system is an adaptive rna-based microbial immune system against invasive genetic elements. casb is an essential protein component in type i-e cascade. here, we characterize casb proteins from three different organisms as non-specific nucleic acid binding proteins. the thermobifida fusca casb crystal structure reveals conserved positive surface charges, which we show are important for its nucleic acid binding function. em docking reveals that casb dimerization aligns individual nucleic ...201223079036
recent advances in the structural mechanisms of dna glycosylases.dna glycosylases safeguard the genome by locating and excising a diverse array of aberrant nucleobases created from oxidation, alkylation, and deamination of dna. since the discovery 28years ago that these enzymes employ a base flipping mechanism to trap their substrates, six different protein architectures have been identified to perform the same basic task. work over the past several years has unraveled details for how the various dna glycosylases survey dna, detect damage within the duplex, s ...201223076011
recent advances in the structural mechanisms of dna glycosylases.dna glycosylases safeguard the genome by locating and excising a diverse array of aberrant nucleobases created from oxidation, alkylation, and deamination of dna. since the discovery 28years ago that these enzymes employ a base flipping mechanism to trap their substrates, six different protein architectures have been identified to perform the same basic task. work over the past several years has unraveled details for how the various dna glycosylases survey dna, detect damage within the duplex, s ...201223076011
alternative ground states enable pathway switching in biological electron transfer.electron transfer is the simplest chemical reaction and constitutes the basis of a large variety of biological processes, such as photosynthesis and cellular respiration. nature has evolved specific proteins and cofactors for these functions. the mechanisms optimizing biological electron transfer have been matter of intense debate, such as the role of the protein milieu between donor and acceptor sites. here we propose a mechanism regulating long-range electron transfer in proteins. specifically ...201223054836
archaeal jab1/mpn/mov34 metalloenzyme (hvjamm1) cleaves ubiquitin-like small archaeal modifier proteins (samps) from protein-conjugates.proteins with jab1/mpn/mov34 metalloenzyme (jamm/mpn+) domains are widespread among all domains of life, yet poorly understood. here we report the purification and characterization of an archaeal jamm/mpn+ domain protein (hvjamm1) from haloferax volcanii that cleaves ubiquitin-like small archaeal modifier proteins (samp1/2) from protein conjugates. hvjamm1 cleaved samp1/2 conjugates generated in h. volcanii as well as isopeptide- and linear-linked samp1-moae in purified form. cleavage of linear ...201222970855
proteins of unknown function in the protein data bank (pdb): an inventory of true uncharacterized proteins and computational tools for their analysis.proteins of uncharacterized functions form a large part of many of the currently available biological databases and this situation exists even in the protein data bank (pdb). our analysis of recent pdb data revealed that only 42.53% of pdb entries (1084 coordinate files) that were categorized under "unknown function" are true examples of proteins of unknown function at this point in time. the remainder 1465 entries also annotated as such appear to be able to have their annotations re-assessed, b ...201223202924
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