CiteULike: aalibes's library [269 articles]

Principles of protein-DNA recognition revealed in the structural analysis of Ndt80-MSE DNA complexes.

JS Lamoureux — 2008-07-29T11:21:51-00:00

Structure (London, England : 1993), Vol. 14, No. 3. (March 2006), pp. 555-565.

The Saccharomyces cerevisiae transcription factor Ndt80 selectively binds a DNA consensus sequence (the middle sporulation element [MSE]) to activate gene expression after the successful completion of meiotic recombination. Here we report the X-ray crystal structures of Ndt80 bound to ten distinct MSE variants. Comparison of these structures with the structure of Ndt80 bound to a consensus MSE reveals structural principles that determine the DNA binding specificity of this transcription factor. The 5' GC-rich end of the MSE contains distinct 5'-YpG-3' steps that are recognized by arginine side chains through a combination of hydrogen bonding and cation-pi interactions. The 3' AT-rich region is recognized via minor groove contacts that sterically exclude the N2 atom of GC base pairs. The conformation of the AT-rich region is fixed by interactions with the protein that favor recognition of poly(A)-poly(T) versus mixed AT sequences through an avoidance of major groove steric clashes at 5'-ApT-3' steps.

Design of Compact, Universal DNA Microarrays for Protein Binding Microarray Experiments.

Anthony A Philippakis — 2008-07-28T09:13:44-00:00

Journal of computational biology : a journal of computational molecular cell biology (23 July 2008)

Our group has recently developed a compact, universal protein binding microarray (PBM) that can be used to determine the binding preferences of transcription factors (TFs). This design represents all possible sequence variants of a given length k (i.e., all k-mers) on a single array, allowing a complete characterization of the binding specificities of a given TF. Here, we present the mathematical foundations of this design based on de Bruijn sequences generated by linear feedback shift registers. We show that these sequences represent the maximum number of variants for any given set of array dimensions (i.e., number of spots and spot lengths), while also exhibiting desirable pseudo-randomness properties. Moreover, de Bruijn sequences can be selected that represent gapped sequence patterns, further increasing the coverage of the array. This design yields a powerful experimental platform that allows the binding preferences of TFs to be determined with unprecedented resolution.

Prediction by Graph Theoretic Measures of Structural Effects in Proteins Arising from Non-Synonymous Single Nucleotide Polymorphisms

Tammy Cheng — 2008-07-26T13:54:09-00:00

PLoS Comput Biol, Vol. 4, No. 7. (25 July 2008), e1000135.

Recent analyses of human genome sequences have given rise to impressive advances in identifying non-synonymous single nucleotide polymorphisms (nsSNPs). By contrast, the annotation of nsSNPs and their links to diseases are progressing at a much slower pace. Many of the current approaches to analysing disease-associated nsSNPs use primarily sequence and evolutionary information, while structural information is relatively less exploited. In order to explore the potential of such information, we developed a structure-based approach, Bongo (Bonds ON Graph), to predict structural effects of nsSNPs. Bongo considers protein structures as residue–residue interaction networks and applies graph theoretical measures to identify the residues that are critical for maintaining structural stability by assessing the consequences on the interaction network of single point mutations. Our results show that Bongo is able to identify mutations that cause both local and global structural effects, with a remarkably low false positive rate. Application of the Bongo method to the prediction of 506 disease-associated nsSNPs resulted in a performance (positive predictive value, PPV, 78.5%) similar to that of PolyPhen (PPV, 77.2%) and PANTHER (PPV, 72.2%). As the Bongo method is solely structure-based, our results indicate that the structural changes resulting from nsSNPs are closely associated to their pathological consequences.

Probing potential binding modes of the p53 tetramer to DNA based on the symmetries encoded in p53 response elements.

Buyong Ma — 2007-11-13T15:11:42-00:00

Nucleic Acids Res (5 November 2007)

Symmetries in the p53 response-element (p53RE) encode binding modes for p53 tetramer to recognize DNA. We investigated the molecular mechanisms and biological implications of the possible binding modes. The probabilities evaluated with molecular dynamics simulations and DNA sequence analyses were found to be correlated, indicating that p53 tetramer models studied here are able to read DNA sequence information. The traditionally believed mode with four p53 monomers binding at all four DNA quarter-sites does not cause linear DNA to bend. Alternatively, p53 tetramer can use only two monomers to recognize DNA sequence and induce DNA bending. With an arrangement of dimer of AB dimer observed in p53 trimer-DNA complex crystal, p53 can recognize supercoiled DNA sequence-specifically by binding to quarter-sites one and four (H14 mode) and recognize Holliday junction geometry-specifically. Examining R273H mutation and p53-DNA interactions, we found that at least three R273H monomers are needed to disable the p53 tetramer, consistent with experiments. But just one R273H monomer may greatly shift the binding mode probabilities. Our work suggests that p53 needs balanced binding modes to maintain genome stability. Inverse repeat p53REs favor the H14 mode and direct repeat p53REs may have high possibilities of other modes.

Weight matrices for protein-DNA binding sites from a single co-crystal structure.

RG Endres — 2008-07-24T15:01:04-00:00

Physical review. E, Statistical, nonlinear, and soft matter physics, Vol. 73, No. 6 Pt 1. (June 2006)

Transcription-factor proteins bind to specific DNA sequences to regulate gene expression in cells. DNA-binding sites are often identified using weight matrices calculated from multiple known binding sites. However, in many cases the number of examples is limited. Here, we report on an atomistic method that starts from an x-ray co-crystal structure of the protein bound to one particular DNA sequence, and infers other binding sites, which are used to construct a weight matrix. The emphasis of the paper is on using the Wang-Landau Monte Carlo algorithm to efficiently sample high-affinity binding sites, which demonstrates that sampling can produce accurate weight matrices in analogy to bioinformatics approaches. For cases of low complexity, we compare to the exhaustive (but slow) dead-end elimination algorithm. To recover crystal binding sites, it is important to include bound water in the protein-DNA interface. Our approach can, in principle, even be applied when no native protein-DNA co-crystal structure is available, only the structure of a closely related homologous protein whose amino-acid sequence is changed to the protein of interest.

Protein-DNA recognition patterns and predictions.

A Sarai — 2006-05-09T21:30:27-00:00

Annu Rev Biophys Biomol Struct, Vol. 34 (2005), pp. 379-398.

Structural data on protein-DNA complexes provide clues for understanding the mechanism of protein-DNA recognition. Although the structures of a large number of protein-DNA complexes are known, the mechanisms underlying their specific binding are still only poorly understood. Analysis of these structures has shown that there is no simple one-to-one correspondence between bases and amino acids within protein-DNA complexes; nevertheless, the observed patterns of interaction carry important information on the mechanisms of protein-DNA recognition. In this review, we show how the patterns of interaction, either observed in known structures or derived from computer simulations, confer recognition specificity, and how they can be used to examine the relationship between structure and specificity and to predict target DNA sequences used by regulatory proteins.

Ab initio prediction of transcription factor binding sites.

LA Liu — 2007-11-12T09:14:17-00:00

Pac Symp Biocomput (2007), pp. 484-495.

Transcription factors are DNA-binding proteins that control gene transcription by binding specific short DNA sequences. Experiments that identify transcription factor binding sites are often laborious and expensive, and the binding sites of many transcription factors remain unknown. We present a computational scheme to predict the binding sites directly from transcription factor sequence using all-atom molecular simulations. This method is a computational counterpart to recent high-throughput experimental technologies that identify transcription factor binding sites (ChIP-chip and protein-dsDNA binding microarrays). The only requirement of our method is an accurate 3D structural model of a transcription factor-DNA complex. We apply free energy calculations by thermodynamic integration to compute the change in binding energy of the complex due to a single base pair mutation. By calculating the binding free energy differences for all possible single mutations, we construct a position weight matrix for the predicted binding sites that can be directly compared with experimental data. As water-bridged hydrogen bonds between the transcription factor and DNA often contribute to the binding specificity, we include explicit solvent in our simulations. We present successful predictions for the yeast MAT-alpha2 homeodomain and GCN4 bZIP proteins. Water-bridged hydrogen bonds are found to be more prevalent than direct protein-DNA hydrogen bonds at the binding interfaces, indicating why empirical potentials with implicit water may be less successful in predicting binding. Our methodology can be applied to a variety of DNA-binding proteins.

Three-dimensional intricacies in protein-DNA recognition and transcriptional control

Stephen Harrison — 2007-12-06T17:53:39-00:00

Nature Structural & Molecular Biology, Vol. 14, No. 12., pp. 1118-1119.

Understanding the molecular machinery of genetics through 3D structures.

Roman A Laskowski — 2008-01-10T10:50:30-00:00

Nat Rev Genet (27 December 2007)

Detailed knowledge of the three-dimensional structures of biological molecules has had an enormous impact on all areas of biological science, including genetics, as structure can reveal the fine details of how molecules perform their biological functions. Here we consider how changes in protein sequence affect the corresponding 3D structure, and describe how structural information about proteins, DNA and chromatin has shed light on gene regulatory mechanisms and the storage and transmission of epigenetic information. Finally, we describe how structure determination is benefiting from the high-throughput technologies of the worldwide structural genomics projects.

From the first protein structures to our current knowledge of protein folding: delights and scepticisms.

Alan R Fersht — 2008-06-27T07:04:03-00:00

Nature reviews. Molecular cell biology (25 June 2008)

Every breakthrough that opens new vistas also removes the ground from under the feet of other scientists. The scientific joy of those who have seen the new light is accompanied by the dismay of those whose way of life has been changed for ever. The publication of the first structures of proteins at atomic resolution 50 years ago astounded and inspired scientists in every field, but caused others to flee or scoff. That advance and every subsequent paradigm-shifting breakthrough in protein science have met with some resistance before universal acceptance. I relate these events and their impact on the field of protein folding.

Computational prediction of transcription-factor binding site locations.

ML Bulyk — 2006-06-26T19:39:49-00:00

Genome Biol, Vol. 5, No. 1. (2003)

Identifying genomic locations of transcription-factor binding sites, particularly in higher eukaryotic genomes, has been an enormous challenge. Various experimental and computational approaches have been used to detect these sites; methods involving computational comparisons of related genomes have been particularly successful.

Protein-DNA binding specificity predictions with structural models

Alexandre Morozov — 2005-10-31T12:05:28-00:00

Nucleic Acids Research, Vol. 33, No. 18. (2005), pp. 5781-5798.

Context-dependent DNA recognition code for C2H2 zinc-finger transcription factors.

Jiajian Liu — 2008-07-07T11:10:34-00:00

Bioinformatics (Oxford, England) (27 June 2008)

MOTIVATION: Modeling and identifying the DNA-protein recognition code is one of the most challenging problems in computational biology. Several quantitative methods have been developed to model DNA-protein interactions with specific focus on the C2H2 zinc finger proteins, the largest transcription factor family in eukaryotic genomes. In many cases, they performed well. But the overall the predictive accuracy of these methods is still limited. One of the major reasons is all these methods used weight matrix models to represent DNA-protein interactions, assuming all base-amino acid contacts contribute independently to the total free energy of binding. RESULTS: We present a context-dependent model for DNA-zinc finger protein interactions that allows us to identify inter-positional dependencies in the DNA recognition code for C2H2 zinc finger proteins. The degree of non-independence was detected by comparing the linear perceptron model with the non-linear neural net (NN) model for their predictions of DNA-zinc finger protein interactions. This dependency is supported by the complex base-amino acid contacts observed in DNA-zinc finger interactions from structural analyses. Using extensive published qualitative and quantitative experimental data, we demonstrated that the context-dependent model developed in this study can significantly improves predictions of DNA binding profiles and free energies of binding for both individual zinc fingers and proteins with multiple zinc fingers when comparing to previous positional-independent models. This approach can be extended to other protein families with complex base-amino acid residue interactions that would help to further understand the transcriptional regulation in eukaryotic genomes. AVAILABILITY: The software implemented as c programs and are available by request. http://ural.wustl.edu/softwares.html CONTACT: stormo@ural.wustl.edu.

Two strategies for gene regulation by promoter nucleosomes

Itay Tirosh — 2008-04-30T23:22:56-00:00

Genome Res. (30 April 2008), gr.076059.108.

Chromatin structure is central for the regulation of gene expression, but its genome-wide organization is only beginning to be understood. Here, we examine the connection between patterns of nucleosome occupancy and the capacity to modulate gene expression upon changing conditions, i.e. transcriptional plasticity. By analyzing a genome-wide data of nucleosome positioning in yeast, we find that the presence of nucleosomes close to the transcription start site is associated with high transcriptional plasticity, while nucleosomes at more distant upstream positions are negatively correlated with transcriptional plasticity. Based on this, we identify two typical promoter structures associated with low or high plasticity, respectively. The first class is characterized by a relatively large nucleosome free region close to the start site coupled with well-positioned nucleosomes further upstream, whereas the second class displays a more evenly distributed and dynamic nucleosome positioning, with high occupancy close to the start site. The two classes are further distinguished by multiple promoter features, including histone turnover, binding sites location, H2A.Z occupancy, expression noise and expression diversity. Analysis of nucleosome positioning in human promoters reproduce the main observations. Our results suggest two distinct strategies for gene regulation by chromatin, which are selectively employed by different genes. 10.1101/gr.076059.108

A high-resolution map of nucleosome positioning on a fission yeast centromere

Jun Song — 2008-07-02T16:01:56-00:00

Genome Res., Vol. 18, No. 7. (1 July 2008), pp. 1064-1072.

A key element for defining the centromere identity is the incorporation of a specific histone H3, CENPA, known as Cnp1p in Schizosaccharomyces pombe. Previous studies have suggested that functional S. pombe centromeres lack regularly positioned nucleosomes and may involve chromatin remodeling as a key step of kinetochore assembly. We used tiling microarrays to show that nucleosomes are, in fact, positioned in regular intervals in the core of centromere 2, providing the first high-resolution map of regional centromere chromatin. Nucleosome locations are not disrupted by mutations in kinetochore protein genes cnp1, mis18, mis12, nuf2, mal2; overexpression of cnp1; or the deletion of ams2, which encodes a GATA-like factor participating in CENPA incorporation. Bioinformatics analysis of the centromere sequence indicates certain enriched motifs in linker regions between nucleosomes and reveals a sequence bias in nucleosome positioning. In addition, sequence analysis of nucleosome-free regions identifies novel binding sites of Ams2p. We conclude that centromeric nucleosome positions are stable and may be derived from the underlying DNA sequence. 10.1101/gr.075374.107

Spatial effects on the speed and reliability of protein-DNA search

Zeba Wunderlich — 2008-06-26T09:48:19-00:00

Nucl. Acids Res., Vol. 36, No. 11. (1 June 2008), pp. 3570-3578.

Strong experimental and theoretical evidence shows that transcription factors (TFs) and other specific DNA-binding proteins find their sites using a two-mode search: alternating between three-dimensional (3D) diffusion through the cell and one-dimensional (1D) sliding along the DNA. We show that, due to the 1D component of the search process, the search time of a TF can depend on the initial position of the TF. We formalize this effect by discriminating between two types of searches: global and local. Using analytical calculations and simulations, we estimate how close a TF and binding site need to be to make a local search likely. We then use our model to interpret the wide range of experimental measurements of this parameter. We also show that local and global searches differ significantly in average search time and the variability of search time. These results lead to a number of biological implications, including suggestions of how prokaryotes achieve rapid gene regulation and the relationship between the search mechanism and noise in gene expression. Lastly, we propose a number of experiments to verify the existence and quantify the extent of spatial effects on the TF search process in prokaryotes. 10.1093/nar/gkn173

DNA physical properties determine nucleosome occupancy from yeast to fly

Vincent Miele — 2008-06-18T08:19:58-00:00

Nucl. Acids Res. (17 May 2008), gkn262.

Nucleosome positioning plays an essential role in cellular processes by modulating accessibility of DNA to proteins. Here, using only sequence-dependent DNA flexibility and intrinsic curvature, we predict the nucleosome occupancy along the genomes of Saccharomyces cerevisiae and Drosophila melanogaster and demonstrate the predictive power and universality of our model through its correlation with experimentally determined nucleosome occupancy data. In yeast promoter regions, the computed average nucleosome occupancy closely superimposes with experimental data, exhibiting a <200 bp region unfavourable for nucleosome formation bordered by regions that facilitate nucleosome formation. In the fly, our model faithfully predicts promoter strength as encoded in distinct chromatin architectures characteristic of strongly and weakly expressed genes. We also predict that nucleosomes are repositioned by active mechanisms at the majority of fly promoters. Our model uses only basic physical properties to describe the wrapping of DNA around the histone core, yet it captures a substantial part of chromatin's structural complexity, thus leading to a much better prediction of nucleosome occupancy than methods based merely on periodic curved DNA motifs. Our results indicate that the physical properties of the DNA chain, and not just the regulatory factors and chromatin-modifying enzymes, play key roles in eukaryotic transcription. 10.1093/nar/gkn262

Inaugural Article: The direction of gut looping is established by changes in the extracellular matrix and in cell:cell adhesion

Natasza Kurpios — 2008-06-25T09:21:50-00:00

Proceedings of the National Academy of Sciences, Vol. 105, No. 25. (24 June 2008), pp. 8499-8506.

The counterclockwise coiling of the intestines is initiated by a leftward tilt of the primitive gut tube, imparted by left-right asymmetries in the architecture of the dorsal mesentery. In silico analysis suggests that this is achieved by synergistic changes in its epithelium and mesenchyme. Within the mesenchymal compartment, cells are more densely packed on the left than on the right. In silico results indicate that this property can result from asymmetries in both extracellular matrix (ECM) and cell:cell adhesion. We find that the dorsal mesentery ECM is indeed left-right asymmetric and moreover that the adhesion molecule N-cadherin is expressed exclusively on the left side. These asymmetries are regulated by the asymmetrically expressed transcription factors Pitx2 and Isl1. Functional studies demonstrate that N-cadherin acts upstream of the changes in the ECM and is both necessary and sufficient to explain the asymmetric packing of the mesenchymal cells. 10.1073/pnas.0803578105

Statistical distributions of nucleosomes: nonrandom locations by a stochastic mechanism.

RD Kornberg — 2008-06-20T15:19:00-00:00

Nucleic acids research, Vol. 16, No. 14A. (25 July 1988), pp. 6677-6690.

Expressions are derived for distributions of nucleosomes in chromatin. Nucleosomes are placed on DNA at the densities found in bulk chromatin, and their locations are allowed to vary at random. No further assumptions are required to simulate the periodic patterns of digestion obtained with various nucleases. The introduction of a boundary constraint, due for example to sequence-specific protein binding, results in an array of regularly spaced nucleosomes at nonrandom locations, similar to the arrays reported for some genes and other chromosomal regions.

Crystal structures of I-SceI complexed to nicked DNA substrates: snapshots of intermediates along the DNA cleavage reaction pathway

Carmen Moure — 2008-06-19T14:43:20-00:00

Nucl. Acids Res., Vol. 36, No. 10. (1 June 2008), pp. 3287-3296.

I-SceI is a homing endonuclease that specifically cleaves an 18-bp double-stranded DNA. I-SceI exhibits a strong preference for cleaving the bottom strand DNA. The published structure of I-SceI bound to an uncleaved DNA substrate provided a mechanism for bottom strand cleavage but not for top strand cleavage. To more fully elucidate the I-SceI catalytic mechanism, we determined the X-ray structures of I-SceI in complex with DNA substrates that are nicked in either the top or bottom strands. The structures resemble intermediates along the DNA cleavage reaction. In a structure containing a nick in the top strand, the spatial arrangement of metal ions is similar to that observed in the structure that contains uncleaved DNA, suggesting that cleavage of the bottom strand occurs by a common mechanism regardless of whether this strand is cleaved first or second. In the structure containing a nick in the bottom strand, a new metal binding site is present in the active site that cleaves the top strand. This new metal and a candidate nucleophilic water molecule are correctly positioned to cleave the top strand following bottom strand cleavage, providing a plausible mechanism for top strand cleavage. 10.1093/nar/gkn178

Zinc-finger Nucleases: The Next Generation Emerges.

Toni Cathomen — 2008-06-17T12:13:30-00:00

Molecular therapy : the journal of the American Society of Gene Therapy (10 June 2008)

Methods of modifying the human genome precisely and efficiently hold great promise for revolutionizing the gene therapy arena. One particularly promising technology is based on the homologous recombination (HR) pathway and is known as gene targeting. Until recently, the low frequency of HR in mammalian cells, and the resulting dependence on selection to identify these rare events, has prevented gene targeting from being applied in a therapeutic context. However, recent advances in generating customized zinc-finger nucleases (ZFNs) that can create a DNA double-strand break (DSB) at preselected sites in the human genome have paved the way for HR-based strategies in gene therapy. By introducing a DSB into a target locus of interest, ZFNs stimulate gene targeting by several orders of magnitude through activation of cellular DNA repair pathways. The capability of this technology to achieve gene conversion frequencies of up to 29% in the absence of selection demonstrates its potential power. In this paper we review recent advances in, and upcoming challenges for, this emerging technology and discuss future experimental work that will be needed to bring ZFNs safely into a clinical setting.Molecular Therapy (2008); doi:10.1038/mt.2008.114.

A barrier nucleosome model for statistical positioning of nucleosomes throughout the yeast genome

Travis Mavrich — 2008-06-13T03:04:32-00:00

Genome Res. (12 June 2008), gr.078261.108.

Most nucleosomes are well-organized at the 5' ends of S. cerevisiae genes where "-1" and "+1" nucleosomes bracket a nucleosome-free promoter region (NFR). How nucleosomal organization is specified by the genome is less clear. Here we establish and inter-relate rules governing genomic nucleosome organization by sequencing DNA from more than one million immunopurified S. cerevisiae nucleosomes (displayed at http://atlas.bx.psu.edu/). Evidence is presented that the organization of nucleosomes throughout genes is largely a consequence of statistical packing principles. The genomic sequence specifies the location of the -1 and +1 nucleosomes. The +1 nucleosome forms a barrier against which nucleosomes are packed, resulting in uniform positioning, which decays at farther distances from the barrier. We present evidence for a novel 3' NFR that is present at >95% of all genes. 3' NFRs may be important for transcription termination and anti-sense initiation. We present a high-resolution genome-wide map of TFIIB locations that implicates 3' NFRs in gene looping. 10.1101/gr.078261.108

A high-resolution, nucleosome position map of C. elegans reveals a lack of universal sequence-dictated positioning

Anton Valouev — 2008-05-15T22:59:25-00:00

Genome Res. (13 May 2008), gr.076463.108.

Using the massively parallel technique of Sequencing by Oligonucleotide Ligation and Detection (SOLiD) we have assessed the in vivo positions of more than 44 million putative nucleosome cores in the multicellular genetic model organism Caenorhabditis elegans. These analyses provide a global view of the chromatin architecture of a multicellular animal at extremely high density and resolution. While we observe some degree of reproducible positioning throughout the genome in our mixed stage population of animals, we note that the major chromatin feature in the worm is a diversity of allowed nucleosome positions at the vast majority of individual loci. While absolute positioning of nucleosomes can vary substantially, relative positioning of nucleosomes (in a repeated array structure likely to be maintained at least in part by steric constraints) appears to be a significant property of chromatin structure. The high density of nucleosomal reads enabled a substantial extension of previous analysis describing the usage of individual oligonucleotide sequences along the span of the nucleosome core and linker. We release this dataset, via the UCSC Genome Browser, as a resource for the high-resolution analysis of chromatin conformation and DNA accessibility at individual loci within the C. elegans genome. 10.1101/gr.076463.108

Specific DNA-binding by Apicomplexan AP2 transcription factors

Erandi De Silva — 2008-06-10T12:36:17-00:00

Proceedings of the National Academy of Sciences (9 June 2008), 0801993105.

Malaria remains one of the most prevalent infectious diseases worldwide, affecting more than half a billion people annually. Despite many years of research, the mechanisms underlying transcriptional regulation in the malaria-causing Plasmodium spp., and in Apicomplexan parasites generally, remain poorly understood. In Plasmodium, few regulatory elements sufficient to drive gene expression have been characterized, and their cognate DNA-binding proteins remain unknown. This study characterizes the DNA-binding specificities of two members of the recently identified Apicomplexan AP2 (ApiAP2) family of putative transcriptional regulators from Plasmodium falciparum. The ApiAP2 proteins contain AP2 domains homologous to the well characterized plant AP2 family of transcriptional regulators, which play key roles in development and environmental stress response pathways. We assayed ApiAP2 protein-DNA interactions using protein-binding microarrays and combined these results with computational predictions of coexpressed target genes to couple these putative trans factors to corresponding cis-regulatory motifs in Plasmodium. Furthermore, we show that protein-DNA sequence specificity is conserved in orthologous proteins between phylogenetically distant Apicomplexan species. The identification of the DNA-binding specificities for ApiAP2 proteins lays the foundation for the exploration of their role as transcriptional regulators during all stages of parasite development. Because of their origin in the plant lineage, ApiAP2 proteins have no homologues in the human host and may prove to be ideal antimalarial targets. 10.1073/pnas.0801993105

Cell cycle-specified fluctuation of nucleosome occupancy at gene promoters.

GJ Hogan — 2008-06-03T08:10:50-00:00

PLoS genetics, Vol. 2, No. 9. (22 September 2006)

The packaging of DNA into nucleosomes influences the accessibility of underlying regulatory information. Nucleosome occupancy and positioning are best characterized in the budding yeast Saccharomyces cerevisiae, albeit in asynchronous cell populations or on individual promoters such as PHO5 and GAL1-10. Using FAIRE (formaldehyde-assisted isolation of regulatory elements) and whole-genome microarrays, we examined changes in nucleosome occupancy throughout the mitotic cell cycle in synchronized populations of S. cerevisiae. Perhaps surprisingly, nucleosome occupancy did not exhibit large, global variation between cell cycle phases. However, nucleosome occupancy at the promoters of cell cycle-regulated genes was reduced specifically at the cell cycle phase in which that gene exhibited peak expression, with the notable exception of S-phase genes. We present data that establish FAIRE as a high-throughput method for assaying nucleosome occupancy. For the first time in any system, nucleosome occupancy was mapped genome-wide throughout the cell cycle. Fluctuation of nucleosome occupancy at promoters of most cell cycle-regulated genes provides independent evidence that periodic expression of these genes is controlled mainly at the level of transcription. The promoters of G2/M genes are distinguished from other cell cycle promoters by an unusually low baseline nucleosome occupancy throughout the cell cycle. This observation, coupled with the maintenance throughout the cell cycle of the stereotypic nucleosome occupancy states between coding and non-coding loci, suggests that the largest component of variation in nucleosome occupancy is "hard wired," perhaps at the level of DNA sequence.

DBD-Hunter: a knowledge-based method for the prediction of DNA-protein interactions

Mu Gao — 2008-06-02T10:25:02-00:00

Nucl. Acids Res. (31 May 2008), gkn332.

The structures of DNA-protein complexes have illuminated the diversity of DNA-protein binding mechanisms shown by different protein families. This lack of generality could pose a great challenge for predicting DNA-protein interactions. To address this issue, we have developed a knowledge-based method, DNA-binding Domain Hunter (DBD-Hunter), for identifying DNA-binding proteins and associated binding sites. The method combines structural comparison and the evaluation of a statistical potential, which we derive to describe interactions between DNA base pairs and protein residues. We demonstrate that DBD-Hunter is an accurate method for predicting DNA-binding function of proteins, and that DNA-binding protein residues can be reliably inferred from the corresponding templates if identified. In benchmark tests on [~]4000 proteins, our method achieved an accuracy of 98% and a precision of 84%, which significantly outperforms three previous methods. We further validate the method on DNA-binding protein structures determined in DNA-free (apo) state. We show that the accuracy of our method is only slightly affected on apo-structures compared to the performance on holo-structures cocrystallized with DNA. Finally, we apply the method to [~]1700 structural genomics targets and predict that 37 targets with previously unknown function are likely to be DNA-binding proteins. DBD-Hunter is freely available at http://cssb.biology.gatech.edu/skolnick/webservice/DBD-Hunter/. 10.1093/nar/gkn332

Re-cracking the nucleosome positioning code.

MR Segal — 2008-05-13T15:11:54-00:00

Statistical applications in genetics and molecular biology, Vol. 7 (2008)

Nucleosomes, the fundamental repeating subunits of all eukaryotic chromatin, are responsible for packaging DNA into chromosomes inside the cell nucleus and controlling gene expression. While it has been well established that nucleosomes exhibit higher affinity for select DNA sequences, until recently it was unclear whether such preferences exerted a significant, genome-wide effect on nucleosome positioning in vivo. This question was seemingly and recently resolved in the affirmative: a wide-ranging series of experimental and computational analyses provided extensive evidence that the instructions for wrapping DNA around nucleosomes are contained in the DNA itself. This subsequently labeled second genetic code was based on data-driven, structural, and biophysical considerations. It was subjected to an extensive suite of validation procedures, with one conclusion being that intrinsic, genome-encoded, nucleosome organization explains approximately 50% of in vivo nucleosome positioning. Here, we revisit both the nature of the underlying sequence preferences, and the performance of the proposed code. A series of new analyses, employing spectral envelope (Fourier transform) methods for assessing key sequence periodicities, classification techniques for evaluating predictive performance, and discriminatory motif finding methods for devising alternate models, are applied. The findings from the respective analyses indicate that signature dinucleotide periodicities are absent from the bulk of the high affinity nucleosome-bound sequences, and that the predictive performance of the code is modest. We conclude that further exploration of the role of sequence-based preferences in genome-wide nucleosome positioning is warranted. This work offers a methodologic counterpart to a recent, high resolution determination of nucleosome positioning that also questions the accuracy of the proposed code and, further, provides illustrations of techniques useful in assessing sequence periodicity and predictive performance.

DNA conformations and their sequence preferences

Daniel Svozil — 2008-05-13T15:06:22-00:00

Nucl. Acids Res. (13 May 2008), gkn260.

The geometry of the phosphodiester backbone was analyzed for 7739 dinucleotides from 447 selected crystal structures of naked and complexed DNA. Ten torsion angles of a near-dinucleotide unit have been studied by combining Fourier averaging and clustering. Besides the known variants of the A-, B- and Z-DNA forms, we have also identified combined A + B backbone-deformed conformers, e.g. with alpha/gamma switches, and a few conformers with a syn orientation of bases occurring e.g. in G-quadruplex structures. A plethora of A- and B-like conformers show a close relationship between the A- and B-form double helices. A comparison of the populations of the conformers occurring in naked and complexed DNA has revealed a significant broadening of the DNA conformational space in the complexes, but the conformers still remain within the limits defined by the A- and B- forms. Possible sequence preferences, important for sequence-dependent recognition, have been assessed for the main A and B conformers by means of statistical goodness-of-fit tests. The structural properties of the backbone in quadruplexes, junctions and histone-core particles are discussed in further detail. 10.1093/nar/gkn260

How protein stability and new functions trade off.

N Tokuriki — 2008-05-13T09:08:34-00:00

PLoS computational biology, Vol. 4, No. 2. (February 2008)

Numerous studies have noted that the evolution of new enzymatic specificities is accompanied by loss of the protein's thermodynamic stability (DeltaDeltaG), thus suggesting a tradeoff between the acquisition of new enzymatic functions and stability. However, since most mutations are destabilizing (DeltaDeltaG>0), one should ask how destabilizing mutations that confer new or altered enzymatic functions relative to all other mutations are. We applied DeltaDeltaG computations by FoldX to analyze the effects of 548 mutations that arose from the directed evolution of 22 different enzymes. The stability effects, location, and type of function-altering mutations were compared to DeltaDeltaG changes arising from all possible point mutations in the same enzymes. We found that mutations that modulate enzymatic functions are mostly destabilizing (average DeltaDeltaG = +0.9 kcal/mol), and are almost as destabilizing as the "average" mutation in these enzymes (+1.3 kcal/mol). Although their stability effects are not as dramatic as in key catalytic residues, mutations that modify the substrate binding pockets, and thus mediate new enzymatic specificities, place a larger stability burden than surface mutations that underline neutral, non-adaptive evolutionary changes. How are the destabilizing effects of functional mutations balanced to enable adaptation? Our analysis also indicated that many mutations that appear in directed evolution variants with no obvious role in the new function exert stabilizing effects that may compensate for the destabilizing effects of the crucial function-altering mutations. Thus, the evolution of new enzymatic activities, both in nature and in the laboratory, is dependent on the compensatory, stabilizing effect of apparently "silent" mutations in regions of the protein that are irrelevant to its function.

Generation and analysis of mesophilic variants of the thermostable archaeal I-DmoI homing endonuclease.

J Prieto — 2008-05-13T08:42:52-00:00

The Journal of biological chemistry, Vol. 283, No. 7. (15 February 2008), pp. 4364-4374.

The hyperthermophilic archaeon Desulfurococcus mobilis I-DmoI protein belongs to the family of proteins known as homing endonucleases (HEs). HEs are highly specific DNA-cleaving enzymes that recognize long stretches of DNA and are powerful tools for genome engineering. Because of its monomeric nature, I-DmoI is an ideal scaffold for generating mutant enzymes with novel DNA specificities, similarly reported for homodimeric HEs, but providing single chain endonucleases instead of dimers. However, this would require the use of a mesophilic variant cleaving its substrate at temperatures of 37 degrees C and below. We have generated mesophilic mutants of I-DmoI, using a single round of directed evolution that relies on a functional assay in yeast. The effect of mutations identified in the novel proteins has been investigated. These mutations are located distant to the DNA-binding site and cause changes in the size and polarity of buried residues, suggesting that they act by destabilizing the protein. Two of the novel proteins have been produced and analyzed in vitro. Their overall structures are similar to that of the parent protein, but they are destabilized against thermal and chemical denaturation. The temperature-dependent activity profiles for the mutants shifted toward lower temperatures with respect to the wild-type activity profile. However, the most destabilized mutant was not the most active at low temperatures, suggesting that other effects, like local structural distortions and/or changes in the protein dynamics, also influence their activity. These mesophilic I-DmoI mutants form the basis for generating new variants with tailored DNA specificities.

PaLS: filtering common literature, biological terms and pathway information

Andreu Alibes — 2008-05-08T13:58:16-00:00

Nucl. Acids Res. (8 May 2008), gkn251.

Many biological experiments and their subsequent analysis yield lists of genes or proteins that can potentially be important to the prognosis or diagnosis of certain diseases (e.g. cancer). Nowadays, information about the function of those genes or proteins may be already gathered in some databases, but it is essential to understand if some of the members of those lists have a function in common or if they belong to the same metabolic pathway. To help researchers filter those genes or proteins that have such information in common, we have developed PaLS (pathway and literature strainer, http://pals.bioinfo.cnio.es). PaLS takes a list or a set of lists of gene or protein identifiers and shows which ones share certain descriptors. Four publicly available databases have been used for this purpose: PubMed, which links genes with those articles that make reference to them; Gene Ontology, an annotated ontology of terms related to the cellular component, biological process or molecular function where those genes or proteins are involved; KEGG pathways and Reactome pathways. Those descriptors among these four sources of information that are shared by more members of the list (or lists) are highlighted by PaLS. 10.1093/nar/gkn251

coliSNP database server mapping nsSNPs on protein structures.

H Kono — 2008-05-08T13:40:35-00:00

Nucleic acids research, Vol. 36, No. Database issue. (January 2008)

We have developed coliSNP, a database server (http://yayoi.kansai.jaea.go.jp/colisnp) that maps non-synonymous single nucleotide polymorphisms (nsSNPs) on the three-dimensional (3D) structure of proteins. Once a week, the SNP data from the dbSNP database and the protein structure data from the Protein Data Bank (PDB) are downloaded, and the correspondence of the two data sets is automatically tabulated in the coliSNP database. Given an amino acid sequence, protein name or PDB ID, the server will immediately provide known nsSNP information, including the amino acid mutation caused by the nsSNP, the solvent accessibility, the secondary structure and the flanking residues of the mutated residue in a single page. The position of the nsSNP within the amino acid sequence and on the 3D structure of the protein can also be observed. The database provides key information with which to judge whether an observed nsSNP critically affects protein function and/or stability. As far as we know, this is the only web-based nsSNP database that automatically compiles SNP and protein information in a concise manner.

nuScore: a web-interface for nucleosome positioning predictions

Michael Tolstorukov — 2008-05-05T10:28:39-00:00

Bioinformatics (29 April 2008), btn212.

Summary: Sequence-directed mapping of nucleosome positions is of major biological interest. Here, we present a web-interface for estimation of the affinity of the histone core to DNA and prediction of nucleosome arrangement on a given sequence. Our approach is based on assessment of the energy cost of imposing the deformations required to wrap DNA around the histone surface. The interface allows the user to specify a number of options such as selecting from several structural templates for threading calculations and adding random sequences to the analysis. Availability: The nuScore interface is freely available for use at http://compbio.med.harvard.edu/nuScore. Supplementary information: The site contains user manual, description of the methodology, and examples. Contact: peter_park@harvard.edu; tolstorukov@gmail.com 10.1093/bioinformatics/btn212

Evolvability and hierarchy in rewired bacterial gene networks

Mark Isalan — 2008-04-16T19:45:06-00:00

Nature, Vol. 452, No. 7189. (17 April 2008), pp. 840-845.

FatiGO: a web tool for finding significant associations of Gene Ontology terms with groups of genes

Fatima Al-Shahrour — 2006-06-09T18:22:46-00:00

Bioinformatics, Vol. 20, No. 4. (1 March 2004), pp. 578-580.

Summary: We present a simple but powerful procedure to extract Gene Ontology (GO) terms that are significantly over- or under-represented in sets of genes within the context of a genome-scale experiment (DNA microarray, proteomics, etc.). Said procedure has been implemented as a web application, FatiGO, allowing for easy and interactive querying. FatiGO, which takes the multiple-testing nature of statistical contrast into account, currently includes GO associations for diverse organisms (human, mouse, fly, worm and yeast) and the TrEMBL/Swissprot GOAnnotations@EBI correspondences from the European Bioinformatics Institute. Availability: http://fatigo.bioinfo.cnio.es 10.1093/bioinformatics/btg455

Theoretical and experimental approaches to understand morphogen gradients

Marta Ibanes — 2008-03-26T20:10:23-00:00

Mol Syst Biol, Vol. 4 (25 March 2008)

Dynamic Regulation of Nucleosome Positioning in the Human Genome

Dustin Schones — 2008-03-07T11:55:52-00:00

Cell, Vol. 132, No. 5. (7 March 2008), pp. 887-898.

Summary The positioning of nucleosomes with respect to DNA plays an important role in regulating transcription. However, nucleosome mapping has been performed for only limited genomic regions in humans. We have generated genome-wide maps of nucleosome positions in both resting and activated human CD4+ T cells by direct sequencing of nucleosome ends using the Solexa high-throughput sequencing technique. We find that nucleosome phasing relative to the transcription start sites is directly correlated to RNA polymerase II (Pol II) binding. Furthermore, the first nucleosome downstream of a start site exhibits differential positioning in active and silent genes. TCR signaling induces extensive nucleosome reorganization in promoters and enhancers to allow transcriptional activation or repression. Our results suggest that H2A.Z-containing and modified nucleosomes are preferentially lost from the -1 nucleosome position. Our data provide a comprehensive view of the nucleosome landscape and its dynamic regulation in the human genome.

Structure-based prediction of transcription factor binding sites using a protein-DNA docking approach

Zhijie Liu — 2008-03-05T01:14:47-00:00

Proteins: Structure, Function, and Bioinformatics, Vol. 9999, No. 9999. (2008), NA.

Accurate identification of transcription factor binding sites is critical to our understanding of transcriptional regulatory networks. To overcome the issue of high false-positive predictions that trouble the sequence-based prediction techniques, we have developed a structure-based prediction method that takes into consideration of interactions between the amino acids of a transcription factor and the nucleotides of its DNA binding sequence at structural level, along with an efficient protein-DNA docking algorithm. The docked structures between a protein and a DNA are evaluated using a knowledge-based energy function, in conjunction with van der Waals energy. Our docking algorithm supports quasi-flexible docking, overcoming a number of limiting issues faced by similar docking algorithms. Our rigid-body docking algorithm is tested on a dataset of 141 nonredundant transcription factor-DNA complex structures. The test results show that 63.1% of the 141 complex structures are reconstructed with accuracies better than 1.0 Å RMSDs (root mean square deviation) and 79.4% of the complexes are predicted with accuracies better than 3.0 Å RMSDs when using the native DNA structures. Our quasi-flexible docking algorithm, assuming that the DNA structures are not known, is tested on a separate set of 45 transcription factor-DNA complexes, of which 57.8% of the docked complex conformations achieve better than 1.0 Å RMSDs while 71.1% of the complexes have RMSDs less than 3.0 Å. We have also applied our method to predict the binding motifs of the ferric uptake regulator in E. coli and showed that most of the experimentally identified sites can be predicted accurately. Proteins 2008. © 2008 Wiley-Liss, Inc.

De Novo Computational Design of Retro-Aldol Enzymes

Lin Jiang — 2008-03-07T02:58:37-00:00

Science, Vol. 319, No. 5868. (7 March 2008), pp. 1387-1391.

The creation of enzymes capable of catalyzing any desired chemical reaction is a grand challenge for computational protein design. Using new algorithms that rely on hashing techniques to construct active sites for multistep reactions, we designed retro-aldolases that use four different catalytic motifs to catalyze the breaking of a carbon-carbon bond in a nonnatural substrate. Of the 72 designs that were experimentally characterized, 32, spanning a range of protein folds, had detectable retro-aldolase activity. Designs that used an explicit water molecule to mediate proton shuffling were significantly more successful, with rate accelerations of up to four orders of magnitude and multiple turnovers, than those involving charged side-chain networks. The atomic accuracy of the design process was confirmed by the x-ray crystal structure of active designs embedded in two protein scaffolds, both of which were nearly superimposable on the design model. 10.1126/science.1152692

GroEL stimulates protein folding through forced unfolding

Zong Lin — 2008-03-06T04:15:38-00:00

Nature Structural & Molecular Biology, Vol. 15, No. 3. (02 March 2008), pp. 303-311.

Functional protein divergence in the evolution of Homo sapiens

Nuria Lopez-Bigas — 2008-02-15T13:27:09-00:00

Genome Biology, Vol. 9 (15 February 2008), R33.

Meganucleases and DNA double-strand break-induced recombination: perspectives for gene therapy.

F Pâques — 2008-02-22T16:03:27-00:00

Curr Gene Ther, Vol. 7, No. 1. (February 2007), pp. 49-66.

Meganucleases are sequence-specific endonucleases recognizing large (>12 bp) sequence sites and several laboratories have used these proteins to induce highly efficient gene targeting in mammalian cells. The recent development of artificial endonucleases with tailored specificities has opened the door for a wide range of new applications, including therapeutic ones: redesigned endonucleases cleaving chosen sequences could be used to in gene therapy to correct mutated genes or introduce transgenes in chosen loci. Such "targeted" approaches markedly differ from current gene therapy strategies based on the random insertion of a complementing virus-borne transgene. As a consequence, they should bypass the odds of random insertion. Artificial fusion proteins including Zinc-Finger binding domains have provided important proofs of concept, however the toxicity of these proteins is still an issue. Today custom-designed homing endonucleases, the natural meganucleases, could represent an efficient alternative. After a brief description of the origin of the technology, current systems based on redesigned endonucleases will be presented, with a special emphasis on the recent advances in homing endonuclease engineering. Finally, we will discuss the main issues that will need to be addressed in order to bring this promising technology to the patient.

Protein-nucleic acid complexes: large, small, old, and new

Gregory Van Duyne — 2008-02-20T15:13:28-00:00

Current Opinion in Structural Biology, Vol. 18, No. 1. (February 2008), pp. 67-69.

Computer design of obligate heterodimer meganucleases allows efficient cutting of custom DNA sequences

Emmanuel Fajardo-Sanchez — 2008-02-18T11:53:59-00:00

Nucl. Acids Res. (14 February 2008), gkn059.

Meganucleases cut long (>12 bp) unique sequences in genomes and can be used to induce targeted genome engineering by homologous recombination in the vicinity of their cleavage site. However, the use of natural meganucleases is limited by the repertoire of their target sequences, and considerable efforts have been made to engineer redesigned meganucleases cleaving chosen targets. Homodimeric meganucleases such as I-CreI have provided a scaffold, but can only be modified to recognize new quasi-palindromic DNA sequences, limiting their general applicability. Other groups have used dimer-interface redesign and peptide linkage to control heterodimerization between related meganucleases such as I-DmoI and I-CreI, but until now there has been no application of this aimed specifically at the scaffolds from existing combinatorial libraries of I-CreI. Here, we show that engineering meganucleases to form obligate heterodimers results in functional endonucleases that cut non-palindromic sequences. The protein design algorithm (FoldX v2.7) was used to design specific heterodimer interfaces between two meganuclease monomers, which were themselves engineered to recognize different DNA sequences. The new monomers favour functional heterodimer formation and prevent homodimer site recognition. This design massively increases the potential repertoire of DNA sequences that can be specifically targeted by designed I-CreI meganucleases and opens the way to safer targeted genome engineering. 10.1093/nar/gkn059

Genomic Sequence Is Highly Predictive of Local Nucleosome Depletion

Guo-Cheng Yuan — 2008-01-28T11:37:59-00:00

PLoS Computational Biology, Vol. 4, No. 1. (1 January 2008), e13.

The regulation of DNA accessibility through nucleosome positioning is important for transcription control. Computational models have been developed to predict genome-wide nucleosome positions from DNA sequences, but these models consider only nucleosome sequences, which may have limited their power. We developed a statistical multi-resolution approach to identify a sequence signature, called the N-score, that distinguishes nucleosome binding DNA from non-nucleosome DNA. This new approach has significantly improved the prediction accuracy. The sequence information is highly predictive for local nucleosome enrichment or depletion, whereas predictions of the exact positions are only modestly more accurate than a null model, suggesting the importance of other regulatory factors in fine-tuning the nucleosome positions. The N-score in promoter regions is negatively correlated with gene expression levels. Regulatory elements are enriched in low N-score regions. While our model is derived from yeast data, the N-score pattern computed from this model agrees well with recent high-resolution protein-binding data in human.

dbSNP: the NCBI database of genetic variation.

ST Sherry — 2006-08-10T16:43:06-00:00

Nucleic Acids Res, Vol. 29, No. 1. (1 January 2001), pp. 308-311.

In response to a need for a general catalog of genome variation to address the large-scale sampling designs required by association studies, gene mapping and evolutionary biology, the National Center for Biotechnology Information (NCBI) has established the dbSNP database [S.T.Sherry, M.Ward and K.Sirotkin (1999) Genome Res., 9, 677-679]. Submissions to dbSNP will be integrated with other sources of information at NCBI such as GenBank, PubMed, LocusLink and the Human Genome Project data. The complete contents of dbSNP are available to the public at website: http://www.ncbi.nlm.nih.gov/SNP. The complete contents of dbSNP can also be downloaded in multiple formats via anonymous FTP at ftp://ncbi.nlm.nih.gov/snp/.

High-throughput mapping of the chromatin structure of human promoters

Fatih Ozsolak — 2007-02-08T10:59:37-00:00

Nat Biotech, Vol. 25, No. 2. (February 2007), pp. 244-248.

The Histone Database: a comprehensive resource for histones and histone fold-containing proteins.

L Mariño-Ramírez — 2008-01-24T11:24:48-00:00

Proteins, Vol. 62, No. 4. (1 March 2006), pp. 838-842.

The Histone Database is a curated and searchable collection of full-length sequences and structures of histones and nonhistone proteins containing histone-like folds, compiled from major public databases. Several new histone fold-containing proteins have been identified, including the huntingtin-interacting protein HYPM. Additionally, based on the recent crystal structure of the Son of Sevenless protein, an interpretation of the sequence analysis of the histone fold domain is presented. The database contains an updated collection of multiple sequence alignments for the four core histones (H2A, H2B, H3, and H4) and the linker histones (H1/H5) from a total of 975 organisms. The database also contains information on the human histone gene complement and provides links to three-dimensional structures of histone and histone fold-containing proteins. The Histone Database is a comprehensive bioinformatics resource for the study of structure and function of histones and histone fold-containing proteins. The database is available at http://research.nhgri.nih.gov/histones/.

A high-resolution atlas of nucleosome occupancy in yeast

William Lee — 2007-10-01T04:57:32-00:00

Nature Genetics, Vol. 39, No. 10. (16 September 2007), pp. 1235-1244.

GeneLibrarian: an effective gene-information summarization and visualization system

Jung-Hsien Chiang — 2006-08-29T23:45:30-00:00

BMC Bioinformatics, Vol. 7 (29 August 2006), 392.