CiteULike: barry's library [699 articles]

Progress in Modeling of Protein Structures and Interactions

Ora Schueler-Furman — 2005-10-27T23:14:15-00:00

Science, Vol. 310, No. 5748. (28 October 2005), pp. 638-642.

The prediction of the structures and interactions of biological macromolecules at the atomic level and the design of new structures and interactions are critical tests of our understanding of the interatomic interactions that underlie molecular biology. Equally important, the capability to accurately predict and design macromolecular structures and interactions would streamline the interpretation of genome sequence information and allow the creation of macromolecules with new and useful functions. This review summarizes recent progress in modeling that suggests that we are entering an era in which high-resolution prediction and design will make increasingly important contributions to biology and medicine.

Cargo transport: molecular motors navigate a complex cytoskeleton

Jennifer Ross — 2008-05-30T09:07:53-00:00

Current Opinion in Cell Biology, Vol. 20, No. 1. (February 2008), pp. 41-47.

Intracellular cargo transport requires microtubule-based motors, kinesin and cytoplasmic dynein, and the actin-based myosin motors to maneuver through the challenges presented by the filamentous meshwork that comprises the cytoskeleton. Recent in vitro single molecule biophysical studies have begun to explore this process by characterizing what occurs as these tiny molecular motors happen upon an intersection between two cytoskeletal filaments. These studies, in combination with in vivo work, define the mechanism by which molecular motors exchange cargo while traveling between filamentous tracks and deliver it to its destination when going from the cell center to the periphery and back again.

Ras effectors

Christopher Marshall — 2008-07-18T18:34:28-00:00

Current Opinion in Cell Biology, Vol. 8, No. 2. (April 1996), pp. 197-204.

The search for proteins which interact with the active GTP-bound form of Ras in order to transmit signals for proliferation, differentiation and oncogenesis has been a long one. Now there are several strong candidates for Ras effectors that include protein kinases, lipid kinases and guanine nucleotide exchange factors. Structural information on how one Ras-binding domain in an effector interacts with Ras-GTP has recently been obtained. Recent data show that transformation by Ras oncoproteins requires the activation of several signal transduction pathways, including those which transmit signals via members of the Rho family of GTPases.

Farnesyltransferase inhibitors: antineoplastic mechanism and clinical prospects

George Prendergast — 2008-07-18T18:29:21-00:00

Current Opinion in Cell Biology, Vol. 12, No. 2. (1 April 2000), pp. 166-173.

Recent work suggests that farnesyltransferase inhibitors suppress cancer cell proliferation through mechanisms other than inhibiting Ras isoprenylation, which is not a crucial event. Recent evidence also suggests that the antineoplastic properties of farnesyltransferase inhibitors are due to alterations in the isoprenylation of RhoB, an endosomal Rho protein that functions in receptor trafficking. A shift in conceptual focus from Ras to Rho to understand how farnesyltransferase inhibitors act provides a new vantage to address old questions in the field and suggests strategies to improve and potentially widen clinical applications.

The Ras branch of small Gtpases: Ras family members don't fall far from the tree

Gary Reuther — 2008-07-18T18:27:16-00:00

Current Opinion in Cell Biology, Vol. 12, No. 2. (1 April 2000), pp. 157-165.

The Ras branch of the Ras superfamily consists of small GTPases most closely related to Ras and include the R-Ras, Rap, Ral, Rheb, Rin and Rit proteins. Although our understanding of Ras signaling and biology is now considerable, recent observations suggest that Ras function is more complex than previously believed. First, the three Ras proteins may not be functionally identical. Second, Ras function involves functional cross-talk with their close relatives.

The importance of stupidity in scientific research

Martin Schwartz — 2008-07-16T21:24:00-00:00

J Cell Sci, Vol. 121, No. 11. (1 June 2008), 1771.

10.1242/jcs.033340

Ras oncogenes: split personalities

Antoine Karnoub — 2008-06-23T17:03:33-00:00

Nat Rev Mol Cell Biol, Vol. 9, No. 7. (July 2008), pp. 517-531.

Protein-protein binding is often associated with changes in protonation state

Aaron Mason — 2007-10-12T12:41:41-00:00

Proteins: Structure, Function, and Bioinformatics, Vol. 9999, No. 9999. (2007), NA.

pKa values of ionizable residues have been calculated using the PROPKA method and structures of 75 protein-protein complexes and their corresponding free forms. These pKa values were used to compute changes in protonation state of individual residues, net changes in protonation state of the complex relative to the uncomplexed proteins, and the correction to a binding energy calculated assuming standard protonation states at pH 7. For each complex, two different structures for the uncomplexed form of the proteins were used: the X-ray structures determined for the proteins in the absence of the other protein and the individual protein structures taken from the structure of the complex (referred to as unbound and bound structures, respectively). In 28 and 77% of the cases considered here, protein-protein binding is accompanied by a complete (>95%) or significant (>50%) change in protonation state of at least one residue using unbound structures. Furthermore, in 36 and 61% of the cases, protein-protein binding is accompanied by a complete or significant net change in protonation state of the complex relative to the separated monomers. Using bound structures, the corresponding values are 12, 51, 20, and 48%. Comparison to experimental data suggest that using unbound and bound structures lead to over- and underestimation of binding-induced protonation state changes, respectively. Thus, we conclude that protein-protein binding is often associated with changes in protonation state of amino acid residues and with changes in the net protonation state of the proteins. The pH-dependent correction to the binding energy contributes at least one order of magnitude to the binding constant in 45 and 23%, using unbound and bound structures, respectively. Proteins 2007. © 2007 Wiley-Liss, Inc.

Avian influenza virus, a very sticky situation

Mark von Itzstein — 2008-04-04T00:38:45-00:00

Current Opinion in Chemical Biology, Vol. 12, No. 1. (February 2008), pp. 102-108.

The appearance of the highly pathogenic avian influenza virus H5N1 highlighted the potential impact of influenza virus on humanity. The emergence of this high profile virus stimulated much research towards a better understanding of the key determinants for successful human-to-human transmission and as such has provided new directions for therapeutic intervention strategies. For example, a phylogenetic-based grouping of influenza virus sialidases into either Group 1 or 2 has been proposed. This has provided new opportunity for the development of Group 1-specific anti-influenza drugs. Furthermore, a number of next generation sialidase inhibitors as anti-influenza drugs have also been developed.

X-ray structure of a prokaryotic pentameric ligand-gated ion channel

Ricarda Hilf — 2008-03-06T04:09:10-00:00

Nature (05 March 2008)

Association of Aminoglycosidic Antibiotics with the Ribosomal A-Site Studied with Brownian Dynamics

Maciej Długosz — 2008-05-12T11:07:58-00:00

J. Chem. Theory Comput., Vol. 4, No. 4. (8 April 2008), pp. 549-559.

Abstract: Brownian dynamics methodology was applied to simulate the encounter of aminoglycosidic antibiotics with the ribosomal A-site RNA. Studied antibiotics included neamine, neomycin, ribostamycin, and paromomycin which differ in chemical structure, the number of pseudosugar rings, and the net charge. The influence of structural, electrostatic, and hydrodynamic properties of antibiotics on the kinetics of their association with the ribosomal A-site was analyzed. The computed diffusion limited rates of association are of the order of 1010 1/M·s, and they weakly depend on ionic strength. Prior to binding, antibiotics often slide along the RNA groove with the time scale of approximately 10 ns per base pair in the case of neamine. We observed that upon forming the encounter complex aminoglycosides displace magnesium ions from the binding pocket.

Accelerated Superposition State Molecular Dynamics for Condensed Phase Systems

Michele Ceotto — 2008-07-08T00:20:59-00:00

J. Chem. Theory Comput., Vol. 4, No. 4. (8 April 2008), pp. 560-568.

Abstract: An extension of superposition state molecular dynamics (SSMD) [Venkatnathan and Voth J. Chem. Theory Comput. 2005, 1, 36] is presented with the goal to accelerate timescales and enable the study of long-time phenomena for condensed phase systems. It does not require any a priori knowledge about final and transition state configurations, or specific topologies. The system is induced to explore new configurations by virtue of a fictitious (free-particle-like) accelerating potential. The acceleration method can be applied to all degrees of freedom in the system and can be applied to condensed phases and fluids.

How Efficient Is Replica Exchange Molecular Dynamics? An Analytic Approach

Hugh Nymeyer — 2008-04-09T12:22:40-00:00

J. Chem. Theory Comput., Vol. 4, No. 4. (8 April 2008), pp. 626-636.

Abstract: Replica exchange molecular dynamics (REMD) has become a standard technique for accelerating relaxation in biosimulations. Despite its widespread use, questions remain about its efficiency compared with conventional, constant temperature molecular dynamics (MD). An analytic approach is taken to describe the relative efficiency of REMD with respect to MD. This is applied to several simple two-state models and to several real proteinsprotein L and the B domain of protein Ato predict the relative efficiency of REMD with respect to MD in actual applications. In agreement with others, we find the following: as long as there is a positive activation energy for folding, REMD is more efficient than MD; the effectiveness of REMD is strongly dependent on the activation enthalpy; and the efficiency of REMD for actual proteins is a strong function of the maximum temperature. Choosing the maximum temperature too high can result in REMD becoming significantly less efficient than conventional MD. A good rule of thumb appears to be to choose the maximum temperature of the REMD simulation slightly above the temperature at which the enthalpy for folding vanishes. Additionally, we find that the number of replicas in REMD, while important for simulations shorter than one or two relaxation times, has a minimal effect on the asymptotic efficiency of the method.

Protonation free energy levels in complex molecular systems

Jan Antosiewicz — 2007-09-06T12:15:09-00:00

Biopolymers, Vol. 9999, No. 9999. (2007), NA.

All proteins, nucleic acids and other biomolecules contain residues capable of exchanging protons with their environment. These proton transfer phenomena lead to pH sensitivity of many molecular processes underlying biological phenomena. In the course of biological evolution, Nature has invented some mechanisms to use pH gradients to regulate biomolecular processes inside cells or in interstitial fluids. Therefore, an ability to model protonation equilibria in molecular systems accurately would be of enormous value for our understanding of biological processes and for possible rational influence on them, like in developing pH dependent drugs to treat particular diseases. © 2007 Wiley Periodicals, Inc. Biopolymers, 2007

Novel Method for Probing the Specificity Binding Profile of Ligands: Applications to HIV Protease

Woody Sherman — 2008-05-06T21:33:12-00:00

Chemical Biology & Drug Design, Vol. 71, No. 5. (2008), pp. 387-407.

A detailed understanding of factors influencing the binding specificity of a ligand to a set of desirable targets and undesirable decoys is a key step in the design of potent and selective therapeutics. We have developed a general method for optimizing binding specificity in ligand-receptor complexes based on the theory of electrostatic charge optimization. This methodology can be used to tune the binding of a ligand to a panel of potential targets and decoys, along the continuum from narrow binding to only one partner to broad binding to the entire panel. Using HIV-1 protease as a model system, we probe specificity in three distinct ways. First, we probe interactions that could make the promiscuous protease inhibitor pepstatin more selective toward HIV-1 protease. Next, we study clinically approved HIV-1 protease inhibitors and probe ways to broaden the binding profiles toward both wild-type HIV-1 protease and drug-resistant mutants. Finally, we study a conformational ensemble of wild-type HIV-1 protease to ‘design in’ broad specificity to known drugs before resistance mutations arise. The results from this conformational ensemble were similar to those from the drug-resistant ensemble, suggesting the use of a conformational wild-type ensemble as a tool to develop escape-mutant-resistant inhibitors.

Calculation of protein-ligand binding free energy by using a polarizable potential

Dian Jiao — 2008-04-29T17:41:01-00:00

Proceedings of the National Academy of Sciences, Vol. 105, No. 17. (29 April 2008), pp. 6290-6295.

The binding of charged ligands benzamidine and diazamidine to trypsin was investigated by using a polarizable potential energy function and explicit-water molecular dynamics simulations. The binding free energies were computed from the difference between the free energies of decoupling the ligand from water and protein environments. Both the absolute and the relative free energies from the perturbation simulations agree with experimental measurements to within 0.5 kcalmiddle dotmol-1. Comparison of free-energy components sampled from different thermodynamic paths indicates that electrostatics is the main driving force behind benzamidine recognition of trypsin. The contribution of electronic polarization to binding appears to be crucial. By computing the free-energy contribution caused by the polarization between the ligand and its surroundings, we found that polarization has the opposite effect in dissimilar environments. Although polarization favors ligand solvation in water, it weakens the protein-ligand attraction by screening the electrostatic interaction between trypsin and benzamidine. We also examined the relative binding free energies of a benzamidine analog diazamidine to trypsin. The changes in free energy on benzamidine-diazamidine substitution were tens of kilocalories in both water and trypsin environments; however, the change in the total binding free energy is <2 kcalmiddle dotmol-1 because of cancellation, consistent with the experimental results. Overall, our results suggest that the use of a polarizable force field, given adequate sampling, is capable of achieving chemical accuracy in molecular simulations of protein-ligand recognition. 10.1073/pnas.0711686105

From the Cover: A dry ligand-binding cavity in a solvated protein

Johan Qvist — 2008-05-26T20:32:13-00:00

Proceedings of the National Academy of Sciences, Vol. 105, No. 17. (29 April 2008), pp. 6296-6301.

Ligands usually bind to proteins by displacing water from the binding site. The affinity and kinetics of binding therefore depend on the hydration characteristics of the site. Here, we show that the extreme case of a completely dehydrated free binding site is realized for the large nonpolar binding cavity in bovine -lactoglobulin. Because spatially delocalized water molecules may escape detection by x-ray diffraction, we use water 17O and 2H magnetic relaxation dispersion (MRD), 13C NMR spectroscopy, molecular dynamics simulations, and free energy calculations to establish the absence of water from the binding cavity. Whereas carbon nanotubes of the same diameter are filled by a hydrogen-bonded water chain, the MRD data show that the binding pore in the apo protein is either empty or contains water molecules with subnanosecond residence times. However, the latter possibility is ruled out by the computed hydration free energies, so we conclude that the 315 A3 binding pore is completely empty. The apo protein is thus poised for efficient binding of fatty acids and other nonpolar ligands. The qualitatively different hydration of the -lactoglobulin pore and carbon nanotubes is caused by subtle differences in water-wall interactions and water entropy. 10.1073/pnas.0709844105

Recognition Dynamics Up to Microseconds Revealed from an RDC-Derived Ubiquitin Ensemble in Solution

Oliver Lange — 2008-06-13T00:26:27-00:00

Science, Vol. 320, No. 5882. (13 June 2008), pp. 1471-1475.

Protein dynamics are essential for protein function, and yet it has been challenging to access the underlying atomic motions in solution on nanosecond-to-microsecond time scales. We present a structural ensemble of ubiquitin, refined against residual dipolar couplings (RDCs), comprising solution dynamics up to microseconds. The ensemble covers the complete structural heterogeneity observed in 46 ubiquitin crystal structures, most of which are complexes with other proteins. Conformational selection, rather than induced-fit motion, thus suffices to explain the molecular recognition dynamics of ubiquitin. Marked correlations are seen between the flexibility of the ensemble and contacts formed in ubiquitin complexes. A large part of the solution dynamics is concentrated in one concerted mode, which accounts for most of ubiquitin's molecular recognition heterogeneity and ensures a low entropic complex formation cost. 10.1126/science.1157092

The Poisson-Boltzmann model for tRNA: Assessment of the calculation set-up and ionic concentration cutoff

Magdalena Gruziel — 2008-07-07T23:51:18-00:00

Journal of Computational Chemistry, Vol. 29, No. 12. (2008), pp. 1970-1981.

Using tRNA molecule as an example, we evaluate the applicability of the Poisson-Boltzmann model to highly charged systems such as nucleic acids. Particularly, we describe the effect of explicit crystallographic divalent ions and water molecules, ionic strength of the solvent, and the linear approximation to the Poisson-Boltzmann equation on the electrostatic potential and electrostatic free energy. We calculate and compare typical similarity indices and measures, such as Hodgkin index and root mean square deviation. Finally, we introduce a modification to the nonlinear Poisson-Boltzmann equation, which accounts in a simple way for the finite size of mobile ions, by applying a cutoff in the concentration formula for ionic distribution at regions of high electrostatic potentials. We test the influence of this ionic concentration cutoff on the electrostatic properties of tRNA. © 2008 Wiley Periodicals, Inc. J Comput Chem, 2008

Protein model refinement using an optimized physics-based all-atom force field

Anna Jagielska — 2008-06-19T11:23:04-00:00

Proceedings of the National Academy of Sciences, Vol. 105, No. 24. (17 June 2008), pp. 8268-8273.

One of the greatest challenges in protein structure prediction is the refinement of low-resolution predicted models to high-resolution structures that are close to the native state. Although contemporary structure prediction methods can assemble the correct topology for a large fraction of protein domains, such approximate models are often not of the resolution required for many important applications, including studies of reaction mechanisms and virtual ligand screening. Thus, the development of a method that could bring those structures closer to the native state is of great importance. We recently optimized the relative weights of the components of the Amber ff03 potential on a large set of decoy structures to create a funnel-shaped energy landscape with the native structure at the global minimum. Such an energy function might be able to drive proteins toward their native structure. In this work, for a test set of 47 proteins, with 100 decoy structures per protein that have a range of structural similarities to the native state, we demonstrate that our optimized potential can drive protein models closer to their native structure. Comparing the lowest-energy structure from each trajectory with the starting decoy, structural improvement is seen for 70% of the models on average. The ability to do such systematic structural refinements by using a physics-based all-atom potential represents a promising approach to high-resolution structure prediction. 10.1073/pnas.0800054105

Pathway and Endpoint Free Energy Calculations for Cyclic Nucleotide Binding to HCN Channels

Lei Zhou — 2008-07-02T23:57:15-00:00

Biophys. J., Vol. 94, No. 12. (15 June 2008), pp. L90-92.

cAMP and cGMP differentially bind to and regulate a variety of proteins, including cyclic nucleotide-gated (CNG) channels and hyperpolarization-activated cyclic nucleotide-regulated (HCN) channels. Previous site-directed mutagenesis studies have isolated two conserved residues that are critical for enabling certain channels to selectively bind cGMP relative to cAMP. However, no definitive mechanism has been identified that explains the preferential activation of other channels by cAMP. Here we apply computational binding free energy methods, including thermodynamic integration, linear interaction energy, and continuum electrostatic calculations, to gain insights into the mechanisms of cyclic nucleotide selectivity. Consistent with experimental observations, computational results for the cAMP-selective HCN channels show that the binding free energy of cAMP is lower (more favorable) than that of cGMP. Surprisingly, cAMP selectivity is not due to its preferential contacts with protein, but rather reflects the greater hydration energy of cGMP relative to cAMP, resulting in a greater energetic cost for cGMP binding. 10.1529/biophysj.108.130872

BIOCHEMISTRY: Metamorphic Proteins

Alexey Murzin — 2008-06-27T06:23:05-00:00

Science, Vol. 320, No. 5884. (27 June 2008), pp. 1725-1726.

10.1126/science.1158868

The GAP arginine finger movement into the catalytic site of Ras increases the activation entropy

Carsten Kotting — 2008-05-05T20:25:27-00:00

Proceedings of the National Academy of Sciences, Vol. 105, No. 17. (29 April 2008), pp. 6260-6265.

Members of the Ras superfamily of small G proteins play key roles in signal transduction pathways, which they control by GTP hydrolysis. They are regulated by GTPase activating proteins (GAPs). Mutations that prevent hydrolysis cause severe diseases including cancer. A highly conserved "arginine finger" of GAP is a key residue. Here, we monitor the GTPase reaction of the Rasmiddle dotRasGAP complex at high temporal and spatial resolution by time-resolved FTIR spectroscopy at 260 K. After triggering the reaction, we observe as the first step a movement of the switch-I region of Ras from the nonsignaling "off" to the signaling "on" state with a rate of 3 s-1. The next step is the movement of the "arginine finger" into the active site of Ras with a rate of k2 = 0.8 s-1. Once the arginine points into the binding pocket, cleavage of GTP is fast and the protein-bound Pi intermediate forms. The switch-I reversal to the "off" state, the release of Pi, and the movement of arginine back into an aqueous environment is observed simultaneously with k3 = 0.1 s-1, the rate-limiting step. Arrhenius plots for the partial reactions show that the activation energy for the cleavage reaction is lowered by favorable positive activation entropy. This seems to indicate that protein-bound structured water molecules are pushed by the "arginine finger" movement out of the binding pocket into the bulk water. The proposed mechanism shows how the high activation barrier for phosphoryl transfer can be reduced by splitting into partial reactions separated by a Pi-intermediate. 10.1073/pnas.0712095105

A Simple Model of Backbone Flexibility Improves Modeling of Side-chain Conformational Variability

Gregory Friedland — 2008-06-30T16:35:27-00:00

Journal of Molecular Biology, Vol. 380, No. 4. (18 July 2008), pp. 757-774.

The considerable flexibility of side-chains in folded proteins is important for protein stability and function, and may have a role in mediating allosteric interactions. While sampling side-chain degrees of freedom has been an integral part of several successful computational protein design methods, the predictions of these approaches have not been directly compared to experimental measurements of side-chain motional amplitudes. In addition, protein design methods frequently keep the backbone fixed, an approximation that may substantially limit the ability to accurately model side-chain flexibility. Here, we describe a Monte Carlo approach to modeling side-chain conformational variability and validate our method against a large dataset of methyl relaxation order parameters derived from nuclear magnetic resonance (NMR) experiments (17 proteins and a total of 530 data points). We also evaluate a model of backbone flexibility based on Backrub motions, a type of conformational change frequently observed in ultra-high-resolution X-ray structures that accounts for correlated side-chain backbone movements. The fixed-backbone model performs reasonably well with an overall rmsd between computed and predicted side-chain order parameters of 0.26. Notably, including backbone flexibility leads to significant improvements in modeling side-chain order parameters for ten of the 17 proteins in the set. Greater accuracy of the flexible backbone model results from both increases and decreases in side-chain flexibility relative to the fixed-backbone model. This simple flexible-backbone model should be useful for a variety of protein design applications, including improved modeling of protein-protein interactions, design of proteins with desired flexibility or rigidity, and prediction of correlated motions within proteins.

The 2006 Automated Function Prediction Meeting

Ana Rodrigues — 2008-06-26T19:06:23-00:00

BMC Bioinformatics, Vol. 8, No. Suppl 4. (2007)

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Do's and don't's of poster presentation.

SM Block — 2008-06-24T17:18:41-00:00

Biophysical journal, Vol. 71, No. 6. (December 1996), pp. 3527-3529.

Mortal Sins in Poster Presentations

Thomas Wolcott — 2007-11-30T17:30:20-00:00

SICB Newsletter (Fall 1997)

Poster Presentations as a Genre in Knowledge Communication: A Case Study of Forms, Norms, and Values

Anu Macintosh-Murray — 2008-06-24T17:11:55-00:00

Science Communication, Vol. 28, No. 3. (1 March 2007), pp. 347-376.

Learning to communicate research well through posters involves far more than formatting issues such as font size. The conventions of poster presentations as social practices are part of academic apprenticeship in many health disciplines. This case study examines doctoral students' poster presentations as a research-process genre. The article maps genre knowledge required of novice researchers: poster form, creation processes, presentation practices, and underlying values. Complexity arises from the multiple roles that posters must fulfill, combined with formatting restrictions, the nature of audience interaction, and prestige issues. Posters are often considered as second class compared to oral presentations, perhaps unfairly. The reuse of posters raises questions about publication as academic currency and appropriate knowledge-exchange practices. Poster presentations are evolving with digital media, which may affect competence development in this multimodal form of research communication. Future research should consider how posters' technology-influenced evolution affects interaction, communicative purposes, and the texts themselves. 10.1177/1075547006298251

Principal components analysis of protein structure ensembles calculated using NMR data

Peter Howe — 2008-06-23T21:19:52-00:00

Journal of Biomolecular NMR, Vol. 20, No. 1. (1 May 2001), pp. 61-70.

One important problem when calculating structures of biomolecules from NMR data is distinguishing converged structures from outlier structures. This paper describes how Principal Components Analysis (PCA) has the potential to classify calculated structures automatically, according to correlated structural variation across the population. PCA analysis has the additional advantage that it highlights regions of proteins which are varying across the population. To apply PCA, protein structures have to be reduced in complexity and this paper describes two different representations of protein structures which achieve this. The calculated structures of a 28 amino acid peptide are used to demonstrate the methods. The two different representations of protein structure are shown to give equivalent results, and correct results are obtained even though the ensemble of structures used as an example contains two different protein conformations. The PCA analysis also correctly identifies the structural differences between the two conformations.

RNA dynamics: it is about time

Hashim Al-Hashimi — 2008-06-10T16:09:07-00:00

Current Opinion in Structural Biology, Vol. In Press, Corrected Proof

Many recently discovered RNA functions rely on highly complex multistep conformational transitions that occur in response to an array of cellular signals. These dynamics accompany and guide, for example, RNA cotranscriptional folding, ligand sensing and signaling, site-specific catalysis in ribozymes, and the hierarchically ordered assembly of ribonucleoproteins. RNA dynamics are encoded by both the inherent properties of RNA structure, spanning many motional modes with a large range of amplitudes and timescales, and external trigger factors, ranging from proteins, nucleic acids, metal ions, metabolites, and vitamins to temperature and even directional RNA biosynthesis itself. Here, we review recent advances in our understanding of RNA dynamics as highlighted by biophysical tools.

Mapping the Nucleotide and Isoform-Dependent Structural and Dynamical Features of Ras Proteins

Alemayehu Gorfe — 2008-06-10T21:32:35-00:00

Structure, Vol. 16, No. 6. (11 June 2008), pp. 885-896.

Summary Ras GTPases are conformational switches controlling cell proliferation, differentiation, and development. Despite their prominent role in many forms of cancer, the mechanism of conformational transition between inactive GDP-bound and active GTP-bound states remains unclear. Here we describe a detailed analysis of available experimental structures and molecular dynamics simulations to quantitatively assess the structural and dynamical features of active and inactive states and their interconversion. We demonstrate that GTP-bound and nucleotide-free G12V H-ras sample a wide region of conformational space, and show that the inactive-to-active transition is a multiphase process defined by the relative rearrangement of the two switches and the orientation of Tyr32. We also modeled and simulated N- and K-ras proteins and found that K-ras is more flexible than N- and H-ras. We identified a number of isoform-specific, long-range side chain interactions that define unique pathways of communication between the nucleotide binding site and the C terminus.

Microarray learning with ABC.

Dhammika Amaratunga — 2007-08-02T13:13:29-00:00

Biostatistics (14 June 2007)

Standard clustering algorithms when applied to DNA microarray data often tend to produce erroneous clusters. A major contributor to this divergence is the feature characteristic of microarray data sets that the number of predictors (genes) in such data far exceeds the number of samples by many orders of magnitude, with only a small percentage of predictors being truly informative with regards to the clustering while the rest merely add noise. An additional complication is that the predictors exhibit an unknown complex correlational configuration embedded in a small subspace of the entire predictor space. Under these conditions, standard clustering algorithms fail to find the true clusters even when applied in tandem with some sort of gene filtering or dimension reduction to reduce the number of predictors. We propose, as an alternative, a novel method for unsupervised classification of DNA microarray data. The method, which is based on the idea of aggregating results obtained from an ensemble of randomly resampled data (where both samples and genes are resampled), introduces a way of tilting the procedure so that the ensemble includes minimal representation from less important areas of the gene predictor space. The method produces a measure of dissimilarity between each pair of samples that can be used in conjunction with (a) a method like Ward's procedure to generate a cluster analysis and (b) multidimensional scaling to generate useful visualizations of the data. We call the dissimilarity measures ABC dissimilarities since they are obtained by aggregating bundles of clusters. An extensive comparison of several clustering methods using actual DNA microarray data convincingly demonstrates that classification using ABC dissimilarities offers significantly superior performance.

Structural Basis of the Honey Bee PBP Pheromone and pH-induced Conformational Change

Marion Pesenti — 2008-06-09T16:43:02-00:00

Journal of Molecular Biology, Vol. 380, No. 1. (27 June 2008), pp. 158-169.

The behavior of insects and their perception of their surroundings are driven, in a large part, by odorants and pheromones. This is especially true for social insects, such as the honey bee, where the queen controls the development and the caste status of the other individuals. Pheromone perception is a complex phenomenon relying on a cascade of recognition events, initiated in antennae by pheromone recognition by a pheromone-binding protein and finishing with signal transduction at the axon membrane level. With to the objective of deciphering this initial step, we have determined the structures of the bee antennal pheromone-binding protein (ASP1) in the apo form and in complex with the main component of the queen mandibular pheromonal mixture, 9-keto-2(E)-decenoic acid (9-ODA) and with nonpheromonal components. In the apo protein, the C terminus obstructs the binding site. In contrast, ASP1 complexes have different open conformations, depending on the ligand shape, leading to different volumes of the binding cavity. The binding site integrity depends on the C terminus (111-119) conformation, which involves the interplay of two factors; i.e. the presence of a ligand and a low pH. Ligand binding to ASP1 is favored by low pH, opposite to what is observed with other pheromone-binding proteins, such as those of Bombyx mori and Anopheles gambiae.

Kinesin-8 molecular motors: putting the brakes on chromosome oscillations

Melissa Gardner — 2008-06-02T18:53:57-00:00

Trends in Cell Biology, Vol. In Press, Corrected Proof

Recent studies suggest that the human Kinesin-8 molecular motor Kif18A has a role in chromosome congression. Specifically, these studies find that Kif18A promotes chromosome congression by attenuating chromosome oscillation magnitudes. Together with recent modeling work, in vitro studies, and the analysis of in vivo yeast data, these reports reveal how Kinesin-8 molecular motors might control chromosome oscillation amplitudes by spatially regulating the dynamic instability of microtubule plus-ends within the mitotic spindle.

Structural dynamics of the microtubule binding and regulatory elements in the kinesin-like calmodulin binding protein

Maia Vinogradova — 2008-06-02T18:40:53-00:00

Journal of Structural Biology, Vol. In Press, Corrected Proof

Kinesins are molecular motors that power cell division and transport of various proteins and organelles. Their motor activity is driven by ATP hydrolysis and depends on interactions with microtubule tracks. Essential steps in kinesin movement rely on controlled alternate binding to and detaching from the microtubules. The conformational changes in the kinesin motors induced by nucleotide and microtubule binding are coordinated by structural elements within their motor domains. Loop L11 of the kinesin motor domain interacts with the microtubule and is implicated in both microtubule binding and sensing nucleotide bound to the active site of kinesin. Consistent with its proposed role as a microtubule sensor, loop L11 is rarely seen in crystal structures of unattached kinesins. Here, we report four structures of a regulated plant kinesin, the kinesin-like calmodulin binding protein (KCBP), determined by X-ray crystallography. Although all structures reveal the kinesin motor in the ATP-like conformation, its loop L11 is observed in different conformational states, both ordered and disordered. When structured, loop L11 adds three additional helical turns to the N-terminal part of the following helix [alpha]4. Although interactions with protein neighbors in the crystal support the ordering of loop L11, its observed conformation suggests the conformation for loop L11 in the microtubule-bound kinesin. Variations in the positions of other features of these kinesins were observed. A critical regulatory element of this kinesin, the calmodulin binding helix positioned at the C-terminus of the motor domain, is thought to confer negative regulation of KCBP. Calmodulin binds to this helix and inserts itself between the motor and the microtubule. Comparison of five independent structures of KCBP shows that the positioning of the calmodulin binding helix is not decided by crystal packing forces but is determined by the conformational state of the motor. The observed variations in the position of the calmodulin binding helix fit the regulatory mechanism previously proposed for this kinesin motor.

Conformational dynamics of loops L11 and L12 of kinesin as revealed by spin-labeling EPR

Masafumi Yamada — 2008-06-02T18:33:25-00:00

Biochemical and Biophysical Research Communications, Vol. 364, No. 3. (21 December 2007), pp. 620-626.

The EPR spectra of the spin labels attached to loops L11 and L12 of kinesin were resolved into slow (rotational correlation time, [tau] = 10-45 ns) and fast ([tau] = 2 ns) components. The fraction of the slow component increased considerably when kinesin was complexed with a microtubule (MT). On MT binding and in the presence of nucleotides ADP and AMPPNP, the spin labels on L11, particularly at A252C and L249C, significantly decreased the fraction of the slow component. Moreover, dipolar EPR detected a wide distribution in distance range, 1-2 nm between the two spin labels attached to T242C/A252C or A247C/A252C; this distribution was slightly narrower in the presence of MTs than in their absence. These results suggested that the L11 residues undergo conformational transition on the binding of nucleotides and MT, while these residues remained to fluctuate over a nanometer range.

Darwinian Evolution on a Chip

Brian Paegel — 2008-04-08T18:05:48-00:00

PLoS Biology, Vol. 6, No. 4. (1 April 2008), e85.

Computer control of Darwinian evolution has been demonstrated by propagating a population of RNA enzymes in a microfluidic device. The RNA population was challenged to catalyze the ligation of an oligonucleotide substrate under conditions of progressively lower substrate concentrations. A microchip-based serial dilution circuit automated an exponential growth phase followed by a 10-fold dilution, which was repeated for 500 log-growth iterations. Evolution was observed in real time as the population adapted and achieved progressively faster growth rates over time. The final evolved enzyme contained a set of 11 mutations that conferred a 90-fold improvement in substrate utilization, coinciding with the applied selective pressure. This system reduces evolution to a microfluidic algorithm, allowing the experimenter to observe and manipulate adaptation.

A Microtubule Interactome: Complexes with Roles in Cell Cycle and Mitosis

Julian Hughes — 2008-04-23T13:18:47-00:00

PLoS Biology, Vol. 6, No. 4. (1 April 2008), e98.

The microtubule (MT) cytoskeleton is required for many aspects of cell function, including the transport of intracellular materials, the maintenance of cell polarity, and the regulation of mitosis. These functions are coordinated by MT-associated proteins (MAPs), which work in concert with each other, binding MTs and altering their properties. We have used a MT cosedimentation assay, combined with 1D and 2D PAGE and mass spectrometry, to identify over 250 MAPs from early Drosophila embryos. We have taken two complementary approaches to analyse the cellular function of novel MAPs isolated using this approach. First, we have carried out an RNA interference (RNAi) screen, identifying 21 previously uncharacterised genes involved in MT organisation. Second, we have undertaken a bioinformatics analysis based on binary protein interaction data to produce putative interaction networks of MAPs. By combining both approaches, we have identified and validated MAP complexes with potentially important roles in cell cycle regulation and mitosis. This study therefore demonstrates that biologically relevant data can be harvested using such a multidisciplinary approach, and identifies new MAPs, many of which appear to be important in cell division.

Investigating protein dynamics in collective coordinate space.

A Kitao — 2006-01-27T21:59:21-00:00

Curr Opin Struct Biol, Vol. 9, No. 2. (April 1999), pp. 164-169.

Currently, collective coordinates are commonly employed in order to examine protein dynamics. In recent studies, they have been successfully applied to finding functionally relevant motions, to investigating the physical nature of protein dynamics, to sampling of the conformational space and to the analysis of experimental data. Collective coordinates also have other possible applications.

Collective protein dynamics in relation to function

Herman Berendsen — 2008-04-28T23:02:20-00:00

Current Opinion in Structural Biology, Vol. 10, No. 2. (1 April 2000), pp. 165-169.

Several techniques for the analysis of the internal motions of proteins are available -- separating large collective motions from small, presumably uninteresting motions. Such descriptions are helpful in the characterization of internal motions and provide insight into the energy landscape of proteins. The real challenge, however, is to relate large collective motions to functional properties, such as binding and regulation, or to folding. These issues have been recently addressed in several papers.

Molecular Dynamics Simulation of the Escherichia coli NikR Protein: Equilibrium Conformational Fluctuations Reveal Interdomain Allosteric Communication Pathways

Michael Bradley — 2008-04-26T13:52:38-00:00

Journal of Molecular Biology, Vol. In Press, Corrected Proof

Escherichia coli NikR is a homotetrameric Ni2+- and DNA-binding protein that functions as a transcriptional repressor of the NikABCDE nickel permease. The protein is composed of two distinct domains. The N-terminal 50 amino acids of each chain forms part of the dimeric ribbon-helix-helix (RHH) domains, a well-studied DNA-binding fold. The 83-residue C-terminal nickel-binding domain forms an ACT (aspartokinase, chorismate mutase, and TyrA) fold and contains the tetrameric interface. In this study, we have utilized an equilibrium molecular dynamics simulation in order to explore the conformational dynamics of the NikR tetramer and determine important residue interactions within and between the RHH and ACT domains to gain insight into the effects of Ni2+ on DNA-binding activity. The molecular simulation data were analyzed using two different correlation measures based on fluctuations in atomic position and noncovalent contacts together with a clustering algorithm to define groups of residues with similar correlation patterns for both types of correlation measure. Based on these analyses, we have defined a series of residue interrelationships that describe an allosteric communication pathway between the Ni2+- and DNA-binding sites, which are separated by 40 Å. Several of the residues identified by our analyses have been previously shown experimentally to be important for NikR function. An additional subset of the identified residues structurally connects the experimentally implicated residues and may help coordinate the allosteric communication between the ACT and RHH domains.

Development of polyphosphate parameters for use with the AMBER force field

Kristin Meagher — 2006-07-24T15:42:26-00:00

Journal of Computational Chemistry, Vol. 24, No. 9. (2003), pp. 1016-1025.

Accurate force fields are essential for reproducing the conformational and dynamic behavior of condensed-phase systems. The popular AMBER force field has parameters for monophosphates, but they do not extend well to polyphorylated molecules such as ADP and ATP. This work presents parameters for the partial charges, atom types, bond angles, and torsions in simple polyphosphorylated compounds. The parameters are based on molecular orbital calculations of methyldiphosphate and methyltriphosphate at the RHF/6-31+G* level. The new parameters were fit to the entire potential energy surface (not just minima) with an RMSD of 0.62 kcal/mol. This is exceptional agreement and a significant improvement over the current parameters that produce a potential surface with an RMSD of 7.8 kcal/mol to that of the ab initio calculations. Testing has shown that the parameters are transferable and capable of reproducing the gas-phase conformations of inorganic diphosphate and triphosphate. Also, the parameters are an improvement over existing parameters in the condensed phase as shown by minimizations of ATP bound in several proteins. These parameters are intended for use with the existing AMBER 94/99 force field, and they will permit users to apply AMBER to a wider variety of important enzymatic systems. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1016-1025, 2003

Comparison of multiple Amber force fields and development of improved protein backbone parameters

Viktor Hornak — 2006-12-19T15:02:47-00:00

Proteins: Structure, Function, and Bioinformatics, Vol. 65, No. 3. (2006), pp. 712-725.

The ff94 force field that is commonly associated with the Amber simulation package is one of the most widely used parameter sets for biomolecular simulation. After a decade of extensive use and testing, limitations in this force field, such as over-stabilization of ?-helices, were reported by us and other researchers. This led to a number of attempts to improve these parameters, resulting in a variety of ?Amber? force fields and significant difficulty in determining which should be used for a particular application. We show that several of these continue to suffer from inadequate balance between different secondary structure elements. In addition, the approach used in most of these studies neglected to account for the existence in Amber of two sets of backbone ϕ/? dihedral terms. This led to parameter sets that provide unreasonable conformational preferences for glycine. We report here an effort to improve the ϕ/? dihedral terms in the ff99 energy function. Dihedral term parameters are based on fitting the energies of multiple conformations of glycine and alanine tetrapeptides from high level ab initio quantum mechanical calculations. The new parameters for backbone dihedrals replace those in the existing ff99 force field. This parameter set, which we denote ff99SB, achieves a better balance of secondary structure elements as judged by improved distribution of backbone dihedrals for glycine and alanine with respect to PDB survey data. It also accomplishes improved agreement with published experimental data for conformational preferences of short alanine peptides and better accord with experimental NMR relaxation data of test protein systems. Proteins 2006. © 2006 Wiley-Liss, Inc.

Very fast empirical prediction and rationalization of protein pKa values.

H Li — 2007-10-16T14:56:51-00:00

Proteins, Vol. 61, No. 4. (1 December 2005), pp. 704-721.

A very fast empirical method is presented for structure-based protein pKa prediction and rationalization. The desolvation effects and intra-protein interactions, which cause variations in pKa values of protein ionizable groups, are empirically related to the positions and chemical nature of the groups proximate to the pKa sites. A computer program is written to automatically predict pKa values based on these empirical relationships within a couple of seconds. Unusual pKa values at buried active sites, which are among the most interesting protein pKa values, are predicted very well with the empirical method. A test on 233 carboxyl, 12 cysteine, 45 histidine, and 24 lysine pKa values in various proteins shows a root-mean-square deviation (RMSD) of 0.89 from experimental values. Removal of the 29 pKa values that are upper or lower limits results in an RMSD = 0.79 for the remaining 285 pKa values.

Transition networks for modeling the kinetics of conformational change in macromolecules

Frank Noé — 2008-04-17T18:13:16-00:00

Current Opinion in Structural Biology, Vol. 18, No. 2. (April 2008), pp. 154-162.

The kinetics and thermodynamics of complex transitions in biomolecules can be modeled in terms of a network of transitions between the relevant conformational substates. Such a transition network, which overcomes the fundamental limitations of reaction-coordinate-based methods, can be constructed either based on the features of the energy landscape, or from molecular dynamics simulations. Energy-landscape-based networks are generated with the aid of automated path-optimization methods, and, using graph-theoretical adaptive methods, can now be constructed for large molecules such as proteins. Dynamics-based networks, also called Markov State Models, can be interpreted and adaptively improved using statistical concepts, such as the mean first passage time, reactive flux and sampling error analysis. This makes transition networks powerful tools for understanding large-scale conformational changes.

Tracing evolutionary pressure

Kai Ye — 2008-03-26T14:55:55-00:00

Bioinformatics, Vol. 24, No. 7. (1 April 2008), pp. 908-915.

Motivation: Recent advances in sequencing techniques have yielded enormous amounts of protein sequence data from various species. This large dataset allows sequence comparison between paralogous and orthologous proteins to identify motifs or functional positions that account for the differences of functional subgroups ( specificity' positions). Algorithms such as SDPpred and the two-entropies analysis (TEA) have been developed to detect such specificity positions from a multiple sequence alignment (MSA) grouped into classes according to certain biological functions. Other algorithms such as TreeDet compute a classification and then predict specificity positions associated with it. However, there are still many unresolved questions: Was the optimal subdivision of a protein family achieved? Do the definitions at different levels of the phylogenetic tree affect the prediction of specificity positions? Can the whole phylogenetic tree be used instead of only one level in it to predict specificity positions? Results: Here we present a novel method, TEA-O (Two-entropies analysisObjective), to trace the evolutionary pressure from the root to the branches of the phylogenetic tree. At each level of the tree, a TEA plot is produced to capture the signal of the evolutionary pressure. A consensus TEA-O plot is composed from the whole series of plots to provide a condensed representation. Positions related to functions that evolved early (conserved) or later (specificity) are close to the lower-left or upper-left corner of the TEA-O plot, respectively. This novel approach allows an unbiased, user-independent, analysis of residue relevance in a protein family. We compared our TEA-O method with various algorithms using both synthetic and real protein sequences. The results show that our method is robust, sensitive to subtle differences in evolutionary pressure during evolution and comprehensive because all positions in the MSA are presented in the consensus plot. Availability: All computer programs and datasets used in this work are available at http://nava.liacs.nl/kye/TEA-O/ for academic use Contact: k.ye@lacdr.leidenuniv.nl 10.1093/bioinformatics/btn057

Computational methods for transcriptional regulation.

ED Siggia — 2006-08-25T21:25:29-00:00

Curr Opin Genet Dev, Vol. 15, No. 2. (April 2005), pp. 214-221.

How is the information from a thousand gene-expression arrays, the location of more than two hundred regulatory factors, and nine sequenced genomes to be integrated into a global view of the regulatory network in budding yeast? Computational methods that fit incomplete noisy data provide the outlines of regulatory pathways, but the errors are not quantified. In the fly, embryonic patterning has proved amenable to computational prediction, but only when the DNA-binding preferences of the relevant factors are taken into account. In both these model organisms, simply restricting attention to regulatory sequences that align with related species (i.e. "conserved") discards much information regarding what is functional.

Allostery: Absence of a Change in Shape Does Not Imply that Allostery Is Not at Play.

Chung-Jung Tsai — 2008-03-26T10:26:57-00:00

J Mol Biol (29 February 2008)

Allostery is essential for controlled catalysis, signal transmission, receptor trafficking, turning genes on and off, and apoptosis. It governs the organism's response to environmental and metabolic cues, dictating transient partner interactions in the cellular network. Textbooks taught us that allostery is a change of shape at one site on the protein surface brought about by ligand binding to another. For several years, it has been broadly accepted that the change of shape is not induced; rather, it is observed simply because a larger protein population presents it. Current data indicate that while side chains can reorient and rewire, allostery may not even involve a change of (backbone) shape. Assuming that the enthalpy change does not reverse the free-energy change due to the change in entropy, entropy is mainly responsible for binding.

What is principal component analysis?

Markus Ringnér — 2008-03-09T04:13:08-00:00

Nature Biotechnology, Vol. 26, No. 3., pp. 303-304.

Compensatory and Long-Range Changes in Picosecond-Nanosecond Main-Chain Dynamics upon Complex Formation: 15N Relaxation Analysis of the Free and Bound States of the Ubiquitin-like Domain of Human Plexin-B1 and the Small GTPase Rac1

S Bouguet-Bonnet — 2008-03-21T19:28:52-00:00

Journal of Molecular Biology, Vol. 377, No. 5. (11 April 2008), pp. 1474-1487.

The formation of a complex between Rac1 and the cytoplasmic domain of plexin-B1 is one of the first documented cases of a direct interaction between a small guanosine 5'-triphosphatase (GTPase) and a transmembrane receptor. Structural studies have begun to elucidate the role of this interaction for the signal transduction mechanism of plexins. Mapping of the Rac1 GTPase surface that contacts the Rho GTPase binding domain of plexin-B1 by solution NMR spectroscopy confirms the plexin domain as a GTPase effector protein. Regions neighboring the GTPase switch I and II regions are also involved in the interaction and there is considerable interest to examine the changes in protein dynamics that take place upon complex formation. Here we present main-chain nitrogen-15 relaxation measurements for the unbound proteins as well as for the Rho GTPase binding domain and Rac1 proteins in their complexed state. Derived order parameters, S2, show that considerable motions are maintained in the bound state of plexin. In fact, some of the changes in S2 on binding appear compensatory, exhibiting decreased as well as increased dynamics. Fluctuations in Rac1, already a largely rigid protein on the picosecond-nanosecond timescale, are overall diminished, but isomerization dynamics in the switch I and II regions of the GTPase are retained in the complex and appear to be propagated to the bound plexin domain. Remarkably, fluctuations in the GTPase are attenuated at sites, including helices [alpha]6 (the Rho-specific insert helix), [alpha]7 and [alpha]8, that are spatially distant from the interaction region with plexin. This effect of binding on long-range dynamics appears to be communicated by hinge sites and by subtle conformational changes in the protein. Similar to recent studies on other systems, we suggest that dynamical protein features are affected by allosteric mechanisms. Altered protein fluctuations are likely to prime the Rho GTPase-plexin complex for interactions with additional binding partners.