Publications

probing protein structure by amino acid-specific covalent labeling and mass spectrometry.	for many years, amino acid-specific covalent labeling has been a valuable tool to study protein structure and protein interactions, especially for systems that are difficult to study by other means. these covalent labeling methods typically map protein structure and interactions by measuring the differential reactivity of amino acid side chains. the reactivity of amino acids in proteins generally depends on the accessibility of the side chain to the reagent, the inherent reactivity of the label ...	2009	19016300
marine-derived metabolites of s-adenosylmethionine as templates for new anti-infectives.	s-adenosylmethionine (adomet) is a key biochemical co-factor whose proximate metabolites include methylated macromolecules (e.g., nucleic acids, proteins, phospholipids), methylated small molecules (e.g., sterols, biogenic amines), polyamines (e.g., spermidine, spermine), ethylene, and n-acyl-homoserine lactones. marine organisms produce numerous adomet metabolites whose novel structures can be regarded as lead compounds for anti-infective drug design.	2009	19841722
structural analysis of polarizing indels: an emerging consensus on the root of the tree of life.	the root of the tree of life has been a holy grail ever since darwin first used the tree as a metaphor for evolution. new methods seek to narrow down the location of the root by excluding it from branches of the tree of life. this is done by finding traits that must be derived, and excluding the root from the taxa those traits cover. however the two most comprehensive attempts at this strategy, performed by cavalier-smith and lake et al., have excluded each other's rootings.	2009	19706177
metal uptake by manganese superoxide dismutase.	manganese superoxide dismutase is an important antioxidant defense metalloenzyme that protects cells from damage by the toxic oxygen metabolite, superoxide free radical, formed as an unavoidable by-product of aerobic metabolism. many years of research have gone into understanding how the metal cofactor interacts with small molecules in its catalytic role. in contrast, very little is presently known about how the protein acquires its metal cofactor, an important step in the maturation of the prot ...	2009	19699328
metal uptake by manganese superoxide dismutase.	manganese superoxide dismutase is an important antioxidant defense metalloenzyme that protects cells from damage by the toxic oxygen metabolite, superoxide free radical, formed as an unavoidable by-product of aerobic metabolism. many years of research have gone into understanding how the metal cofactor interacts with small molecules in its catalytic role. in contrast, very little is presently known about how the protein acquires its metal cofactor, an important step in the maturation of the prot ...	2009	19699328
computational design of candida boidinii xylose reductase for altered cofactor specificity.	in this study we introduce a computationally-driven enzyme redesign workflow for altering cofactor specificity from nadph to nadh. by compiling and comparing data from previous studies involving cofactor switching mutations, we show that their effect cannot be explained as straightforward changes in volume, hydrophobicity, charge, or blosum62 scores of the residues populating the cofactor binding site. instead, we find that the use of a detailed cofactor binding energy approximation is needed to ...	2009	19693930
structure of era in complex with the 3' end of 16s rrna: implications for ribosome biogenesis.	era, composed of an n-terminal gtpase domain followed by an rna-binding kh domain, is essential for bacterial cell viability. it binds to 16s rrna and the 30s ribosomal subunit. however, its rna-binding site, the functional relationship between the two domains, and its role in ribosome biogenesis remain unclear. we have determined two crystal structures of era, a binary complex with gdp and a ternary complex with a gtp-analog and the 1531aucaccuccuua1542 sequence at the 3' end of 16s rrna. in th ...	2009	19706445
large facilities and the evolving ribosome, the cellular machine for genetic-code translation.	well-focused x-ray beams, generated by advanced synchrotron radiation facilities, yielded high-resolution diffraction data from crystals of ribosomes, the cellular nano-machines that translate the genetic code into proteins. these structures revealed the decoding mechanism, localized the mrna path and the positions of the trna molecules in the ribosome and illuminated the interactions of the ribosome with initiation, release and recycling factors. they also showed that the ribosome is a ribozyme ...	2009	19656820
studying the salt dependence of the binding of sigma70 and sigma32 to core rna polymerase using luminescence resonance energy transfer.	the study of protein-protein interactions is becoming increasingly important for understanding the regulation of many cellular processes. the ability to quantify the strength with which two binding partners interact is desirable but the accurate determination of equilibrium binding constants is a difficult process. the use of luminescence resonance energy transfer (lret) provides a homogeneous binding assay that can be used for the detection of protein-protein interactions. previously, we develo ...	2009	19649256
biogenesis and homeostasis of nicotinamide adenine dinucleotide cofactor.	universal and ubiquitous redox cofactors, nicotinamide adenine dinucleotide (nad) and its phosphorylated analog (nadp), collectively contribute to approximately 12% of all biochemical reactions included in the metabolic model of escherichia coli k-12. a homeostasis of the nad pool faithfully maintained by the cells results from a dynamic balance in a network of nad biosynthesis, utilization, decomposition, and recycling pathways that is subject to tight regulation at various levels. a brief over ...	2009	26443758
bis{μ-4,4'-dimeth-oxy-2,2'-[propane-1,2-diylbis(nitrilo-methyl-idyne)]diphenolato}bis-({4,4'-dimeth-oxy-2,2'-[propane-1,2-diylbis(nitrilo-methyl-idyne)]diphenol}manganese(iii)) bis-(hexa-fluorido-phosphate).	in the title complex, [mn(2)(c(19)h(20)n(2)o(4))(2)(c(19)h(22)n(2)o(4))(2)](pf(6))(2), the mn(iii) ion is coordinated by two o [mn-o = 1.855 (2) and 1.887 (2) å] and two n [mn-n = 1.982 (3) and 1.977 (3) å] atoms from the tetra-dentate schiff base ligand and a coordinated axial ligand [mn-o = 2.129 (2) å]. the centrosymmetric dimer contains two jahn-teller-distorted mn(iii) ions, each in a nearly octa-hedral geometry, connected through two phenolate bridges from two ligands. there are two stereo ...	2009	21583304
crystal structure of a homolog of mammalian serine racemase from schizosaccharomyces pombe.	d-serine is an endogenous coagonist for the n-methyl-d-aspartate receptor and is involved in excitatory neurotransmission in the brain. mammalian pyridoxal 5'-phosphate-dependent serine racemase, which is localized in the mammalian brain, catalyzes the racemization of l-serine to yield d-serine and vice versa. the enzyme also catalyzes the dehydration of d- and l-serine. both reactions are enhanced by mg.atp in vivo. we have determined the structures of the following three forms of the mammalian ...	2009	19640845
sequence-based analysis of protein energy landscapes reveals nonuniform thermal adaptation within the proteome.	thermal adaptation of individual proteins is often achieved through modulating protein stability, with proteins that are adapted to extreme cold environments having increased conformational flexibility when brought to mesophilic conditions. conversely, proteins adapted to higher temperatures appear less dynamic and are found to be much more stable against thermal denaturation than their mesophilic counterparts. according to the current paradigm, the adaptation of an organism for survival at high ...	2009	19592668
regulation and function of proline oxidase under nutrient stress.	under conditions of nutrient stress, cells switch to a survival mode catabolizing cellular and tissue constituents for energy. proline metabolism is especially important in nutrient stress because proline is readily available from the breakdown of extracellular matrix (ecm), and the degradation of proline through the proline cycle initiated by proline oxidase (pox), a mitochondrial inner membrane enzyme, can generate atp. this degradative pathway generates glutamate and alpha-ketoglutarate, prod ...	2009	19415679
the dimeric proto-ribosome: structural details and possible implications on the origin of life.	a symmetric pocket-like entity, composed of two l-shaped rna units, encircles the peptide synthesis site within the contemporary ribosome. this entity was suggested to be the vestige of a dimeric proto-ribosome, which could have formed spontaneously in the prebiotic world, catalyzing non-coded peptide bond formation and elongation. this structural element, beyond offering the initial step in the evolution of translation, is hypothesized here to be linked to the origin of life. by catalyzing the ...	2009	19742176
carboxylic ester hydrolases from hyperthermophiles.	carboxylic ester hydrolyzing enzymes constitute a large group of enzymes that are able to catalyze the hydrolysis, synthesis or transesterification of an ester bond. they can be found in all three domains of life, including the group of hyperthermophilic bacteria and archaea. esterases from the latter group often exhibit a high intrinsic stability, which makes them of interest them for various biotechnological applications. in this review, we aim to give an overview of all characterized carboxyl ...	2009	19544040
thermodynamic characterization of tandem mismatches found in naturally occurring rna.	although all sequence symmetric tandem mismatches and some sequence asymmetric tandem mismatches have been thermodynamically characterized and a model has been proposed to predict the stability of previously unmeasured sequence asymmetric tandem mismatches [christiansen,m.e. and znosko,b.m. (2008) biochemistry, 47, 4329-4336], experimental thermodynamic data for frequently occurring tandem mismatches is lacking. since experimental data is preferred over a predictive model, the thermodynamic para ...	2009	19509311
limited proteolysis analysis of the ribosome is affected by subunit association.	our understanding of the structural organization of ribosome assembly intermediates, in particular those intermediates that result from misfolding leading to their eventual degradation within the cell, is limited because of the lack of methods available to characterize assembly intermediate structures. because conventional structural approaches, such as nmr, x-ray crystallography, and cryo-em, are not ideally suited to characterize the structural organization of these flexible and sometimes hete ...	2009	19213046
collective dynamics of the ribosomal tunnel revealed by elastic network modeling.	the collective dynamics of the nascent polypeptide exit tunnel are investigated with the computationally efficient elastic network model using normal mode analysis. the calculated normal modes are considered individually and in linear combinations with different coefficients mimicking the phase angles between modes, in order to follow the mechanistic motions of tunnel wall residues. the low frequency fluctuations indicate three distinct regions along the tunnel-the entrance, the neck, and the ex ...	2009	19004020
evidence for phosphatase activity of p27sj and its impact on the cell cycle.	p27sj, a novel protein isolated from st john's wort (hypericum perforatum), belongs to an emerging family of ding proteins that are related to a prokaryotic phosphate-binding protein superfamily. here we demonstrate that p27sj exhibits phosphatase activity and that its expression in cells decreases the level of phosphorylated erk1/2, a key protein of several signaling pathways. treatment of p27sj-expressing cells with phosphatase inhibitors including okadaic acid, maintained erk1/2 in its phosph ...	2009	19343785
a ca(+) pair adjacent to a sheared ga or aa pair stabilizes size-symmetric rna internal loops.	rna internal loops are often important sites for folding and function. residues in internal loops can have pka values shifted close to neutral ph because of the local structural environment. a series of rna internal loops were studied at different ph by uv absorbance versus temperature melting experiments and imino proton nuclear magnetic resonance (nmr). a stabilizing ca pair forms at ph 7 in the cg/aa and ca/aa nearest neighbors when the ca pair is the first noncanonical pair (loop-terminal pa ...	2009	19485416
nucleic acids: how high resolution structural biology help us to understand darwinian evolution.		2009	19482468
conformational changes in redox pairs of protein structures.	disulfides are conventionally viewed as structurally stabilizing elements in proteins but emerging evidence suggests two disulfide subproteomes exist. one group mediates the well known role of structural stabilization. a second redox-active group are best known for their catalytic functions but are increasingly being recognized for their roles in regulation of protein function. redox-active disulfides are, by their very nature, more susceptible to reduction than structural disulfides; and conver ...	2009	19598234
prediction of the rotational tumbling time for proteins with disordered segments.	for well-structured, rigid proteins, the prediction of rotational tumbling time (tau(c)) using atomic coordinates is reasonably accurate, but is inaccurate for proteins with long unstructured sequences. under physiological conditions, many proteins contain long disordered segments that play important regulatory roles in fundamental biological events including signal transduction and molecular recognition. here we describe an ensemble approach to the boundary element method that accurately predic ...	2009	19391622
carboxylation mechanism and stereochemistry of crotonyl-coa carboxylase/reductase, a carboxylating enoyl-thioester reductase.	chemo- and stereoselective reductions are important reactions in chemistry and biology, and reductases from biological sources are increasingly applied in organic synthesis. in contrast, carboxylases are used only sporadically. we recently described crotonyl-coa carboxylase/reductase, which catalyzes the reduction of (e)-crotonyl-coa to butyryl-coa but also the reductive carboxylation of (e)-crotonyl-coa to ethylmalonyl-coa. in this study, the complete stereochemical course of both reactions was ...	2009	19458256
mm-align: a quick algorithm for aligning multiple-chain protein complex structures using iterative dynamic programming.	structural comparison of multiple-chain protein complexes is essential in many studies of protein-protein interactions. we develop a new algorithm, mm-align, for sequence-independent alignment of protein complex structures. the algorithm is built on a heuristic iteration of a modified needleman-wunsch dynamic programming (dp) algorithm, with the alignment score specified by the inter-complex residue distances. the multiple chains in each complex are first joined, in every possible order, and the ...	2009	19443443
crystal structure of iodotyrosine deiodinase, a novel flavoprotein responsible for iodide salvage in thyroid glands.	the flavoprotein iodotyrosine deiodinase (iyd) salvages iodide from mono- and diiodotyrosine formed during the biosynthesis of the thyroid hormone thyroxine. expression of a soluble domain of this membrane-bound enzyme provided sufficient material for crystallization and characterization by x-ray diffraction. the structures of iyd and two co-crystals containing substrates, mono- and diiodotyrosine, alternatively, were solved at resolutions of 2.0, 2.45, and 2.6 a, respectively. the structure of ...	2009	19436071
groel-assisted protein folding: does it occur within the chaperonin inner cavity?	the folding of protein molecules in the groel inner cavity under the co-chaperonin groes lid is widely accepted as a crucial event of groel-assisted protein folding. this review is focused on the data showing that groel-assisted protein folding may proceed out of the complex with the chaperonin. the models of groel-assisted protein folding assuming ligand-controlled dissociation of nonnative proteins from the groel surface and their folding in the bulk solution are also discussed.	2009	19564940
flexible structural protein alignment by a sequence of local transformations.	throughout evolution, homologous proteins have common regions that stay semi-rigid relative to each other and other parts that vary in a more noticeable way. in order to compare the increasing number of structures in the pdb, flexible geometrical alignments are needed, that are reliable and easy to use.	2009	19417057
the eukaryotic ribosome: current status and challenges.	despite having been identified first, their greater degree of complexity has resulted in our understanding of eukaryotic ribosomes lagging behind that of their bacterial and archaeal counterparts. a much more complicated biogenesis program results in ribosomes that are structurally, biochemically, and functionally more complex. however, recent advances in molecular genetics and structural biology are helping to reveal the intricacies of the eukaryotic ribosome and to address many longstanding qu ...	2009	19117941
distinct functions of elongation factor g in ribosome recycling and translocation.	elongation factor g (ef-g) promotes the translocation step in bacterial protein synthesis and, together with ribosome recycling factor (rrf), the disassembly of the post-termination ribosome. unlike translocation, ribosome disassembly strictly requires gtp hydrolysis by ef-g. here we report that ribosome disassembly is strongly inhibited by vanadate, an analog of inorganic phosphate (pi), indicating that pi release is required for ribosome disassembly. in contrast, the function of ef-g in single ...	2009	19324963
biogenesis of beta-barrel membrane proteins in bacteria and eukaryotes: evolutionary conservation and divergence.	membrane-embedded beta-barrel proteins span the membrane via multiple amphipathic beta-strands arranged in a cylindrical shape. these proteins are found in the outer membranes of gram-negative bacteria, mitochondria and chloroplasts. this situation is thought to reflect the evolutionary origin of mitochondria and chloroplasts from gram-negative bacterial endosymbionts. beta-barrel proteins fulfil a variety of functions; among them are pore-forming proteins that allow the flux of metabolites acro ...	2009	19399587
detection of biochemical pathways by probabilistic matching of phyletic vectors.	a phyletic vector, also known as a phyletic (or phylogenetic) pattern, is a binary representation of the presences and absences of orthologous genes in different genomes. joint occurrence of two or more genes in many genomes results in closely similar binary vectors representing these genes, and this similarity between gene vectors may be used as a measure of functional association between genes. better understanding of quantitative properties of gene co-occurrences is needed for systematic stud ...	2009	19390636
gene composer: database software for protein construct design, codon engineering, and gene synthesis.	to improve efficiency in high throughput protein structure determination, we have developed a database software package, gene composer, which facilitates the information-rich design of protein constructs and their codon engineered synthetic gene sequences. with its modular workflow design and numerous graphical user interfaces, gene composer enables researchers to perform all common bio-informatics steps used in modern structure guided protein engineering and synthetic gene engineering.	2009	19383142
the evolution of cyclodextrin glucanotransferase product specificity.	cyclodextrin glucanotransferases (cgtases) have attracted major interest from industry due to their unique capacity of forming large quantities of cyclic alpha-(1,4)-linked oligosaccharides (cyclodextrins) from starch. cgtases produce a mixture of cyclodextrins from starch consisting of 6 (alpha), 7 (beta) and 8 (gamma) glucose units. in an effort to identify the structural factors contributing to the evolutionary diversification of product specificity amongst this group of enzymes, we selected ...	2009	19367403
real-time dna microarrays: reality check.	dna microarrays are plagued with inconsistent quantifications and false-positive results. using established mechanisms of surface reactions, we argue that these problems are inherent to the current technology. in particular, the problem of multiplex non-equilibrium reactions cannot be resolved within the framework of the existing paradigm. we discuss the advantages and limitations of changing the paradigm to real-time data acquisition similar to real-time pcr methodology. our analysis suggests t ...	2009	19290884
the rna-induced silencing complex: a versatile gene-silencing machine.	rna interference is a powerful mechanism of gene silencing that underlies many aspects of eukaryotic biology. on the molecular level, rna interference is mediated by a family of ribonucleoprotein complexes called rna-induced silencing complexes (riscs), which can be programmed to target virtually any nucleic acid sequence for silencing. the ability of risc to locate target rnas has been co-opted by evolution many times to generate a broad spectrum of gene-silencing pathways. here, we review the ...	2009	19342379
identification of optimal protocols for sequencing difficult templates: results of the 2008 abrf dna sequencing research group difficult template study 2008.	the 2008 abrf dna sequencing research group (dsrg) difficult template sequencing study was designed to identify a general set of guidelines that would constitute the best approaches for sequencing difficult templates. this was a continuation of previous dsrg difficult template studies performed in 1996, 1997, and 2003. the distinguishing factors in the present study were the number of dna templates used, the number of different types of difficult regions tested, and the inclusion of a follow-up ...	2009	19503623
early events, kinetic intermediates and the mechanism of protein folding in cytochrome c.	kinetic studies of the early events in cytochrome c folding are reviewed with a focus on the evidence for folding intermediates on the submillisecond timescale. evidence from time-resolved absorption, circular dichroism, magnetic circular dichroism, fluorescence energy and electron transfer, small-angle x-ray scattering and amide hydrogen exchange studies on the t < or = 1 ms timescale reveals a picture of cytochrome c folding that starts with the approximately 1-micros conformational diffusion ...	2009	19468320
extension of the tryptophan chi2,1 dihedral angle-w3 band frequency relationship to a full rotation: correlations and caveats.	the correlation of the uvrr nuw3 mode with the tryptophan chi(2,1) dihedral angle [maruyama and takeuchi (1995) j. raman spectrosc. 26, 319; miura et al. (1989) j. raman spectrosc. 20, 667; takeuchi (2003) biopolymers 72, 305] has been extended to a full, 360 degrees rotation. the 3-fold periodicity of the relationship (cos 3chi(2,1)) over 360 degrees results in up to six dihedral angles for a given nuw3. consideration of a newman plot of dihedral angles for proteinaceous tryptophans taken from ...	2009	19267450
length variations amongst protein domain superfamilies and consequences on structure and function.	related protein domains of a superfamily can be specified by proteins of diverse lengths. the structural and functional implications of indels in a domain scaffold have been examined.	2009	19333395
recent developments in parameter estimation and structure identification of biochemical and genomic systems.	the organization, regulation and dynamical responses of biological systems are in many cases too complex to allow intuitive predictions and require the support of mathematical modeling for quantitative assessments and a reliable understanding of system functioning. all steps of constructing mathematical models for biological systems are challenging, but arguably the most difficult task among them is the estimation of model parameters and the identification of the structure and regulation of the ...	2009	19327372
slow unfolding of monomeric proteins from hyperthermophiles with reversible unfolding.	based on the differences in their optimal growth temperatures microorganisms can be classified into psychrophiles, mesophiles, thermophiles, and hyperthermophiles. proteins from hyperthermophiles generally exhibit greater stability than those from other organisms. in this review, we collect data about the stability and folding of monomeric proteins from hyperthermophilies with reversible unfolding, from the equilibrium and kinetic aspects. the results indicate that slow unfolding is a general st ...	2009	19399254
co-evolution of primordial membranes and membrane proteins.	studies of the past several decades have provided major insights into the structural organization of biological membranes and mechanisms of many membrane molecular machines. however, the origin(s) of the membrane(s) and membrane proteins remains enigmatic. we discuss different concepts of the origin and early evolution of membranes with a focus on the evolution of the (im)permeability to charged molecules such as proteins, nucleic acids and small ions. reconstruction of the evolution of f-type a ...	2009	19303305
copper trafficking in biology: an nmr approach.	copper ions are essential for living organisms because they are involved in several fundamental biological processes. biomolecules interacting with copper ions have to be characterized as such, when bound to the metal ion, and when they interact with other biomolecules or substrates. the characterization is both structural and dynamic. in this context, nmr is a preferred tool of investigation because it allows shedding light on what happens in solution. here, the nmr contribution to the copper t ...	2009	19949444
oxidatively modified proteins in alzheimer's disease (ad), mild cognitive impairment and animal models of ad: role of abeta in pathogenesis.	oxidative stress has been implicated in the pathogenesis of a number of diseases including alzheimer's disease (ad). the oxidative stress hypothesis of ad pathogenesis, in part, is based on beta-amyloid peptide (abeta)-induced oxidative stress in both in vitro and in vivo studies. oxidative modification of the protein may induce structural changes in a protein that might lead to its functional impairment. a number of oxidatively modified brain proteins were identified using redox proteomics in a ...	2009	19288120
primary structure revision and active site mapping of e. coli isoleucyl-trna synthetase by means of maldi mass spectrometry.	the correct amino acid sequence of e. coli isoleucyl-trna synthetase (ilers) was established by means of peptide mapping by maldi mass spectrometry, using a set of four endoproteases (trypsin, lysc, aspn and gluc). thereafter, the active site of ilers was mapped by affinity labeling with reactive analogs of the substrates. for the atp binding site, the affinity labeling reagent was pyridoxal 5'-diphospho-5'-adenosine (adp-pl), whereas periodate-oxidized trna(ile), the 2',3'-dialdehyde derivative ...	2009	19557155
identification of rna molecules by specific enzyme digestion and mass spectrometry: software for and implementation of rna mass mapping.	the idea of identifying or characterizing an rna molecule based on a mass spectrum of specifically generated rna fragments has been used in various forms for well over a decade. we have developed software-named rrm for 'rna mass mapping'-which can search whole prokaryotic genomes or rna fasta sequence databases to identify the origin of a given rna based on a mass spectrum of rna fragments. as input, the program uses the masses of specific rnase cleavage of the rna under investigation. rnase t1 ...	2009	19264806
the role of disordered ribosomal protein extensions in the early steps of eubacterial 50 s ribosomal subunit assembly.	although during the past decade research has shown the functional importance of disorder in proteins, many of the structural and dynamics properties of intrinsically unstructured proteins (iups) remain to be elucidated. this review is focused on the role of the extensions of the ribosomal proteins in the early steps of the assembly of the eubacterial 50 s subunit. the recent crystallographic structures of the ribosomal particles have revealed the picture of a complex assembly pathway that conden ...	2009	19399222
properties of the nucleic-acid bases in free and watson-crick hydrogen-bonded states: computational insights into the sequence-dependent features of double-helical dna.	the nucleic-acid bases carry structural and energetic signatures that contribute to the unique features of genetic sequences. here we review the connection between the chemical structure of the constituent nucleotides and the polymeric properties of dna. the sequence-dependent accumulation of charge on the major- and minor-groove edges of the watson-crick base pairs, obtained from ab initio calculations, presents unique motifs for direct sequence recognition. the optimization of base interaction ...	2009	21218180
chemically modified primers for improved multiplex polymerase chain reaction.	multiplex polymerase chain reaction (pcr), the amplification of multiple targets in a single reaction, presents a new set of challenges that further complicate more traditional pcr setups. these complications include a greater probability for nonspecific amplicon formation and for imbalanced amplification of different targets, each of which can compromise quantification and detection of multiple targets. despite these difficulties, multiplex pcr is frequently used in applications such as pathoge ...	2009	19258004
inhibition of bacterial ribosome assembly: a suitable drug target?	the assembly of bacterial ribosomes is viewed with increasing interest as a potential target for new antibiotics. the in vivo synthesis and assembly of ribosomes are briefly reviewed here, highlighting the many ways in which assembly can be perturbed. the process is compared with the model in vitro process from which much of our knowledge is derived. the coordinate synthesis of the ribosomal components is essential for their ordered and efficient assembly; antibiotics interfere with this coordin ...	2009	19258531
crystallization and preliminary x-ray diffraction analysis of a putative two-domain-type laccase from a metagenome.	a putative two-domain-type laccase retrieved from a metagenome was successfully crystallized using the sitting-drop vapour-diffusion method. data were collected to a resolution of 1.7 a at 100 k using synchrotron radiation. the crystal belonged to space group p2(1)2(1)2(1), with unit-cell parameters a = 74.67, b = 100.95, c = 124.11 a. the self-rotation function showed the presence of a noncrystallographic threefold axis in the structure. the presence of one trimer in the asymmetric unit yielded ...	2009	19255479
frequency and isostericity of rna base pairs.	most of the hairpin, internal and junction loops that appear single-stranded in standard rna secondary structures form recurrent 3d motifs, where non-watson-crick base pairs play a central role. non-watson-crick base pairs also play crucial roles in tertiary contacts in structured rna molecules. we previously classified rna base pairs geometrically so as to group together those base pairs that are structurally similar (isosteric) and therefore able to substitute for each other by mutation withou ...	2009	19240142
regulation of translation initiation in eukaryotes: mechanisms and biological targets.	translational control in eukaryotic cells is critical for gene regulation during nutrient deprivation and stress, development and differentiation, nervous system function, aging, and disease. we describe recent advances in our understanding of the molecular structures and biochemical functions of the translation initiation machinery and summarize key strategies that mediate general or gene-specific translational control, particularly in mammalian systems.	2009	19239892
origins and mechanisms of mirnas and sirnas.	over the last decade, approximately 20-30 nucleotide rna molecules have emerged as critical regulators in the expression and function of eukaryotic genomes. two primary categories of these small rnas--short interfering rnas (sirnas) and micrornas (mirnas)--act in both somatic and germline lineages in a broad range of eukaryotic species to regulate endogenous genes and to defend the genome from invasive nucleic acids. recent advances have revealed unexpected diversity in their biogenesis pathways ...	2009	19239886
evolution of mutation rates: phylogenomic analysis of the photolyase/cryptochrome family.	photoreactivation, one of the first dna repair pathways to evolve, is the direct reversal of premutagenic lesions caused by ultraviolet (uv) irradiation, catalyzed by photolyases in a light-dependent, single-enzyme reaction. it has been experimentally shown that photoreactivation prevents uv mutagenesis in a broad range of species. in the absence of photoreactivation, uv-induced photolesions are repaired by the more complex and much less efficient nucleotide excision repair pathway. despite thei ...	2009	19228922
comparison of molecular dynamics and superfamily spaces of protein domain deformation.	it is well known the strong relationship between protein structure and flexibility, on one hand, and biological protein function, on the other hand. technically, protein flexibility exploration is an essential task in many applications, such as protein structure prediction and modeling. in this contribution we have compared two different approaches to explore the flexibility space of protein domains: i) molecular dynamics (md-space), and ii) the study of the structural changes within superfamily ...	2009	19220918
molecular dynamics guided study of salt bridge length dependence in both fluorinated and non-fluorinated parallel dimeric coiled-coils.	the alpha-helical coiled-coil is one of the most common oligomerization motifs found in both native and engineered proteins. to better understand the stability and dynamics of the coiled-coil motifs, including those modified by fluorination, several fluorinated and nonfluorinated parallel dimeric coiled-coil protein structures were designed and modeled. we also attempt to investigate how changing the length and geometry of the important stabilizing salt bridges influences the coiled-coil protein ...	2009	18704948
recurring cluster and operon assembly for phenylacetate degradation genes.	a large number of theories have been advanced to explain why genes involved in the same biochemical processes are often co-located in genomes. most of these theories have been dismissed because empirical data do not match the expectations of the models. in this work we test the hypothesis that cluster formation is most likely due to a selective pressure to gradually co-localise protein products and that operon formation is not an inevitable conclusion of the process.	2009	19208251
predicting peptide structures in native proteins from physical simulations of fragments.	it has long been proposed that much of the information encoding how a protein folds is contained locally in the peptide chain. here we present a large-scale simulation study designed to examine the extent to which conformations of peptide fragments in water predict native conformations in proteins. we perform replica exchange molecular dynamics (remd) simulations of 872 8-mer, 12-mer, and 16-mer peptide fragments from 13 proteins using the amber 96 force field and the obc implicit solvent model. ...	2009	19197352
synthesizing non-natural parts from natural genomic template.	the current knowledge of genes and proteins comes from 'naturally designed' coding and non-coding regions. it would be interesting to move beyond natural boundaries and make user-defined parts. to explore this possibility we made six non-natural proteins in e. coli. we also studied their potential tertiary structure and phenotypic outcomes.	2009	19187561
the ribosome returned.	since the mid-1990s, insights obtained from electron microscopy and x-ray crystallography have transformed our understanding of how the most important ribozyme in the cell, the ribosome, catalyzes protein synthesis. this review provides a brief account of how this structural revolution came to pass, and the impact it has had on our understanding of how the ribosome decodes messenger rnas.	2009	19222865
new information content in rna base pairing deduced from quantitative analysis of high-resolution structures.	non-canonical base pairs play important roles in organizing the complex three-dimensional folding of rna. here, we outline methodology developed both to analyze the spatial patterns of interacting base pairs in known rna structures and to reconstruct models from the collective experimental information. we focus attention on the structural context and deformability of the seven pairing patterns found in greatest abundance in the helical segments in a set of well-resolved crystal structures, inclu ...	2009	19150407
regulation of translation initiation by rna binding proteins.	rna binding proteins are capable of regulating translation initiation by a variety of mechanisms. although the vast majority of these regulatory mechanisms involve translational repression, one example of translational activation has been characterized in detail. the rna recognition targets of these regulatory proteins exhibit a wide range in structural complexity, with some proteins recognizing complex pseudoknot structures and others binding to simple rna hairpins and/or short repeated single- ...	2009	19385727
determining structures of rna aptamers and riboswitches by x-ray crystallography.	structural biology plays a central role in gaining a full understanding of the myriad roles of rna in biology. in recent years, innovative approaches in rna purification and crystallographic methods have lead to the visualization of an increasing number of unique structures, providing new insights into its function at the atomic level. this article presents general protocols which have streamlined the process of obtaining a homogeneous sample of properly folded and active rna in high concentrati ...	2009	19377976
high-throughput haplotype determination over long distances by haplotype fusion pcr and ligation haplotyping.	when combined with haplotype fusion pcr (hf-pcr), ligation haplotyping is a robust, high-throughput method for empirical determination of haplotypes, which can be applied to assaying both sequence and structural variation over long distances. unlike alternative approaches to haplotype determination, such as allele-specific pcr and long pcr, hf-pcr and ligation haplotyping do not suffer from mispriming or template-switching errors. in this method, hf-pcr is used to juxtapose dna sequences from si ...	2009	20010928
lessons from structural genomics.	a decade of structural genomics, the large-scale determination of protein structures, has generated a wealth of data and many important lessons for structural biology and for future large-scale projects. these lessons include a confirmation that it is possible to construct large-scale facilities that can determine the structures of a hundred or more proteins per year, that these structures can be of high quality, and that these structures can have an important impact. technology development has ...	2009	19416074
pharmacological targeting of the hsp70 chaperone.	the molecular chaperone, heat shock protein 70 (hsp70), acts at multiple steps in a protein's life cycle, including during the processes of folding, trafficking, remodeling and degradation. to accomplish these various tasks, the activity of hsp70 is shaped by a host of co-chaperones, which bind to the core chaperone and influence its functions. genetic studies have strongly linked hsp70 and its co-chaperones to numerous diseases, including cancer, neurodegeneration and microbial pathogenesis, ye ...	2009	19860737
the biogenesis and function of piwi proteins and pirnas: progress and prospect.	the evolutionarily conserved argonaute/piwi (ago/piwi, also known as paz-piwi domain or ppd) family of proteins is crucial for the biogenesis and function of small noncoding rnas (ncrnas). this family can be divided into ago and piwi subfamilies. the ago proteins are ubiquitously present in diverse tissues. they bind to small interfering rnas (sirnas) and micrornas (mirnas). in contrast, the piwi proteins are predominantly present in the germline and associate with a novel class of small rnas kn ...	2009	19575643
stabilization of alpha-chymotrypsin upon pegylation correlates with reduced structural dynamics.	protein stability remains one of the main factors limiting the realization of the full potential of protein therapeutics. poly(ethylene glycol) (peg) conjugation to proteins has evolved into an important tool to overcome instability issues associated with proteins. the observed increase in thermodynamic stability of several proteins upon pegylation has been hypothesized to arise from reduced protein structural dynamics, although experimental evidence for this hypothesis is currently missing. to ...	2008	18781698
how mitochondria produce reactive oxygen species.	the production of ros (reactive oxygen species) by mammalian mitochondria is important because it underlies oxidative damage in many pathologies and contributes to retrograde redox signalling from the organelle to the cytosol and nucleus. superoxide (o2(-)) is the proximal mitochondrial ros, and in the present review i outline the principles that govern o2(-) production within the matrix of mammalian mitochondria. the flux of o2(*-) is related to the concentration of potential electron donors, ...	2008	19061483
how mitochondria produce reactive oxygen species.	the production of ros (reactive oxygen species) by mammalian mitochondria is important because it underlies oxidative damage in many pathologies and contributes to retrograde redox signalling from the organelle to the cytosol and nucleus. superoxide (o2(-)) is the proximal mitochondrial ros, and in the present review i outline the principles that govern o2(-) production within the matrix of mammalian mitochondria. the flux of o2(*-) is related to the concentration of potential electron donors, ...	2008	19061483
upf201 archaeal specific family members reveal structural similarity to rna-binding proteins but low likelihood for rna-binding function.	we have determined x-ray crystal structures of four members of an archaeal specific family of proteins of unknown function (upf0201; pfam classification: duf54) to advance our understanding of the genetic repertoire of archaea. despite low pairwise amino acid sequence identities (10-40%) and the absence of conserved sequence motifs, the three-dimensional structures of these proteins are remarkably similar to one another. their common polypeptide chain fold, encompassing a five-stranded antiparal ...	2008	19079550
implementation of a flash-photolysis system for time-resolved cryo-electron microscopy.	we describe here the implementation of a flash-photolysis system for time-resolved cryo-electron microscopy. a previously designed computer-controlled cryo-plunging apparatus [white, h.d., thirumurugan, k., walker, m.l., trinick, j., 2003. a second generation apparatus for time-resolved electron cryo-microscopy using stepper motors and electrospray. j. struct. biol. 144, 246-252] was used as a hardware platform, onto which a xenon flash lamp and liquid light pipe were mounted. the irradiation in ...	2008	19114106
implementation of a flash-photolysis system for time-resolved cryo-electron microscopy.	we describe here the implementation of a flash-photolysis system for time-resolved cryo-electron microscopy. a previously designed computer-controlled cryo-plunging apparatus [white, h.d., thirumurugan, k., walker, m.l., trinick, j., 2003. a second generation apparatus for time-resolved electron cryo-microscopy using stepper motors and electrospray. j. struct. biol. 144, 246-252] was used as a hardware platform, onto which a xenon flash lamp and liquid light pipe were mounted. the irradiation in ...	2008	19114106
a bridge to transcription by rna polymerase.	a comprehensive survey of single amino-acid substitution mutations critical for rna polymerase function published in journal of biology supports a proposed mechanism for polymerase action in which movement of the polymerase 'bridge helix' promotes transcriptional activity in cooperation with a critical substrate-interaction domain, the 'trigger loop'.	2008	19090964
effect of primer proximity to a difficult-to-sequence region on read length and sequence quality.	anecdotal and not well-established evidence implies that there could be some effect of primer proximity in relation to a difficult region on read length and sequence quality. in this paper we sequenced many different categories of difficult regions where primers were located at various distances in relation to such regions and we found that there is only weak, if any, correlation between primer proximity and read length or sequence quality. the occasional improvements observed in some studies co ...	2008	19183797
the ua_handle: a versatile submotif in stable rna architectures.	stable rnas are modular and hierarchical 3d architectures taking advantage of recurrent structural motifs to form extensive non-covalent tertiary interactions. sequence and atomic structure analysis has revealed a novel submotif involving a minimal set of five nucleotides, termed the ua_handle motif (5'xu/an(n)x3'). it consists of a u:a watson-crick: hoogsteen trans base pair stacked over a classic watson-crick base pair, and a bulge of one or more nucleotides that can act as a handle for making ...	2008	19036788
the ua_handle: a versatile submotif in stable rna architectures.	stable rnas are modular and hierarchical 3d architectures taking advantage of recurrent structural motifs to form extensive non-covalent tertiary interactions. sequence and atomic structure analysis has revealed a novel submotif involving a minimal set of five nucleotides, termed the ua_handle motif (5'xu/an(n)x3'). it consists of a u:a watson-crick: hoogsteen trans base pair stacked over a classic watson-crick base pair, and a bulge of one or more nucleotides that can act as a handle for making ...	2008	19036788
genome-wide survey of prokaryotic serine proteases: analysis of distribution and domain architectures of five serine protease families in prokaryotes.	serine proteases are one of the most abundant groups of proteolytic enzymes found in all the kingdoms of life. while studies have established significant roles for many prokaryotic serine proteases in several physiological processes, such as those associated with metabolism, cell signalling, defense response and development, functional associations for a large number of prokaryotic serine proteases are relatively unknown. current analysis is aimed at understanding the distribution and probable b ...	2008	19019219
the linkage between ribosomal crystallography, metal ions, heteropolytungstates and functional flexibility.	crystallography of ribosomes, the universal cell nucleoprotein assemblies facilitating the translation of the genetic-code into proteins, met with severe problems owing to their large size, complex structure, inherent flexibility and high conformational variability. for the case of the small ribosomal subunit, which caused extreme difficulties, post crystallization treatment by minute amounts of a heteropolytungstate cluster allowed structure determination at atomic resolution. this cluster play ...	2008	19915655
high tolerance for ionizable residues in the hydrophobic interior of proteins.	internal ionizable groups are quite rare in water-soluble globular proteins. presumably, this reflects the incompatibility between charges and the hydrophobic environment in the protein interior. here we show that proteins can have an inherently high tolerance for internal ionizable groups. the 25 internal positions in staphylococcal nuclease were substituted one at a time with lys, glu, or asp without abolishing enzymatic activity and without detectable changes in the conformation of the protei ...	2008	19004768
recognition of a common rdna target site in archaea and eukarya by analogous laglidadg and his-cys box homing endonucleases.	the presence of a homing endonuclease gene (heg) within a microbial intron or intein empowers the entire element with the ability to invade genomic targets. the persistence of a homing endonuclease lineage depends in part on conservation of its dna target site. one such rdna sequence has been invaded both in archaea and in eukarya, by laglidadg and his-cys box homing endonucleases, respectively. the bases encoded by this target include a universally conserved ribosomal structure, termed helix 69 ...	2008	18984620
molecular characterization of organelle-type nudix hydrolases in arabidopsis.	nudix (for nucleoside diphosphates linked to some moiety x) hydrolases act to hydrolyze ribonucleoside and deoxyribonucleoside triphosphates, nucleotide sugars, coenzymes, or dinucleoside polyphosphates. arabidopsis (arabidopsis thaliana) contains 27 genes encoding nudix hydrolase homologues (atnudx1 to -27) with a predicted distribution in the cytosol, mitochondria, and chloroplasts. previously, cytosolic nudix hydrolases (atnudx1 to -11 and -25) were characterized. here, we conducted a charact ...	2008	18815383
rna in motion.	although rna duplex regions are highly structured and inflexible, other elements of an rna molecule are capable of dynamic motions. these flexible regions are the sites of interactions with small molecules, proteins, and other rnas, yet there are few descriptions of these regions that include the timescale and amplitude of their motions. no one technique is sufficient to accurately describe these motions, but the combination of in vitro methods, particularly nmr relaxation methods, and more robu ...	2008	18957331
higher-order association states of cellular erbb3 probed with photo-cross-linkable aptamers.	nucleic acid aptamers are rapidly gaining prominence as diagnostic tools, targeting reagents, and potential therapeutics. to extend the use of aptamers into the biochemical analysis of protein interactions on the surface of live cells, we converted an enzymatically generated rna aptamer into a photo-cross-linkable affinity tag through the replacement of all uracils with 4-thiouracil. specifically, we converted a previously selected, inhibitory aptamer that binds the soluble extracellular domains ...	2008	18942860
structural basis for specific, high-affinity tetracycline binding by an in vitro evolved aptamer and artificial riboswitch.	the tetracycline aptamer is an in vitro selected rna that binds to the antibiotic with the highest known affinity of an artificial rna for a small molecule (kd approximately 0.8 nm). it is one of few aptamers known to be capable of modulating gene expression in vivo. the 2.2 a resolution cocrystal structure of the aptamer reveals a pseudoknot-like fold formed by tertiary interactions between an 11 nucleotide loop and the minor groove of an irregular helix. tetracycline binds within this interfac ...	2008	18940672
rna folding and ribosome assembly.	ribosome synthesis is a tightly regulated process that is crucial for cell survival. chemical footprinting, mass spectrometry, and cryo-electron microscopy are revealing how these complex cellular machines are assembled. rapid folding of the rrna provides a platform for protein-induced assembly of the bacterial 30s ribosome. multiple assembly pathways increase the flexibility of the assembly process, while accessory factors and modification enzymes chaperone the late stages of assembly and contr ...	2008	18935976
dna polymerases and aminoacyl-trna synthetases: shared mechanisms for ensuring the fidelity of gene expression.	dna polymerases and aminoacyl-trna synthetases (arss) represent large enzyme families with critical roles in the transformation of genetic information from dna to rna to protein. dna polymerases carry out replication and collaborate in the repair of the genome, while arss provide aminoacylated trna precursors for protein synthesis. enzymes of both families face the common challenge of selecting their cognate small molecule substrates from a pool of chemically related molecules, achieving high le ...	2008	18850722
conformationally gated metal uptake by apomanganese superoxide dismutase.	metal uptake by apomanganese superoxide dismutase in vitro is a complex process exhibiting multiphase "gated" reaction kinetics and a striking sigmoidal temperature profile that has led to a model of conformationally gated metal binding, requiring conversion between "closed" and "open" forms. this work systematically explores the structural determinants of metal binding in both wild-type (wt) apoprotein and mutational variants as a test of mechanistic models. the ph dependence of metalation unde ...	2008	18841998
the higher level of organization of the oxidative phosphorylation system: mitochondrial supercomplexes.	the organization of the oxidative phosphorylation (oxphos) system within the inner mitochondrial membrane appears to be far more complicated than previously thought. in particular, the individual protein complexes of the oxphos system (complexes i to v) were found to specifically interact forming defined supramolecular structures. blue-native polyacrylamide gel electrophoresis and single particle electron microscopy proved to be especially valuable in studying the so-called "respiratory supercom ...	2008	18839290
the pretranslocation ribosome is targeted by gtp-bound ef-g in partially activated form.	translocation of the trna x mrna complex through the bacterial ribosome is driven by the multidomain guanosine triphosphatase elongation factor g (ef-g). we have used isothermal titration calorimetry to characterize the binding of gdp and gtp to free ef-g at 4 degrees c, 20 degrees c, and 37 degrees c. the binding affinity of ef-g is higher to gdp than to gtp at 4 degrees c, but lower at 37 degrees c. the binding enthalpy and entropy change little with temperature in the case of gdp binding but ...	2008	18836081
the unique nature of mg2+ channels.	considering the biological abundance and importance of mg2+, there is a surprising lack of information regarding the proteins that transport mg2+, the mechanisms by which they do so, and their physiological roles within the cell. the best characterized mg2+ channel to date is the bacterial protein cora, present in a wide range of bacterial species. the cora homolog mrs2 forms the mitochondrial mg2+ channel in all eukaryotes. physiologically, cora is involved in bacterial pathogenesis, and the mr ...	2008	18927203
the metabolism of proline as microenvironmental stress substrate.	proline, a unique proteogenic secondary amino acid, has its own metabolic system with special features. recent findings defining the regulation of this system led us to propose that proline is a stress substrate in the microenvironment of inflammation and tumorigenesis. the criteria for proline as a stress substrate are: 1) the enzymes utilizing proline respond to stress signaling; 2) there is a large, mobilizable pool of proline; and 3) the metabolism of proline serves special stress functions. ...	2008	18806116
the medium-chain dehydrogenase/reductase engineering database: a systematic analysis of a diverse protein family to understand sequence-structure-function relationship.	the medium-chain dehydrogenase/reductase engineering database (mdred, http://www.mdred.uni-stuttgart.de) has been established to serve as an analysis tool for a systematic investigation of sequence-structure-function relationships. it includes sequence and structure information of 2684 and 42 medium-chain dehydrogenases/reductases (mdrs), respectively. although mdrs are very diverse in sequence, they have a conserved tertiary structure. mdrs are assigned to 199 homologous families and 29 superfa ...	2008	18614751
visualization of the eef2-80s ribosome transition-state complex by cryo-electron microscopy.	in an attempt to understand ribosome-induced gtp hydrolysis on eef2, we determined a 12.6-a cryo-electron microscopy reconstruction of the eef2-bound 80s ribosome in the presence of aluminum tetrafluoride and gdp, with aluminum tetrafluoride mimicking the gamma-phosphate during hydrolysis. this is the first visualization of a structure representing a transition-state complex on the ribosome. tight interactions are observed between the factor's g domain and the large ribosomal subunit, as well as ...	2008	18644383
effects of protein subunits removal on the computed motions of partial 30s structures of the ribosome.	the anisotropic network model (anm) is used to study motions of the 30s small ribosomal subunit. the effect of the absence of certain subunits on the motions of the remaining partial structures was investigated by removing one protein, pairs of proteins and selected sets of proteins at a time. our results show that the removal of some proteins doesn't change the large-scale dynamics of the partial structures, but the removal of certain subunits does cause significant changes in motion of the rem ...	2008	19771145
genome signature analysis of thermal virus metagenomes reveals archaea and thermophilic signatures.	metagenomic analysis provides a rich source of biological information for otherwise intractable viral communities. however, study of viral metagenomes has been hampered by its nearly complete reliance on blast algorithms for identification of dna sequences. we sought to develop algorithms for examination of viral metagenomes to identify the origin of sequences independent of blast algorithms. we chose viral metagenomes obtained from two hot springs, bear paw and octopus, in yellowstone national ...	2008	18798991
evolution of catalases from bacteria to humans.	excessive hydrogen peroxide is harmful for almost all cell components, so its rapid and efficient removal is of essential importance for aerobically living organisms. conversely, hydrogen peroxide acts as a second messenger in signal-transduction pathways. h(2)o(2) is degraded by peroxidases and catalases, the latter being able both to reduce h(2)o(2) to water and to oxidize it to molecular oxygen. nature has evolved three protein families that are able to catalyze this dismutation at reasonable ...	2008	18498226