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Sample records for macromolecular structure solution

  1. Phenix - a comprehensive python-based system for macromolecular structure solution

    SciTech Connect

    Terwilliger, Thomas C; Hung, Li - Wei; Adams, Paul D; Afonine, Pavel V; Bunkoczi, Gabor; Chen, Vincent B; Davis, Ian; Echols, Nathaniel; Headd, Jeffrey J; Grosse Kunstleve, Ralf W; Mccoy, Airlie J; Moriarty, Nigel W; Oeffner, Robert; Read, Randy J; Richardson, David C; Richardson, Jane S; Zwarta, Peter H

    2009-01-01

    Macromolecular X-ray crystallography is routinely applied to understand biological processes at a molecular level. However, significant time and effort are still required to solve and complete many of these structures because of the need for manual interpretation of complex numerical data using many software packages, and the repeated use of interactive three-dimensional graphics. Phenix has been developed to provide a comprehensive system for crystallographic structure solution with an emphasis on automation of all procedures. This has relied on the development of algorithms that minimize or eliminate subjective input, the development of algorithms that automate procedures that are traditionally performed by hand, and finally the development of a framework that allows a tight integration between the algorithms.

  2. Data Mining of Macromolecular Structures.

    PubMed

    van Beusekom, Bart; Perrakis, Anastassis; Joosten, Robbie P

    2016-01-01

    The use of macromolecular structures is widespread for a variety of applications, from teaching protein structure principles all the way to ligand optimization in drug development. Applying data mining techniques on these experimentally determined structures requires a highly uniform, standardized structural data source. The Protein Data Bank (PDB) has evolved over the years toward becoming the standard resource for macromolecular structures. However, the process selecting the data most suitable for specific applications is still very much based on personal preferences and understanding of the experimental techniques used to obtain these models. In this chapter, we will first explain the challenges with data standardization, annotation, and uniformity in the PDB entries determined by X-ray crystallography. We then discuss the specific effect that crystallographic data quality and model optimization methods have on structural models and how validation tools can be used to make informed choices. We also discuss specific advantages of using the PDB_REDO databank as a resource for structural data. Finally, we will provide guidelines on how to select the most suitable protein structure models for detailed analysis and how to select a set of structure models suitable for data mining.

  3. Fractal Dimensions of Macromolecular Structures

    PubMed Central

    Todoroff, Nickolay; Kunze, Jens; Schreuder, Herman; Hessler, Gerhard; Baringhaus, Karl-Heinz; Schneider, Gisbert

    2014-01-01

    Quantifying the properties of macromolecules is a prerequisite for understanding their roles in biochemical processes. One of the less-explored geometric features of macromolecules is molecular surface irregularity, or ‘roughness’, which can be measured in terms of fractal dimension (D). In this study, we demonstrate that surface roughness correlates with ligand binding potential. We quantified the surface roughnesses of biological macromolecules in a large-scale survey that revealed D values between 2.0 and 2.4. The results of our study imply that surface patches involved in molecular interactions, such as ligand-binding pockets and protein-protein interfaces, exhibit greater local fluctuations in their fractal dimensions than ‘inert’ surface areas. We expect approximately 22 % of a protein’s surface outside of the crystallographically known ligand binding sites to be ligandable. These findings provide a fresh perspective on macromolecular structure and have considerable implications for drug design as well as chemical and systems biology. PMID:26213587

  4. Ordered macromolecular structures in ferrofluid mixtures

    SciTech Connect

    Hayter, J.B.; Pynn, R.; Charles, S.; Skjeltorp, A.T.; Trewhella, J.; Stubbs, G.; Timmins, P.

    1989-04-03

    We have observed ordering of dilute dispersions of spherical and cylindrical macromolecules in magnetized ferrofluids. The order results from structural correlations between macromolecular and ferrofluid particles rather than from macroscopic magnetostatic effects. We have aligned elongated macromolecules by this technique and have obtained anisotropic neutron-diffraction patterns, which reflect the internal structure of the macromolecules. The method provides a tool for orienting suspended macromolecular assemblies which are not amenable to conventional alignment techniques.

  5. Long-range correlations, geometrical structure, and transport properties of macromolecular solutions. The equivalence of configurational statistics and geometrodynamics of large molecules.

    PubMed

    Mezzasalma, Stefano A

    2007-12-01

    A special theory of Brownian relativity was previously proposed to describe the universal picture arising in ideal polymer solutions. In brief, it redefines a Gaussian macromolecule in a 4-dimensional diffusive spacetime, establishing a (weak) Lorentz-Poincaré invariance between liquid and polymer Einstein's laws for Brownian movement. Here, aimed at inquiring into the effect of correlations, we deepen the extension of the special theory to a general formulation. The previous statistical equivalence, for dynamic trajectories of liquid molecules and static configurations of macromolecules, and rather obvious in uncorrelated systems, is enlarged by a more general principle of equivalence, for configurational statistics and geometrodynamics. Accordingly, the three geodesic motion, continuity, and field equations could be rewritten, and a number of scaling behaviors were recovered in a spacetime endowed with general static isotropic metric (i.e., for equilibrium polymer solutions). We also dealt with universality in the volume fraction and, unexpectedly, found that a hyperscaling relation of the form, (average size) x (diffusivity) x (viscosity)1/2 ~f(N0, phi0) is fulfilled in several regimes, both in the chain monomer number (N) and polymer volume fraction (phi). Entangled macromolecular dynamics was treated as a geodesic light deflection, entaglements acting in close analogy to the field generated by a spherically symmetric mass source, where length fluctuations of the chain primitive path behave as azimuth fluctuations of its shape. Finally, the general transformation rule for translational and diffusive frames gives a coordinate gauge invariance, suggesting a widened Lorentz-Poincaré symmetry for Brownian statistics. We expect this approach to find effective applications to solutions of arbitrarily large molecules displaying a variety of structures, where the effect of geometry is more explicit and significant in itself (e.g., surfactants, lipids, proteins). PMID

  6. Solution-Phase Processes of Macromolecular Crystallization

    NASA Technical Reports Server (NTRS)

    Pusey, Marc L.; Minamitani, Elizabeth Forsythe

    2004-01-01

    We have proposed, for the tetragonal form of chicken egg lysozyme, that solution phase assembly processes are needed to form the growth units for crystal nucleation and growth. The starting point for the self-association process is the monomeric protein, and the final crystallographic symmetry is defined by the initial dimerization interactions of the monomers and subsequent n-mers formed, which in turn are a function of the crystallization conditions. It has been suggested that multimeric proteins generally incorporate the underlying multimers symmetry into the final crystallographic symmetry. We posed the question of what happens to a protein that is known to grow as an n-mer when it is placed in solution conditions where it is monomeric. The trypsin-treated, or cut, form of the protein canavalin (CCAN) has been shown to nucleate and grow crystals as a trimer from neutral to slightly acidic solutions. Under these conditions the solution is composed almost wholly of trimers. The insoluble protein can be readily dissolved by weakly basic solution, which results in a solution that is monomeric. There are three possible outcomes to an attempt at crystallization of the protein under monomeric (high pH) conditions: 1) we will obtain the same crystals as under trimer conditions, but at different protein concentrations governed by the self association equilibria; 2) we will obtain crystals having a different symmetry, based upon a monomeric growth unit; 3) we will not obtain crystals. Obtaining the first result would be indicative that the solution-phase self-association process is critical to the crystal nucleation and growth process. The second result would be less clear, as it may also reflect a pH-dependent shift in the trimer-trimer molecular interactions. The third result, particularly for experiments in the transition pH's between trimeric and monomeric CCAN, would indicate that the monomer does not crystallize, and that solution phase self association is not part

  7. Identifying and Visualizing Macromolecular Flexibility in Structural Biology

    PubMed Central

    Palamini, Martina; Canciani, Anselmo; Forneris, Federico

    2016-01-01

    Structural biology comprises a variety of tools to obtain atomic resolution data for the investigation of macromolecules. Conventional structural methodologies including crystallography, NMR and electron microscopy often do not provide sufficient details concerning flexibility and dynamics, even though these aspects are critical for the physiological functions of the systems under investigation. However, the increasing complexity of the molecules studied by structural biology (including large macromolecular assemblies, integral membrane proteins, intrinsically disordered systems, and folding intermediates) continuously demands in-depth analyses of the roles of flexibility and conformational specificity involved in interactions with ligands and inhibitors. The intrinsic difficulties in capturing often subtle but critical molecular motions in biological systems have restrained the investigation of flexible molecules into a small niche of structural biology. Introduction of massive technological developments over the recent years, which include time-resolved studies, solution X-ray scattering, and new detectors for cryo-electron microscopy, have pushed the limits of structural investigation of flexible systems far beyond traditional approaches of NMR analysis. By integrating these modern methods with powerful biophysical and computational approaches such as generation of ensembles of molecular models and selective particle picking in electron microscopy, more feasible investigations of dynamic systems are now possible. Using some prominent examples from recent literature, we review how current structural biology methods can contribute useful data to accurately visualize flexibility in macromolecular structures and understand its important roles in regulation of biological processes. PMID:27668215

  8. Identifying and Visualizing Macromolecular Flexibility in Structural Biology

    PubMed Central

    Palamini, Martina; Canciani, Anselmo; Forneris, Federico

    2016-01-01

    Structural biology comprises a variety of tools to obtain atomic resolution data for the investigation of macromolecules. Conventional structural methodologies including crystallography, NMR and electron microscopy often do not provide sufficient details concerning flexibility and dynamics, even though these aspects are critical for the physiological functions of the systems under investigation. However, the increasing complexity of the molecules studied by structural biology (including large macromolecular assemblies, integral membrane proteins, intrinsically disordered systems, and folding intermediates) continuously demands in-depth analyses of the roles of flexibility and conformational specificity involved in interactions with ligands and inhibitors. The intrinsic difficulties in capturing often subtle but critical molecular motions in biological systems have restrained the investigation of flexible molecules into a small niche of structural biology. Introduction of massive technological developments over the recent years, which include time-resolved studies, solution X-ray scattering, and new detectors for cryo-electron microscopy, have pushed the limits of structural investigation of flexible systems far beyond traditional approaches of NMR analysis. By integrating these modern methods with powerful biophysical and computational approaches such as generation of ensembles of molecular models and selective particle picking in electron microscopy, more feasible investigations of dynamic systems are now possible. Using some prominent examples from recent literature, we review how current structural biology methods can contribute useful data to accurately visualize flexibility in macromolecular structures and understand its important roles in regulation of biological processes.

  9. Identifying and Visualizing Macromolecular Flexibility in Structural Biology.

    PubMed

    Palamini, Martina; Canciani, Anselmo; Forneris, Federico

    2016-01-01

    Structural biology comprises a variety of tools to obtain atomic resolution data for the investigation of macromolecules. Conventional structural methodologies including crystallography, NMR and electron microscopy often do not provide sufficient details concerning flexibility and dynamics, even though these aspects are critical for the physiological functions of the systems under investigation. However, the increasing complexity of the molecules studied by structural biology (including large macromolecular assemblies, integral membrane proteins, intrinsically disordered systems, and folding intermediates) continuously demands in-depth analyses of the roles of flexibility and conformational specificity involved in interactions with ligands and inhibitors. The intrinsic difficulties in capturing often subtle but critical molecular motions in biological systems have restrained the investigation of flexible molecules into a small niche of structural biology. Introduction of massive technological developments over the recent years, which include time-resolved studies, solution X-ray scattering, and new detectors for cryo-electron microscopy, have pushed the limits of structural investigation of flexible systems far beyond traditional approaches of NMR analysis. By integrating these modern methods with powerful biophysical and computational approaches such as generation of ensembles of molecular models and selective particle picking in electron microscopy, more feasible investigations of dynamic systems are now possible. Using some prominent examples from recent literature, we review how current structural biology methods can contribute useful data to accurately visualize flexibility in macromolecular structures and understand its important roles in regulation of biological processes. PMID:27668215

  10. Identifying and Visualizing Macromolecular Flexibility in Structural Biology.

    PubMed

    Palamini, Martina; Canciani, Anselmo; Forneris, Federico

    2016-01-01

    Structural biology comprises a variety of tools to obtain atomic resolution data for the investigation of macromolecules. Conventional structural methodologies including crystallography, NMR and electron microscopy often do not provide sufficient details concerning flexibility and dynamics, even though these aspects are critical for the physiological functions of the systems under investigation. However, the increasing complexity of the molecules studied by structural biology (including large macromolecular assemblies, integral membrane proteins, intrinsically disordered systems, and folding intermediates) continuously demands in-depth analyses of the roles of flexibility and conformational specificity involved in interactions with ligands and inhibitors. The intrinsic difficulties in capturing often subtle but critical molecular motions in biological systems have restrained the investigation of flexible molecules into a small niche of structural biology. Introduction of massive technological developments over the recent years, which include time-resolved studies, solution X-ray scattering, and new detectors for cryo-electron microscopy, have pushed the limits of structural investigation of flexible systems far beyond traditional approaches of NMR analysis. By integrating these modern methods with powerful biophysical and computational approaches such as generation of ensembles of molecular models and selective particle picking in electron microscopy, more feasible investigations of dynamic systems are now possible. Using some prominent examples from recent literature, we review how current structural biology methods can contribute useful data to accurately visualize flexibility in macromolecular structures and understand its important roles in regulation of biological processes.

  11. Size evolution of highly amphiphilic macromolecular solution assemblies via a distinct bimodal pathway

    PubMed Central

    Kelley, Elizabeth G.; Murphy, Ryan P.; Seppala, Jonathan E.; Smart, Thomas P.; Hann, Sarah D.

    2014-01-01

    The solution self-assembly of macromolecular amphiphiles offers an efficient, bottom-up strategy for producing well--defined nanocarriers, with applications ranging from drug delivery to nanoreactors. Typically, the generation of uniform nanocarrier architecturesis controlled by processing methods that rely upon cosolvent mixtures. These preparation strategies hinge on the assumption that macromolecular solution nanostructures are kinetically stable following transfer from an organic/aqueous cosolvent into aqueous solution. Herein we demonstrate that unequivocal step-change shifts in micelle populations occur over several weeks following transfer into a highly selective solvent. The unexpected micelle growth evolves through a distinct bimodal distribution separated by multiple fusion events and critically depends on solution agitation. Notably, these results underscore fundamental similarities between assembly processes in amphiphilic polymer, small molecule, and protein systems. Moreover, the non-equilibrium micelle size increase can have a major impact on the assumed stability of solution assemblies, for which performance is dictated by nanocarrier size and structure. PMID:24710204

  12. Size evolution of highly amphiphilic macromolecular solution assemblies via a distinct bimodal pathway

    NASA Astrophysics Data System (ADS)

    Kelley, Elizabeth G.; Murphy, Ryan P.; Seppala, Jonathan E.; Smart, Thomas P.; Hann, Sarah D.; Sullivan, Millicent O.; Epps, Thomas H.

    2014-04-01

    The solution self-assembly of macromolecular amphiphiles offers an efficient, bottom-up strategy for producing well-defined nanocarriers, with applications ranging from drug delivery to nanoreactors. Typically, the generation of uniform nanocarrier architectures is controlled by processing methods that rely on cosolvent mixtures. These preparation strategies hinge on the assumption that macromolecular solution nanostructures are kinetically stable following transfer from an organic/aqueous cosolvent into aqueous solution. Herein we demonstrate that unequivocal step-change shifts in micelle populations occur over several weeks following transfer into a highly selective solvent. The unexpected micelle growth evolves through a distinct bimodal distribution separated by multiple fusion events and critically depends on solution agitation. Notably, these results underscore fundamental similarities between assembly processes in amphiphilic polymer, small molecule and protein systems. Moreover, the non-equilibrium micelle size increase can have a major impact on the assumed stability of solution assemblies, for which performance is dictated by nanocarrier size and structure.

  13. New methods of structure refinement for macromolecular structure determination by NMR

    PubMed Central

    Clore, G. Marius; Gronenborn, Angela M.

    1998-01-01

    Recent advances in multidimensional NMR methodology have permitted solution structures of proteins in excess of 250 residues to be solved. In this paper, we discuss several methods of structure refinement that promise to increase the accuracy of macromolecular structures determined by NMR. These methods include the use of a conformational database potential and direct refinement against three-bond coupling constants, secondary 13C shifts, 1H shifts, T1/T2 ratios, and residual dipolar couplings. The latter two measurements provide long range restraints that are not accessible by other solution NMR parameters. PMID:9600889

  14. Isotope labeling for NMR studies of macromolecular structure and interactions

    SciTech Connect

    Wright, P.E.

    1994-12-01

    Implementation of biosynthetic methods for uniform or specific isotope labeling of proteins, coupled with the recent development of powerful heteronuclear multidimensional NMR methods, has led to a dramatic increase in the size and complexity of macromolecular systems that are now amenable to NMR structural analysis. In recent years, a new technology has emerged that combines uniform {sup 13}C, {sup 15}N labeling with heteronuclear multidimensional NMR methods to allow NMR structural studies of systems approaching 25 to 30 kDa in molecular weight. In addition, with the introduction of specific {sup 13}C and {sup 15}N labels into ligands, meaningful NMR studies of complexes of even higher molecular weight have become feasible. These advances usher in a new era in which the earlier, rather stringent molecular weight limitations have been greatly surpassed and NMR can begin to address many central biological problems that involve macromolecular structure, dynamics, and interactions.

  15. Cryo-Electron Tomography for Structural Characterization of Macromolecular Complexes

    PubMed Central

    Cope, Julia; Heumann, John; Hoenger, Andreas

    2011-01-01

    Cryo-electron tomography (cryo-ET) is an emerging 3-D reconstruction technology that combines the principles of tomographic 3-D reconstruction with the unmatched structural preservation of biological material embedded in vitreous ice. Cryo-ET is particularly suited to investigating cell-biological samples and large macromolecular structures that are too polymorphic to be reconstructed by classical averaging-based 3-D reconstruction procedures. This unit aims to make cryo-ET accessible to newcomers and discusses the specialized equipment required, as well as the relevant advantages and hurdles associated with sample preparation by vitrification and cryo-ET. Protocols describe specimen preparation, data recording and 3-D data reconstruction for cryo-ET, with a special focus on macromolecular complexes. A step-by-step procedure for specimen vitrification by plunge freezing is provided, followed by the general practicalities of tilt-series acquisition for cryo-ET, including advice on how to select an area appropriate for acquiring a tilt series. A brief introduction to the underlying computational reconstruction principles applied in tomography is described, along with instructions for reconstructing a tomogram from cryo-tilt series data. Finally, a method is detailed for extracting small subvolumes containing identical macromolecular structures from tomograms for alignment and averaging as a means to increase the signal-to-noise ratio and eliminate missing wedge effects inherent in tomographic reconstructions. PMID:21842467

  16. Cryo-electron tomography for structural characterization of macromolecular complexes.

    PubMed

    Cope, Julia; Heumann, John; Hoenger, Andreas

    2011-08-01

    Cryo-electron tomography (cryo-ET) is an emerging 3-D reconstruction technology that combines the principles of tomographic 3-D reconstruction with the unmatched structural preservation of biological matter embedded in vitreous ice. Cryo-ET is particularly suited to investigating cell-biological samples and large macromolecular structures that are too polymorphic to be reconstructed by classical averaging-based 3-D reconstruction procedures. This unit aims to make cryo-ET accessible to newcomers and discusses the specialized equipment required, as well as relevant advantages and hurdles associated with sample preparation by vitrification and cryo-ET. Protocols describe specimen preparation, data recording and 3-D data reconstruction for cryo-ET, with a special focus on macromolecular complexes. A step-by-step procedure for specimen vitrification by plunge freezing is provided, followed by the general practicalities of tilt-series acquisition for cryo-ET, including advice on how to select an area appropriate for acquiring a tilt series. A brief introduction to the underlying computational reconstruction principles applied in tomography is described, along with instructions for reconstructing a tomogram from cryo-tilt series data. Finally, a method is detailed for extracting small subvolumes containing identical macromolecular structures from tomograms for alignment and averaging as a means to increase the signal-to-noise ratio and eliminate missing wedge effects inherent in tomographic reconstructions.

  17. Macromolecular structure phasing by neutron anomalous diffraction

    PubMed Central

    Cuypers, Maxime G.; Mason, Sax A.; Mossou, Estelle; Haertlein, Michael; Forsyth, V. Trevor; Mitchell, Edward P.

    2016-01-01

    In this report we show for the first time that neutron anomalous dispersion can be used in a practical manner to determine experimental phases of a protein crystal structure, providing a new tool for structural biologists. The approach is demonstrated through the use of a state-of-the-art monochromatic neutron diffractometer at the Institut Laue-Langevin (ILL) in combination with crystals of perdeuterated protein that minimise the level of hydrogen incoherent scattering and enhance the visibility of the anomalous signal. The protein used was rubredoxin in which cadmium replaced the iron at the iron-sulphur site. While this study was carried out using a steady-state neutron beam source, the results will be of major interest for capabilities at existing and emerging spallation neutron sources where time-of-flight instruments provide inherent energy discrimination. In particular this capability may be expected to offer unique opportunities to a rapidly developing structural biology community where there is increasing interest in the identification of protonation states, protein/water interactions and protein-ligand interactions – all of which are of central importance to a wide range of fundamental and applied areas in the biosciences. PMID:27511806

  18. Macromolecular crowding increases structural content of folded proteins.

    PubMed

    Perham, Michael; Stagg, Loren; Wittung-Stafshede, Pernilla

    2007-10-30

    Here we show that increased amount of secondary structure is acquired in the folded states of two structurally-different proteins (alpha-helical VlsE and alpha/beta flavodoxin) in the presence of macromolecular crowding agents. The structural content of flavodoxin and VlsE is enhanced by 33% and 70%, respectively, in 400 mg/ml Ficoll 70 (pH 7, 20 degrees C) and correlates with higher protein-thermal stability. In the same Ficoll range, there are only small effects on the unfolded-state structures of the proteins. This is the first in vitro assessment of crowding effects on the native-state structures at physiological conditions. Our findings imply that for proteins with low intrinsic stability, the functional structures in vivo may differ from those observed in dilute buffers. PMID:17919600

  19. REFMAC5 for the refinement of macromolecular crystal structures

    PubMed Central

    Murshudov, Garib N.; Skubák, Pavol; Lebedev, Andrey A.; Pannu, Navraj S.; Steiner, Roberto A.; Nicholls, Robert A.; Winn, Martyn D.; Long, Fei; Vagin, Alexei A.

    2011-01-01

    This paper describes various components of the macromolecular crystallographic refinement program REFMAC5, which is distributed as part of the CCP4 suite. REFMAC5 utilizes different likelihood functions depending on the diffraction data employed (amplitudes or intensities), the presence of twinning and the availability of SAD/SIRAS experimental diffraction data. To ensure chemical and structural integrity of the refined model, REFMAC5 offers several classes of restraints and choices of model parameterization. Reliable models at resolutions at least as low as 4 Å can be achieved thanks to low-resolution refinement tools such as secondary-structure restraints, restraints to known homologous structures, automatic global and local NCS restraints, ‘jelly-body’ restraints and the use of novel long-range restraints on atomic displacement parameters (ADPs) based on the Kullback–Leibler divergence. REFMAC5 additionally offers TLS parameterization and, when high-resolution data are available, fast refinement of anisotropic ADPs. Refinement in the presence of twinning is performed in a fully automated fashion. REFMAC5 is a flexible and highly optimized refinement package that is ideally suited for refinement across the entire resolution spectrum encountered in macromolecular crystallography. PMID:21460454

  20. Crosslinked macromolecular structures in bituminous coals: Theoretical and experimental considerations

    NASA Astrophysics Data System (ADS)

    Lucht, Lucy M.; Peppas, Nicolaos A.

    1981-02-01

    Ample evidence from physicochemical experiments suggests that bituminous coals can be described as highly crosslinked and entangled networks of macromolecular chains of irregular structure. Theoretically these structures can be analyzed by statistical mechanical models considering non-Gaussian distribution of the macro-molecular chains along with departure from the Flory theories of crosslinked macromolecules. The models of Kovac (1978) and Peppas and Lucht (1979) have been developed in order to describe non-extractable coal matrices and their behavior during swelling in appropriate swelling agents. The molecular weight between cross-links Mc and the crosslinking density ρx can be determined for various solvents and equilibrium swelling ratios. Few experimental data are available to which these models can be applied. Thus, in view of these new theoretical models, experimental research must be directed towards the reexamination of extraction and swelling behavior of bituminous coals. Some of the important parameters to be determined for characterization of the physical structure of coals include the thermodynamic interaction parameter χ, the crosslinking parameters Mc and ρx and the molecular weight distribution of the extractable coal portion.

  1. Macromolecular structure analysis and effective liquefaction pretreatment. Final report

    SciTech Connect

    Suuberg, E.M.; Yun, Y.; Lilly, W.D.; Leung, K.; Gates, T.; Otake, Y.; Deevi, S.C.

    1994-07-01

    This project was concerned with characterizing the changes in coal macromolecular structure, that are of significance for liquefaction pretreatments of coal. The macromolecular structure of the insoluble portion of coal is difficult to characterize. Techniques that do so indirectly (based upon, for example, NMR and FTIR characterizations of atomic linkages) are not particularly sensitive for this purpose. Techniques that characterize the elastic structure (such as solvent swelling) are much more sensitive to subtle changes in the network structure. It is for this reason that we focused upon these techniques. The overall objective involved identifying pretreatments that reduce the crosslinking (physical or chemical) of the network structure, and thus lead to materials that can be handled to a greater extent by traditional liquid-phase processing techniques. These techniques tend to be inherently more efficient at producing desirable products. This report is divided into seven chapters. Chapter II summarizes the main experimental approaches used throughout the project, and summarizes the main findings on the Argonne Premium coal samples. Chapter III considers synergistic effects of solvent pairs. It is divided into two subsections. The first is concerned with mixtures of CS{sub 2} with electron donor solvents. The second subsection is concerned with aromatic hydrocarbon - alcohol or hydrocarbon - alcohol mixtures, as might be of interest for preliquefaction delivery of catalysts into bituminous coals. Chapter IV deals with questions of how oxidation might influence the results that are obtained. Chapter V briefly details what conclusions may be drawn concerning the elastic behavior of coals, and the effects of thermal treatments on this behavior. Chapter VI is concerned with theories to describe the action of solvents that are capable of dissociating non-covalent crosslinks. Finally, Chapter VII discusses the practical implications of the study.

  2. Conformational States of Macromolecular Assemblies Explored by Integrative Structure Calculation

    PubMed Central

    Thalassinos, Konstantinos; Pandurangan, Arun Prasad; Xu, Min; Alber, Frank; Topf, Maya

    2013-01-01

    Summary A detailed description of macromolecular assemblies in multiple conformational states can be very valuable for understanding cellular processes. At present, structural determination of most assemblies in different biologically relevant conformations cannot be achieved by a single technique and thus requires an integrative approach that combines information from multiple sources. Different techniques require different computational methods to allow efficient and accurate data processing and analysis. Here, we summarize the latest advances and future challenges in computational methods that help the interpretation of data from two techniques—mass spectrometry and three-dimensional cryo-electron microscopy (with focus on alignment and classification of heterogeneous subtomograms from cryo-electron tomography). We evaluate how new developments in these two broad fields will lead to further integration with atomic structures to broaden our picture of the dynamic behavior of assemblies in their native environment. PMID:24010709

  3. Cryo electron microscopy to determine the structure of macromolecular complexes.

    PubMed

    Carroni, Marta; Saibil, Helen R

    2016-02-15

    Cryo-electron microscopy (cryo-EM) is a structural molecular and cellular biology technique that has experienced major advances in recent years. Technological developments in image recording as well as in processing software make it possible to obtain three-dimensional reconstructions of macromolecular assemblies at near-atomic resolution that were formerly obtained only by X-ray crystallography or NMR spectroscopy. In parallel, cryo-electron tomography has also benefitted from these technological advances, so that visualization of irregular complexes, organelles or whole cells with their molecular machines in situ has reached subnanometre resolution. Cryo-EM can therefore address a broad range of biological questions. The aim of this review is to provide a brief overview of the principles and current state of the cryo-EM field.

  4. Timely deposition of macromolecular structures is necessary for peer review

    PubMed Central

    Joosten, Robbie P.; Soueidan, Hayssam; Wessels, Lodewyk F. A.; Perrakis, Anastassis

    2013-01-01

    Most of the macromolecular structures in the Protein Data Bank (PDB), which are used daily by thousands of educators and scientists alike, are determined by X-ray crystallography. It was examined whether the crystallographic models and data were deposited to the PDB at the same time as the publications that describe them were submitted for peer review. This condition is necessary to ensure pre-publication validation and the quality of the PDB public archive. It was found that a significant proportion of PDB entries were submitted to the PDB after peer review of the corresponding publication started, and many were only submitted after peer review had ended. It is argued that clear description of journal policies and effective policing is important for pre-publication validation, which is key in ensuring the quality of the PDB and of peer-reviewed literature. PMID:24311569

  5. Macromolecular crowding can account for RNase-sensitive constraint of bacterial nucleoid structure

    SciTech Connect

    Foley, Patricia L.; Wilson, David B.; Shuler, Michael L.

    2010-04-23

    The shape and compaction of the bacterial nucleoid may affect the accessibility of genetic material to the transcriptional machinery in natural and synthetic systems. To investigate this phenomenon, the nature and contribution of RNA and protein to the compaction of nucleoids that had been gently released from Escherichia coli cells were investigated using fluorescent and transmission electron microscopy. We propose that the removal of RNA from the bacterial nucleoid affects nucleoid compaction by altering the branching density and molecular weight of the nucleoid. We show that a common detergent in nucleoid preparations, Brij 58, plays a previously unrecognized role as a macromolecular crowding agent. RNA-free nucleoids adopt a compact structure similar in size to exponential-phase nucleoids when the concentration of Brij 58 is increased, consistent with our hypothesis. We present evidence that control and protein-free nucleoids behave similarly in solutions containing a macromolecular crowding agent. These results show that the contribution to DNA compaction by nucleoid-associated proteins is small when compared to macromolecular crowding effects.

  6. Macromolecular Crystallography and Structural Biology Databases at NIST

    PubMed Central

    Gilliland, Gary L.

    2001-01-01

    In the late 1970s, macromolecular crystallography at NIST began with collaboration between NIST and NIH to establish a single-crystal neutron diffractometer. This instrument was constructed and employed to solve a number of crystal structures: bovine ribonuclease A, bovine-ribonuclease-uridine vanadate complex, and porcine insulin. In the mid 1980s a Biomolecular Structure Group was created establishing NIST capabilities in biomolecular singe-crystal x-ray diffraction. The group worked on a variety of structural problems until joining the NIST/UMBI Center for Advanced Research in Biotechnology (CARB) in 1987. Crystallographic studies at CARB were then focused on protein engineering efforts that included among others chymosin, subtilisin BPN', interleukin 1β, and glutathione S-transferase. Recently, the structural biology efforts have centered on enzymes in the chorismate metabolic pathways involved in amino acid biosynthesis and in structural genomics that involves determining the structures of “hypothetical” proteins to aid in assigning function. In addition to crystallographic studies, structural biology database activities began with the formal establishment of the Biological Macro-molecule Crystallization Database in 1989. Later, in 1997, NIST in partnership with Rutgers and UCSD formed the Research Collaboratory for Structural Bioinformatics that successfully acquired the Protein Data Bank. The NIST efforts in these activities have focused on data uniformity, establishing and maintaining the physical archive, and working with the NMR community. PMID:27500071

  7. Timely deposition of macromolecular structures is necessary for peer review

    SciTech Connect

    Joosten, Robbie P.; Soueidan, Hayssam; Wessels, Lodewyk F. A.; Perrakis, Anastassis

    2013-12-01

    Deposition of crystallographic structures should be concurrent with or prior to manuscript submission for peer review, enabling validation and increasing reliability of the PDB. Most of the macromolecular structures in the Protein Data Bank (PDB), which are used daily by thousands of educators and scientists alike, are determined by X-ray crystallography. It was examined whether the crystallographic models and data were deposited to the PDB at the same time as the publications that describe them were submitted for peer review. This condition is necessary to ensure pre-publication validation and the quality of the PDB public archive. It was found that a significant proportion of PDB entries were submitted to the PDB after peer review of the corresponding publication started, and many were only submitted after peer review had ended. It is argued that clear description of journal policies and effective policing is important for pre-publication validation, which is key in ensuring the quality of the PDB and of peer-reviewed literature.

  8. Automated identification of elemental ions in macromolecular crystal structures

    SciTech Connect

    Echols, Nathaniel Morshed, Nader; Afonine, Pavel V.; McCoy, Airlie J.; Read, Randy J.; Terwilliger, Thomas C.; Adams, Paul D.

    2014-04-01

    The solvent-picking procedure in phenix.refine has been extended and combined with Phaser anomalous substructure completion and analysis of coordination geometry to identify and place elemental ions. Many macromolecular model-building and refinement programs can automatically place solvent atoms in electron density at moderate-to-high resolution. This process frequently builds water molecules in place of elemental ions, the identification of which must be performed manually. The solvent-picking algorithms in phenix.refine have been extended to build common ions based on an analysis of the chemical environment as well as physical properties such as occupancy, B factor and anomalous scattering. The method is most effective for heavier elements such as calcium and zinc, for which a majority of sites can be placed with few false positives in a diverse test set of structures. At atomic resolution, it is observed that it can also be possible to identify tightly bound sodium and magnesium ions. A number of challenges that contribute to the difficulty of completely automating the process of structure completion are discussed.

  9. Microelectrophoretic study of calcium oxalate monohydrate in macromolecular solutions

    NASA Technical Reports Server (NTRS)

    Curreri, P. A.; Onoda, G. Y., Jr.; Finlayson, B.

    1987-01-01

    Electrophoretic mobilities were measured for calcium oxalate monohydrate (COM) in solutions containing macromolecules. Two mucopolysaccharides (sodium heparin and chondroitin sulfate) and two proteins (positively charged lysozyme and negatively charged bovine serum albumin) were studied as adsorbates. The effects of pH, calcium oxalate surface charge (varied by calcium or oxalate ion activity), and citrate concentration were investigated. All four macromolecules showed evidence for adsorption. The macromolecule concentrations needed for reversing the surface charge indicated that the mucopolysaccharides have greater affinity for the COM surface than the proteins. Citrate ions at high concentrations appear to compete effectively with the negative protein for surface sites but show no evidence for competing with the positively charged protein.

  10. The electrokinetic behavior of calcium oxalate monohydrate in macromolecular solutions

    NASA Technical Reports Server (NTRS)

    Curreri, P. A.; Onoda, G. Y., Jr.; Finlayson, B.

    1988-01-01

    Electrophoretic mobilities were measured for calcium oxalate monohydrate (COM) in solutions containing macromolecules. Two mucopolysaccharides (sodium heparin and chrondroitin sulfate) and two proteins (positively charged lysozyme and negatively charged bovine serum albumin) were studied as adsorbates. The effects of pH, calcium oxalate surface charge (varied by calcium or oxalate ion activity), and citrate concentration were investigated. All four macromolecules showed evidence for chemical adsorption. The macromolecule concentrations needed for reversing the surface charge indicated that the mucopopolysacchrides have greater affinity for the COM surface than the proteins. The amount of proteins that can chemically adsorb appears to be limited to approximately one monomolecular layer. When the surface charge is high, an insufficient number of proteins can chemically adsorb to neutralize or reverse the surface charge. The remaining surface charge is balanced by proteins held near the surface by longer range electrostatic forces only. Citrate ions at high concentrations appear to compete effectively with the negative protein for surface sites but show no evidence for competing with the positively charged protein.

  11. Macromolecular coal structure as revealed by novel diffusion tests

    SciTech Connect

    Peppas, N.A.; Olivares, J.; Drummond, R.; Lustig, S.

    1990-01-01

    The main goal of the present work was the elucidation of the mechanistic characteristics of dynamic transport of various penetrants (solvents) in thin sections of coals by examining their penetrant uptake, front swelling and stress development. An important objective of this work was the study of coal network structure in different thermodynamically compatible penetrants and the analysis of dynamic swelling in terms of present anomalous transport theories. Interferometry/polariscopy, surface image analysis and related techniques were used to quantify the stresses and solvent concentration profiles in these sections. Dynamic and equilibrium swelling behavior were correlated using the polar interaction contributions of the solvent solubility parameters. The penetrant front position was followed in thin coal sections as a function of time. The initial front velocity was calculated for various coals and penetrants. Our penetrant studies with thin coal section from the same coal sample but with different thickness show that within the range of 150 {mu}m to 1500{mu}m the transport mechanism of dimethyl formamide in the macromolecular coal network is non-Fickian. In fact, for the thickest samples the transport mechanism is predominately Case-II whereas in the thinner samples penetrant uptake may be diffusion-controlled. Studies in various penetrants such as acetone, cyclohexane, methanol, methyl ethyl ketone, toluene and methylene chloride indicated that penetrant transport is a non-Fickian phenomenon. Stresses and cracks were observed for transport of methylene chloride. 73 refs., 88 figs., 15 tabs.

  12. Macromolecular ab initio phasing enforcing secondary and tertiary structure

    PubMed Central

    Millán, Claudia; Sammito, Massimo; Usón, Isabel

    2015-01-01

    Ab initio phasing of macromolecular structures, from the native intensities alone with no experimental phase information or previous particular structural knowledge, has been the object of a long quest, limited by two main barriers: structure size and resolution of the data. Current approaches to extend the scope of ab initio phasing include use of the Patterson function, density modification and data extrapolation. The authors’ approach relies on the combination of locating model fragments such as polyalanine α-helices with the program PHASER and density modification with the program SHELXE. Given the difficulties in discriminating correct small substructures, many putative groups of fragments have to be tested in parallel; thus calculations are performed in a grid or supercomputer. The method has been named after the Italian painter Arcimboldo, who used to compose portraits out of fruit and vegetables. With ARCIMBOLDO, most collections of fragments remain a ‘still-life’, but some are correct enough for density modification and main-chain tracing to reveal the protein’s true portrait. Beyond α-helices, other fragments can be exploited in an analogous way: libraries of helices with modelled side chains, β-strands, predictable fragments such as DNA-binding folds or fragments selected from distant homologues up to libraries of small local folds that are used to enforce nonspecific tertiary structure; thus restoring the ab initio nature of the method. Using these methods, a number of unknown macromolecules with a few thousand atoms and resolutions around 2 Å have been solved. In the 2014 release, use of the program has been simplified. The software mediates the use of massive computing to automate the grid access required in difficult cases but may also run on a single multicore workstation (http://chango.ibmb.csic.es/ARCIMBOLDO_LITE) to solve straightforward cases. PMID:25610631

  13. Macromolecular ab initio phasing enforcing secondary and tertiary structure.

    PubMed

    Millán, Claudia; Sammito, Massimo; Usón, Isabel

    2015-01-01

    Ab initio phasing of macromolecular structures, from the native intensities alone with no experimental phase information or previous particular structural knowledge, has been the object of a long quest, limited by two main barriers: structure size and resolution of the data. Current approaches to extend the scope of ab initio phasing include use of the Patterson function, density modification and data extrapolation. The authors' approach relies on the combination of locating model fragments such as polyalanine α-helices with the program PHASER and density modification with the program SHELXE. Given the difficulties in discriminating correct small substructures, many putative groups of fragments have to be tested in parallel; thus calculations are performed in a grid or supercomputer. The method has been named after the Italian painter Arcimboldo, who used to compose portraits out of fruit and vegetables. With ARCIMBOLDO, most collections of fragments remain a 'still-life', but some are correct enough for density modification and main-chain tracing to reveal the protein's true portrait. Beyond α-helices, other fragments can be exploited in an analogous way: libraries of helices with modelled side chains, β-strands, predictable fragments such as DNA-binding folds or fragments selected from distant homologues up to libraries of small local folds that are used to enforce nonspecific tertiary structure; thus restoring the ab initio nature of the method. Using these methods, a number of unknown macromolecules with a few thousand atoms and resolutions around 2 Å have been solved. In the 2014 release, use of the program has been simplified. The software mediates the use of massive computing to automate the grid access required in difficult cases but may also run on a single multicore workstation (http://chango.ibmb.csic.es/ARCIMBOLDO_LITE) to solve straightforward cases.

  14. Macromolecular ab initio phasing enforcing secondary and tertiary structure.

    PubMed

    Millán, Claudia; Sammito, Massimo; Usón, Isabel

    2015-01-01

    Ab initio phasing of macromolecular structures, from the native intensities alone with no experimental phase information or previous particular structural knowledge, has been the object of a long quest, limited by two main barriers: structure size and resolution of the data. Current approaches to extend the scope of ab initio phasing include use of the Patterson function, density modification and data extrapolation. The authors' approach relies on the combination of locating model fragments such as polyalanine α-helices with the program PHASER and density modification with the program SHELXE. Given the difficulties in discriminating correct small substructures, many putative groups of fragments have to be tested in parallel; thus calculations are performed in a grid or supercomputer. The method has been named after the Italian painter Arcimboldo, who used to compose portraits out of fruit and vegetables. With ARCIMBOLDO, most collections of fragments remain a 'still-life', but some are correct enough for density modification and main-chain tracing to reveal the protein's true portrait. Beyond α-helices, other fragments can be exploited in an analogous way: libraries of helices with modelled side chains, β-strands, predictable fragments such as DNA-binding folds or fragments selected from distant homologues up to libraries of small local folds that are used to enforce nonspecific tertiary structure; thus restoring the ab initio nature of the method. Using these methods, a number of unknown macromolecules with a few thousand atoms and resolutions around 2 Å have been solved. In the 2014 release, use of the program has been simplified. The software mediates the use of massive computing to automate the grid access required in difficult cases but may also run on a single multicore workstation (http://chango.ibmb.csic.es/ARCIMBOLDO_LITE) to solve straightforward cases. PMID:25610631

  15. PDBe: improved accessibility of macromolecular structure data from PDB and EMDB.

    PubMed

    Velankar, Sameer; van Ginkel, Glen; Alhroub, Younes; Battle, Gary M; Berrisford, John M; Conroy, Matthew J; Dana, Jose M; Gore, Swanand P; Gutmanas, Aleksandras; Haslam, Pauline; Hendrickx, Pieter M S; Lagerstedt, Ingvar; Mir, Saqib; Fernandez Montecelo, Manuel A; Mukhopadhyay, Abhik; Oldfield, Thomas J; Patwardhan, Ardan; Sanz-García, Eduardo; Sen, Sanchayita; Slowley, Robert A; Wainwright, Michael E; Deshpande, Mandar S; Iudin, Andrii; Sahni, Gaurav; Salavert Torres, Jose; Hirshberg, Miriam; Mak, Lora; Nadzirin, Nurul; Armstrong, David R; Clark, Alice R; Smart, Oliver S; Korir, Paul K; Kleywegt, Gerard J

    2016-01-01

    The Protein Data Bank in Europe (http://pdbe.org) accepts and annotates depositions of macromolecular structure data in the PDB and EMDB archives and enriches, integrates and disseminates structural information in a variety of ways. The PDBe website has been redesigned based on an analysis of user requirements, and now offers intuitive access to improved and value-added macromolecular structure information. Unique value-added information includes lists of reviews and research articles that cite or mention PDB entries as well as access to figures and legends from full-text open-access publications that describe PDB entries. A powerful new query system not only shows all the PDB entries that match a given query, but also shows the 'best structures' for a given macromolecule, ligand complex or sequence family using data-quality information from the wwPDB validation reports. A PDBe RESTful API has been developed to provide unified access to macromolecular structure data available in the PDB and EMDB archives as well as value-added annotations, e.g. regarding structure quality and up-to-date cross-reference information from the SIFTS resource. Taken together, these new developments facilitate unified access to macromolecular structure data in an intuitive way for non-expert users and support expert users in analysing macromolecular structure data.

  16. DOMMINO 2.0: integrating structurally resolved protein-, RNA-, and DNA-mediated macromolecular interactions.

    PubMed

    Kuang, Xingyan; Dhroso, Andi; Han, Jing Ginger; Shyu, Chi-Ren; Korkin, Dmitry

    2016-01-01

    Macromolecular interactions are formed between proteins, DNA and RNA molecules. Being a principle building block in macromolecular assemblies and pathways, the interactions underlie most of cellular functions. Malfunctioning of macromolecular interactions is also linked to a number of diseases. Structural knowledge of the macromolecular interaction allows one to understand the interaction's mechanism, determine its functional implications and characterize the effects of genetic variations, such as single nucleotide polymorphisms, on the interaction. Unfortunately, until now the interactions mediated by different types of macromolecules, e.g. protein-protein interactions or protein-DNA interactions, are collected into individual and unrelated structural databases. This presents a significant obstacle in the analysis of macromolecular interactions. For instance, the homogeneous structural interaction databases prevent scientists from studying structural interactions of different types but occurring in the same macromolecular complex. Here, we introduce DOMMINO 2.0, a structural Database Of Macro-Molecular INteractiOns. Compared to DOMMINO 1.0, a comprehensive database on protein-protein interactions, DOMMINO 2.0 includes the interactions between all three basic types of macromolecules extracted from PDB files. DOMMINO 2.0 is automatically updated on a weekly basis. It currently includes ∼1,040,000 interactions between two polypeptide subunits (e.g. domains, peptides, termini and interdomain linkers), ∼43,000 RNA-mediated interactions, and ∼12,000 DNA-mediated interactions. All protein structures in the database are annotated using SCOP and SUPERFAMILY family annotation. As a result, protein-mediated interactions involving protein domains, interdomain linkers, C- and N- termini, and peptides are identified. Our database provides an intuitive web interface, allowing one to investigate interactions at three different resolution levels: whole subunit network

  17. PDBe: improved accessibility of macromolecular structure data from PDB and EMDB

    PubMed Central

    Velankar, Sameer; van Ginkel, Glen; Alhroub, Younes; Battle, Gary M.; Berrisford, John M.; Conroy, Matthew J.; Dana, Jose M.; Gore, Swanand P.; Gutmanas, Aleksandras; Haslam, Pauline; Hendrickx, Pieter M. S.; Lagerstedt, Ingvar; Mir, Saqib; Fernandez Montecelo, Manuel A.; Mukhopadhyay, Abhik; Oldfield, Thomas J.; Patwardhan, Ardan; Sanz-García, Eduardo; Sen, Sanchayita; Slowley, Robert A.; Wainwright, Michael E.; Deshpande, Mandar S.; Iudin, Andrii; Sahni, Gaurav; Salavert Torres, Jose; Hirshberg, Miriam; Mak, Lora; Nadzirin, Nurul; Armstrong, David R.; Clark, Alice R.; Smart, Oliver S.; Korir, Paul K.; Kleywegt, Gerard J.

    2016-01-01

    The Protein Data Bank in Europe (http://pdbe.org) accepts and annotates depositions of macromolecular structure data in the PDB and EMDB archives and enriches, integrates and disseminates structural information in a variety of ways. The PDBe website has been redesigned based on an analysis of user requirements, and now offers intuitive access to improved and value-added macromolecular structure information. Unique value-added information includes lists of reviews and research articles that cite or mention PDB entries as well as access to figures and legends from full-text open-access publications that describe PDB entries. A powerful new query system not only shows all the PDB entries that match a given query, but also shows the ‘best structures’ for a given macromolecule, ligand complex or sequence family using data-quality information from the wwPDB validation reports. A PDBe RESTful API has been developed to provide unified access to macromolecular structure data available in the PDB and EMDB archives as well as value-added annotations, e.g. regarding structure quality and up-to-date cross-reference information from the SIFTS resource. Taken together, these new developments facilitate unified access to macromolecular structure data in an intuitive way for non-expert users and support expert users in analysing macromolecular structure data. PMID:26476444

  18. Macromolecular properties and polymeric structure of canine tracheal mucins.

    PubMed Central

    Shankar, V; Virmani, A K; Naziruddin, B; Sachdev, G P

    1991-01-01

    Two high-Mr mucus glycoproteins (mucins), CTM-A and CTM-B, were highly purified from canine tracheal pouch secretions, and their macromolecular properties as well as polymeric structure were investigated. On SDS/composite-gel electrophoresis, a diffuse band was observed for each mucin. Polyacrylamide-gel electrophoresis using 6% gels also showed the absence of low-Mr contaminants in the mucins. Comparison of chemical and amino acid compositions revealed significant differences between the two mucins. Using a static-laser-light-scattering technique, CTM-A and CTM-B were found to have weight-average Mr values of about 11.0 x 10(6) and 1.4 x 10(6) respectively. Both mucins showed concentration-dependent aggregation in buffer containing 6 M-guanidine hydrochloride. Under similar experimental conditions, reduced-alkylated CTM-A had an Mr of 5.48 x 10(6) and showed no concentration-dependent aggregation. Hydrophobic properties of the mucins, investigated by the fluorescent probe technique using mansylphenylalanine as the probe, showed the presence of a large number of low-affinity (KD approx. 10(5) M) binding sites. These sites appeared to be located on the non-glycosylated regions of the protein core, since Pronase digestion of the mucins almost completely eliminated probe binding. Reduction of disulphide bonds of CTM-A and CTM-B did not significantly alter the probe-binding properties. Also, addition of increasing NaCl concentrations (0.03-1.0 M) to the buffer caused only a small change in the hydrophobic properties of native and reduced-alkylated mucins. CTM-A was deglycosylated, without notable in the hydrophobic properties of native and reduced-alkylated mucins. CTM-A was deglycosylated, without notable degradation, using a combination of chemical and enzymic methods. On SDS/PAGE the protein core was estimated to have an Mr of approx. 60,000. On the basis of the protein and carbohydrate contents of the major mucin CTM-A, the mucin monomer was calculated to have an

  19. Recent developments in phasing and structure refinement for macromolecular crystallography

    PubMed Central

    Adams, Paul D.; Afonine, Pavel V.; Grosse-Kunstleve, Ralf W.; Read, Randy J.; Richardson, Jane S.; Richardson, David C.; Terwilliger, Thomas C.

    2009-01-01

    Summary Central to crystallographic structure solution is obtaining accurate phases in order to build a molecular model, ultimately followed by refinement of that model to optimize its fit to the experimental diffraction data and prior chemical knowledge. Recent advances in phasing and model refinement and validation algorithms make it possible to arrive at better electron density maps and more accurate models. PMID:19700309

  20. The Neurobiologist's Guide to Structural Biology: A Primer on Why Macromolecular Structure Matters and How to Evaluate Structural Data

    PubMed Central

    Minor, Daniel L.

    2010-01-01

    Structural biology now plays a prominent role in addressing questions central to understanding how excitable cells function. Although interest in the insights gained from the definition and dissection of macromolecular anatomy is high, many neurobiologists remain unfamiliar with the methods employed. This primer aims to help neurobiologists understand approaches for probing macromolecular structure and where the limits and challenges remain. Using examples of macromolecules with neurobiological importance, the review covers X-ray crystallography, electron microscopy (EM), small-angle X-ray scattering (SAXS), and nuclear magnetic resonance (NMR) and biophysical methods with which these approaches are often paired: isothermal titration calorimetry (ITC), equilibrium analytical ultracentifugation, and molecular dynamics (MD). PMID:17521566

  1. Macromolecular Stabilization by Excluded Cosolutes: Mean Field Theory of Crowded Solutions.

    PubMed

    Sapir, Liel; Harries, Daniel

    2015-07-14

    We propose a mean field theory to account for the experimentally determined temperature dependence of protein stabilization that emerges in solutions crowded by preferentially excluded cosolutes. Based on regular solution theory and employing the Flory-Huggins approximation, our model describes cosolutes in terms of their size, and two temperature-dependent microscopic parameters that correspond to macromolecule-cosolute and bulk solution interactions. The theory not only predicts a "depletion force" that can account for the experimentally observed stabilization of protein folding or association in the presence of excluded cosolutes but also predicts the full range of associated entropic and enthalpic components. Remarkably, depending on cosolute identity and in accordance with experiments, the theory describes entropically as well as enthalpically dominated depletion forces, even those disfavored by entropy. This emerging depletion attraction cannot be simply linked to molecular volumes. Instead, the relevant parameter is an effective volume that represents an interplay between solvent, cosolute, and macromolecular interactions. We demonstrate that the apparent depletion free energy is often accompanied by significant yet compensating entropy and enthalpy terms that, although having a net zero contribution to stabilization, can obscure the underlying molecular mechanism. This study underscores the importance of including often-neglected free energy terms that correspond to solvent-cosolute and cosolute-macromolecule interactions, which for most typical cosolutes are expected to be temperature dependent. We propose that experiments specifically aimed at resolving the temperature-dependence of cosolute exclusion from macromolecular surfaces should help reveal the full range of the underlying molecular mechanisms of the depletion force.

  2. Dynamic simulation of concentrated macromolecular solutions with screened long-range hydrodynamic interactions: Algorithm and limitations

    PubMed Central

    Ando, Tadashi; Chow, Edmond; Skolnick, Jeffrey

    2013-01-01

    Hydrodynamic interactions exert a critical effect on the dynamics of macromolecules. As the concentration of macromolecules increases, by analogy to the behavior of semidilute polymer solutions or the flow in porous media, one might expect hydrodynamic screening to occur. Hydrodynamic screening would have implications both for the understanding of macromolecular dynamics as well as practical implications for the simulation of concentrated macromolecular solutions, e.g., in cells. Stokesian dynamics (SD) is one of the most accurate methods for simulating the motions of N particles suspended in a viscous fluid at low Reynolds number, in that it considers both far-field and near-field hydrodynamic interactions. This algorithm traditionally involves an O(N3) operation to compute Brownian forces at each time step, although asymptotically faster but more complex SD methods are now available. Motivated by the idea of hydrodynamic screening, the far-field part of the hydrodynamic matrix in SD may be approximated by a diagonal matrix, which is equivalent to assuming that long range hydrodynamic interactions are completely screened. This approximation allows sparse matrix methods to be used, which can reduce the apparent computational scaling to O(N). Previously there were several simulation studies using this approximation for monodisperse suspensions. Here, we employ newly designed preconditioned iterative methods for both the computation of Brownian forces and the solution of linear systems, and consider the validity of this approximation in polydisperse suspensions. We evaluate the accuracy of the diagonal approximation method using an intracellular-like suspension. The diffusivities of particles obtained with this approximation are close to those with the original method. However, this approximation underestimates intermolecular correlated motions, which is a trade-off between accuracy and computing efficiency. The new method makes it possible to perform large-scale and

  3. Macromolecular Stabilization by Excluded Cosolutes: Mean Field Theory of Crowded Solutions.

    PubMed

    Sapir, Liel; Harries, Daniel

    2015-07-14

    We propose a mean field theory to account for the experimentally determined temperature dependence of protein stabilization that emerges in solutions crowded by preferentially excluded cosolutes. Based on regular solution theory and employing the Flory-Huggins approximation, our model describes cosolutes in terms of their size, and two temperature-dependent microscopic parameters that correspond to macromolecule-cosolute and bulk solution interactions. The theory not only predicts a "depletion force" that can account for the experimentally observed stabilization of protein folding or association in the presence of excluded cosolutes but also predicts the full range of associated entropic and enthalpic components. Remarkably, depending on cosolute identity and in accordance with experiments, the theory describes entropically as well as enthalpically dominated depletion forces, even those disfavored by entropy. This emerging depletion attraction cannot be simply linked to molecular volumes. Instead, the relevant parameter is an effective volume that represents an interplay between solvent, cosolute, and macromolecular interactions. We demonstrate that the apparent depletion free energy is often accompanied by significant yet compensating entropy and enthalpy terms that, although having a net zero contribution to stabilization, can obscure the underlying molecular mechanism. This study underscores the importance of including often-neglected free energy terms that correspond to solvent-cosolute and cosolute-macromolecule interactions, which for most typical cosolutes are expected to be temperature dependent. We propose that experiments specifically aimed at resolving the temperature-dependence of cosolute exclusion from macromolecular surfaces should help reveal the full range of the underlying molecular mechanisms of the depletion force. PMID:26575781

  4. Boundary element solution of macromolecular electrostatics: interaction energy between two proteins.

    PubMed Central

    Zhou, H X

    1993-01-01

    The boundary element technique is implemented to solve for the electrostatic potential of macromolecules in an ionic solution. This technique entails solving surface integral equations that are equivalent to the Poisson and the Poisson-Boltzmann equations governing the electrostatic potential inside the macromolecules and and in the solvent. A simple but robust method is described for discretizing the macromolecular surfaces in order to approximate the integral equations by linear algebraic equations. Particular attention is paid to the interaction energy between two macromolecules, and an iterative procedure is devised to make the calculation more efficient. This iterative procedure is illustrated in the electron transfer system of cytochrome c and cytochrome c peroxidase. PMID:8218918

  5. Probing the Interplay of Size, Shape, and Solution Environment in Macromolecular Diffusion Using a Simple Refraction Experiment

    ERIC Educational Resources Information Center

    Mankidy, Bijith D.; Coutinho, Cecil A.; Gupta, Vinay K.

    2010-01-01

    The diffusion coefficient of polymers is a critical parameter in biomedicine, catalysis, chemical separations, nanotechnology, and other industrial applications. Here, measurement of macromolecular diffusion in solutions is described using a visually instructive, undergraduate-level optical refraction experiment based on Weiner's method. To…

  6. Protein crystallography for aspiring crystallographers or how to avoid pitfalls and traps in macromolecular structure determination

    PubMed Central

    Wlodawer, Alexander; Minor, Wladek; Dauter, Zbigniew; Jaskolski, Mariusz

    2014-01-01

    The number of macromolecular structures deposited in the Protein Data Bank now approaches 100 000, with the vast majority of them determined by crystallographic methods. Thousands of papers describing such structures have been published in the scientific literature, and 20 Nobel Prizes in chemistry or medicine have been awarded for discoveries based on macromolecular crystallography. New hardware and software tools have made crystallography appear to be an almost routine (but still far from being analytical) technique and many structures are now being determined by scientists with very limited experience in the practical aspects of the field. However, this apparent ease is sometimes illusory and proper procedures need to be followed to maintain high standards of structure quality. In addition, many noncrystallographers may have problems with the critical evaluation and interpretation of structural results published in the scientific literature. The present review provides an outline of the technical aspects of crystallography for less experienced practitioners, as well as information that might be useful for users of macromolecular structures, aiming to show them how to interpret (but not overinterpret) the information present in the coordinate files and in their description. A discussion of the extent of information that can be gleaned from the atomic coordinates of structures solved at different resolution is provided, as well as problems and pitfalls encountered in structure determination and interpretation. PMID:24034303

  7. Protein crystallography for aspiring crystallographers or how to avoid pitfalls and traps in macromolecular structure determination.

    PubMed

    Wlodawer, Alexander; Minor, Wladek; Dauter, Zbigniew; Jaskolski, Mariusz

    2013-11-01

    The number of macromolecular structures deposited in the Protein Data Bank now approaches 100,000, with the vast majority of them determined by crystallographic methods. Thousands of papers describing such structures have been published in the scientific literature, and 20 Nobel Prizes in chemistry or medicine have been awarded for discoveries based on macromolecular crystallography. New hardware and software tools have made crystallography appear to be an almost routine (but still far from being analytical) technique and many structures are now being determined by scientists with very limited experience in the practical aspects of the field. However, this apparent ease is sometimes illusory and proper procedures need to be followed to maintain high standards of structure quality. In addition, many noncrystallographers may have problems with the critical evaluation and interpretation of structural results published in the scientific literature. The present review provides an outline of the technical aspects of crystallography for less experienced practitioners, as well as information that might be useful for users of macromolecular structures, aiming to show them how to interpret (but not overinterpret) the information present in the coordinate files and in their description. A discussion of the extent of information that can be gleaned from the atomic coordinates of structures solved at different resolution is provided, as well as problems and pitfalls encountered in structure determination and interpretation.

  8. 3DEM Loupe: Analysis of macromolecular dynamics using structures from electron microscopy.

    PubMed

    Nogales-Cadenas, R; Jonic, S; Tama, F; Arteni, A A; Tabas-Madrid, D; Vázquez, M; Pascual-Montano, A; Sorzano, C O S

    2013-07-01

    Electron microscopy (EM) provides access to structural information of macromolecular complexes in the 3-20 Å resolution range. Normal mode analysis has been extensively used with atomic resolution structures and successfully applied to EM structures. The major application of normal modes is the identification of possible conformational changes in proteins. The analysis can throw light on the mechanism following ligand binding, protein-protein interactions, channel opening and other functional macromolecular movements. In this article, we present a new web server, 3DEM Loupe, which allows normal mode analysis of any uploaded EM volume using a user-friendly interface and an intuitive workflow. Results can be fully explored in 3D through animations and movies generated by the server. The application is freely available at http://3demloupe.cnb.csic.es.

  9. High-Resolution Macromolecular Structure Determination by MicroED, a cryo-EM Method.

    PubMed

    Rodriguez, J A; Gonen, T

    2016-01-01

    Microelectron diffraction (MicroED) is a new cryo-electron microscopy (cryo-EM) method capable of determining macromolecular structures at atomic resolution from vanishingly small 3D crystals. MicroED promises to solve atomic resolution structures from even the tiniest of crystals, less than a few hundred nanometers thick. MicroED complements frontier advances in crystallography and represents part of the rebirth of cryo-EM that is making macromolecular structure determination more accessible for all. Here we review the concept and practice of MicroED, for both the electron microscopist and crystallographer. Where other reviews have addressed specific details of the technique (Hattne et al., 2015; Shi et al., 2016; Shi, Nannenga, Iadanza, & Gonen, 2013), we aim to provide context and highlight important features that should be considered when performing a MicroED experiment.

  10. High-Resolution Macromolecular Structure Determination by MicroED, a cryo-EM Method.

    PubMed

    Rodriguez, J A; Gonen, T

    2016-01-01

    Microelectron diffraction (MicroED) is a new cryo-electron microscopy (cryo-EM) method capable of determining macromolecular structures at atomic resolution from vanishingly small 3D crystals. MicroED promises to solve atomic resolution structures from even the tiniest of crystals, less than a few hundred nanometers thick. MicroED complements frontier advances in crystallography and represents part of the rebirth of cryo-EM that is making macromolecular structure determination more accessible for all. Here we review the concept and practice of MicroED, for both the electron microscopist and crystallographer. Where other reviews have addressed specific details of the technique (Hattne et al., 2015; Shi et al., 2016; Shi, Nannenga, Iadanza, & Gonen, 2013), we aim to provide context and highlight important features that should be considered when performing a MicroED experiment. PMID:27572734

  11. Principles and Overview of Sampling Methods for Modeling Macromolecular Structure and Dynamics

    PubMed Central

    Moffatt, Ryan; Ma, Buyong; Nussinov, Ruth

    2016-01-01

    Investigation of macromolecular structure and dynamics is fundamental to understanding how macromolecules carry out their functions in the cell. Significant advances have been made toward this end in silico, with a growing number of computational methods proposed yearly to study and simulate various aspects of macromolecular structure and dynamics. This review aims to provide an overview of recent advances, focusing primarily on methods proposed for exploring the structure space of macromolecules in isolation and in assemblies for the purpose of characterizing equilibrium structure and dynamics. In addition to surveying recent applications that showcase current capabilities of computational methods, this review highlights state-of-the-art algorithmic techniques proposed to overcome challenges posed in silico by the disparate spatial and time scales accessed by dynamic macromolecules. This review is not meant to be exhaustive, as such an endeavor is impossible, but rather aims to balance breadth and depth of strategies for modeling macromolecular structure and dynamics for a broad audience of novices and experts. PMID:27124275

  12. A New Program for Detecting the Geometrical Core of a Set of Structures of Macromolecular Complexes.

    PubMed

    Vakulenko, Yu A; Nagaev, B E; Alexeevski, A V; Karyagina, A S; Spirin, S A

    2016-04-01

    Comparison of structures of homological proteins often helps to understand functionally significant features of these structures. This concerns not only structures of separate protein chains, but also structures of macromolecular complexes. In particular, a comparison of complexes of homologous proteins with DNA is significant for analysis of the recognition of DNA by proteins. We present program LCore for detecting geometrical cores of a family of structures; a geometrical core is a set of amino acid residues and nucleotides that disposed similarly in all structures of the family. We describe the algorithm of the program, its web interface, and an example of its application to analysis of complexes of homeodomains with DNA.

  13. The emerging role of native mass spectrometry in characterizing the structure and dynamics of macromolecular complexes

    PubMed Central

    Boeri Erba, Elisabetta; Petosa, Carlo

    2015-01-01

    Mass spectrometry (MS) is a powerful tool for determining the mass of biomolecules with high accuracy and sensitivity. MS performed under so-called “native conditions” (native MS) can be used to determine the mass of biomolecules that associate noncovalently. Here we review the application of native MS to the study of protein−ligand interactions and its emerging role in elucidating the structure of macromolecular assemblies, including soluble and membrane protein complexes. Moreover, we discuss strategies aimed at determining the stoichiometry and topology of subunits by inducing partial dissociation of the holo-complex. We also survey recent developments in "native top-down MS", an approach based on Fourier Transform MS, whereby covalent bonds are broken without disrupting non-covalent interactions. Given recent progress, native MS is anticipated to play an increasingly important role for researchers interested in the structure of macromolecular complexes. PMID:25676284

  14. Accurate macromolecular structures using minimal measurements from X-ray free-electron lasers.

    PubMed

    Hattne, Johan; Echols, Nathaniel; Tran, Rosalie; Kern, Jan; Gildea, Richard J; Brewster, Aaron S; Alonso-Mori, Roberto; Glöckner, Carina; Hellmich, Julia; Laksmono, Hartawan; Sierra, Raymond G; Lassalle-Kaiser, Benedikt; Lampe, Alyssa; Han, Guangye; Gul, Sheraz; DiFiore, Dörte; Milathianaki, Despina; Fry, Alan R; Miahnahri, Alan; White, William E; Schafer, Donald W; Seibert, M Marvin; Koglin, Jason E; Sokaras, Dimosthenis; Weng, Tsu-Chien; Sellberg, Jonas; Latimer, Matthew J; Glatzel, Pieter; Zwart, Petrus H; Grosse-Kunstleve, Ralf W; Bogan, Michael J; Messerschmidt, Marc; Williams, Garth J; Boutet, Sébastien; Messinger, Johannes; Zouni, Athina; Yano, Junko; Bergmann, Uwe; Yachandra, Vittal K; Adams, Paul D; Sauter, Nicholas K

    2014-05-01

    X-ray free-electron laser (XFEL) sources enable the use of crystallography to solve three-dimensional macromolecular structures under native conditions and without radiation damage. Results to date, however, have been limited by the challenge of deriving accurate Bragg intensities from a heterogeneous population of microcrystals, while at the same time modeling the X-ray spectrum and detector geometry. Here we present a computational approach designed to extract meaningful high-resolution signals from fewer diffraction measurements.

  15. Chirality as a physical aspect of structure formation in biological macromolecular systems

    NASA Astrophysics Data System (ADS)

    Malyshko, E. V.; Tverdislov, V. A.

    2016-08-01

    A novel regularity of hierarchical structures is found in the formation of chiral biological macromolecular systems. The formation of structures with alternating chirality (helical structures) serves as an instrument of stratification. The ability of a carbon atom to form chiral compounds is an important factor that determined the carbon basis of living systems on the Earth as well as their development through a series of chiral bifurcations. In the course of biological evolution, the helical structures became basic elements of the molecular machines in the cell. The discreteness of structural levels allowed the mechanical degrees of freedom formation in the molecular machines in the cell.

  16. The chemical structure of macromolecular fractions of a sulfur-rich oil

    NASA Astrophysics Data System (ADS)

    Richnow, Hans H.; Jenisch, Angela; Michaelis, Walter

    1993-06-01

    A selective stepwise chemical degradation has been developed for structural studies of highmolecularweight (HMW) fractions of sulfur-rich oils. The degradation steps are: (i) desulfurization (ii) cleavage of oxygen-carbon bonds (iii) oxidation of aromatic structural units. After each step, the remaining macromolecular matter was subjected to the subsequent reaction. This degradation scheme was applied to the asphaltene, the resin and a macromolecular fraction of low polarity (LPMF) of the Rozel Point oil. Total amounts of degraded low-molecular-weight compounds increased progressively in the order asphaltene < resin < LPMF. Desulfurization yielded mainly phytane, steranes and triterpanes. Oxygen-carbon bond cleavage resulted in hydrocarbon fractions predominated by n-alkanes and acyclic isoprenoids. The oxidation step afforded high amounts of linear carboxylic acids in the range of C 11 to C 33. The released compounds provide a more complete picture of the molecular structure of the oil fractions than previously available. Labelling experiments with deuterium atoms allowed to characterize the site of bonding and the type of linkage for the released compounds. Evidence is presented that subunits of the macromolecular network are attached simultaneously by oxygen and sulfur (n-alkanes, hopanes) or by sulfur and aromatic units ( n-alkanes, steranes).

  17. The R-factor gap in macromolecular crystallography: an untapped potential for insights on accurate structures

    PubMed Central

    Holton, James M; Classen, Scott; Frankel, Kenneth A; Tainer, John A

    2014-01-01

    In macromolecular crystallography, the agreement between observed and predicted structure factors (Rcryst and Rfree) is seldom better than 20%. This is much larger than the estimate of experimental error (Rmerge). The difference between Rcryst and Rmerge is the R-factor gap. There is no such gap in small-molecule crystallography, for which calculated structure factors are generally considered more accurate than the experimental measurements. Perhaps the true noise level of macromolecular data is higher than expected? Or is the gap caused by inaccurate phases that trap refined models in local minima? By generating simulated diffraction patterns using the program MLFSOM, and including every conceivable source of experimental error, we show that neither is the case. Processing our simulated data yielded values that were indistinguishable from those of real data for all crystallographic statistics except the final Rcryst and Rfree. These values decreased to 3.8% and 5.5% for simulated data, suggesting that the reason for high R-factors in macromolecular crystallography is neither experimental error nor phase bias, but rather an underlying inadequacy in the models used to explain our observations. The present inability to accurately represent the entire macromolecule with both its flexibility and its protein-solvent interface may be improved by synergies between small-angle X-ray scattering, computational chemistry and crystallography. The exciting implication of our finding is that macromolecular data contain substantial hidden and untapped potential to resolve ambiguities in the true nature of the nanoscale, a task that the second century of crystallography promises to fulfill. Database Coordinates and structure factors for the real data have been submitted to the Protein Data Bank under accession 4tws. PMID:25040949

  18. D Macromolecular Structure Analyses: Applications in Plant Proteins

    NASA Astrophysics Data System (ADS)

    Dede, Filiz; Dinler, Gizem; Sayers, Zehra

    Attempts to relate the function to the structural features of biological macromolecules have intensified through the use of synchrotron radiation (SR) based techniques. Small Angle X-ray Scattering (SXAS) is a technique that has become readily accessible through several bamlines on SR sources all over the world. SAXS is used for obtaining low resolution structural information and is particularly useful for macromolecules that do not easily crystallize or alternatively when time-resolved structural information is required. In this paper use of SAXS for determination of structural parameters of a wheat metallothionein as a fusion protein with glutathione-stransferase (GST) is presented together with low resolution models. Results are discussed in the framework of functional roles of wheat metallothionein.

  19. Methods for displaying macromolecular structural uncertainty: application to the globins.

    PubMed

    Altman, R B; Hughes, C; Gerstein, M B

    1995-06-01

    Most molecular graphics programs ignore any uncertainty in the atomic coordinates being displayed. Structures are displayed in terms of perfect points, spheres, and lines with no uncertainty. However, all experimental methods for defining structures, and many methods for predicting and comparing structures, associate uncertainties with each atomic coordinate. We have developed graphical representations that highlight these uncertainties. These representations are encapsulated in a new interactive display program, PROTEAND. PROTEAND represents structural uncertainty in three ways: (1) The traditional way: The program shows a collection of structures as superposed and overlapped stick-figure models. (2) Ellipsoids: At each atom position, the program shows an ellipsoid derived from a three-dimensional Gaussian model of uncertainty. This probabilistic model provides additional information about the relationship between atoms that can be displayed as a correlation matrix. (3) Rigid-body volumes: Using clouds of dots, the program can show the range of rigid-body motion of selected substructures, such as individual alpha helices. We illustrate the utility of these display modalities by the applying PROTEAND to the globin family of proteins, and show that certain types of structural variation are best illustrated with different methods of display.

  20. Macromolecular Crystallography for Synthetic Abiological Molecules: Combining xMDFF and PHENIX for Structure Determination of Cyanostar Macrocycles

    PubMed Central

    Singharoy, Abhishek; Venkatakrishnan, Balasubramanian; Liu, Yun; Mayne, Christopher G.; Lee, Semin; Chen, Chun-Hsing; Zlotnick, Adam; Schulten, Klaus; Flood, Amar H.

    2015-01-01

    Crystal structure determination has long provided insight into structure and bonding of small molecules. When those same small molecules are designed to come together in multi-molecular assemblies, such as in coordination cages, supramolecular architectures and organic-based frameworks, their crystallographic characteristics closely resemble biological macromolecules. This resemblance suggests that bio-macromolecular refinement approaches be used for structure determination of abiological molecular complexes that arise in an aggregate state. Following this suggestion we investigated the crystal structure of a pentagonal macrocycle, cyanostar, by means of biological structure analysis methods and compared results to traditional small molecule methods. Cyanostar presents difficulties seen in supramolecular crystallography including whole molecule disorder and highly flexible solvent molecules sitting in macrocyclic and intermolecule void spaces. We used the force-field assisted refinement method, molecular dynamics flexible fitting algorithm for X-ray crystallography (xMDFF), along with tools from the macromolecular structure determination suite PHENIX. We found that a standard implementation of PHENIX, namely one without xMDFF, either fails to produce a solution by molecular replacement alone or produces an inaccurate structure when using generic geometry restraints, even at a very high diffraction data resolution of 0.84 Å. The problems disappear when taking advantage of xMDFF, which applies an optimized force field to re-align molecular models during phasing by providing accurate restraints. The structure determination for this model system shows excellent agreement with the small-molecule methods. Therefore, the joint xMDFF-PHENIX refinement protocol provides a new strategy that uses macromolecule methods for structure determination of small molecules and their assemblies. PMID:26121416

  1. The PDB_REDO server for macromolecular structure model optimization.

    PubMed

    Joosten, Robbie P; Long, Fei; Murshudov, Garib N; Perrakis, Anastassis

    2014-07-01

    The refinement and validation of a crystallographic structure model is the last step before the coordinates and the associated data are submitted to the Protein Data Bank (PDB). The success of the refinement procedure is typically assessed by validating the models against geometrical criteria and the diffraction data, and is an important step in ensuring the quality of the PDB public archive [Read et al. (2011 ▶), Structure, 19, 1395-1412]. The PDB_REDO procedure aims for 'constructive validation', aspiring to consistent and optimal refinement parameterization and pro-active model rebuilding, not only correcting errors but striving for optimal interpretation of the electron density. A web server for PDB_REDO has been implemented, allowing thorough, consistent and fully automated optimization of the refinement procedure in REFMAC and partial model rebuilding. The goal of the web server is to help practicing crystallo-graphers to improve their model prior to submission to the PDB. For this, additional steps were implemented in the PDB_REDO pipeline, both in the refinement procedure, e.g. testing of resolution limits and k-fold cross-validation for small test sets, and as new validation criteria, e.g. the density-fit metrics implemented in EDSTATS and ligand validation as implemented in YASARA. Innovative ways to present the refinement and validation results to the user are also described, which together with auto-generated Coot scripts can guide users to subsequent model inspection and improvement. It is demonstrated that using the server can lead to substantial improvement of structure models before they are submitted to the PDB. PMID:25075342

  2. The PDB_REDO server for macromolecular structure model optimization

    PubMed Central

    Joosten, Robbie P.; Long, Fei; Murshudov, Garib N.; Perrakis, Anastassis

    2014-01-01

    The refinement and validation of a crystallographic structure model is the last step before the coordinates and the associated data are submitted to the Protein Data Bank (PDB). The success of the refinement procedure is typically assessed by validating the models against geometrical criteria and the diffraction data, and is an important step in ensuring the quality of the PDB public archive [Read et al. (2011 ▶), Structure, 19, 1395–1412]. The PDB_REDO procedure aims for ‘constructive validation’, aspiring to consistent and optimal refinement parameterization and pro-active model rebuilding, not only correcting errors but striving for optimal interpretation of the electron density. A web server for PDB_REDO has been implemented, allowing thorough, consistent and fully automated optimization of the refinement procedure in REFMAC and partial model rebuilding. The goal of the web server is to help practicing crystallo­graphers to improve their model prior to submission to the PDB. For this, additional steps were implemented in the PDB_REDO pipeline, both in the refinement procedure, e.g. testing of resolution limits and k-fold cross-validation for small test sets, and as new validation criteria, e.g. the density-fit metrics implemented in EDSTATS and ligand validation as implemented in YASARA. Innovative ways to present the refinement and validation results to the user are also described, which together with auto-generated Coot scripts can guide users to subsequent model inspection and improvement. It is demonstrated that using the server can lead to substantial improvement of structure models before they are submitted to the PDB. PMID:25075342

  3. Recovering a representative conformational ensemble from underdetermined macromolecular structural data.

    PubMed

    Berlin, Konstantin; Castañeda, Carlos A; Schneidman-Duhovny, Dina; Sali, Andrej; Nava-Tudela, Alfredo; Fushman, David

    2013-11-01

    Structural analysis of proteins and nucleic acids is complicated by their inherent flexibility, conferred, for example, by linkers between their contiguous domains. Therefore, the macromolecule needs to be represented by an ensemble of conformations instead of a single conformation. Determining this ensemble is challenging because the experimental data are a convoluted average of contributions from multiple conformations. As the number of the ensemble degrees of freedom generally greatly exceeds the number of independent observables, directly deconvolving experimental data into a representative ensemble is an ill-posed problem. Recent developments in sparse approximations and compressive sensing have demonstrated that useful information can be recovered from underdetermined (ill-posed) systems of linear equations by using sparsity regularization. Inspired by these advances, we designed the Sparse Ensemble Selection (SES) method for recovering multiple conformations from a limited number of observations. SES is more general and accurate than previously published minimum-ensemble methods, and we use it to obtain representative conformational ensembles of Lys48-linked diubiquitin, characterized by the residual dipolar coupling data measured at several pH conditions. These representative ensembles are validated against NMR chemical shift perturbation data and compared to maximum-entropy results. The SES method reproduced and quantified the previously observed pH dependence of the major conformation of Lys48-linked diubiquitin, and revealed lesser-populated conformations that are preorganized for binding known diubiquitin receptors, thus providing insights into possible mechanisms of receptor recognition by polyubiquitin. SES is applicable to any experimental observables that can be expressed as a weighted linear combination of data for individual states.

  4. Automated MAD and MIR structure solution.

    PubMed

    Terwilliger, T C; Berendzen, J

    1999-04-01

    Obtaining an electron-density map from X-ray diffraction data can be difficult and time-consuming even after the data have been collected, largely because MIR and MAD structure determinations currently require many subjective evaluations of the qualities of trial heavy-atom partial structures before a correct heavy-atom solution is obtained. A set of criteria for evaluating the quality of heavy-atom partial solutions in macromolecular crystallography have been developed. These have allowed the conversion of the crystal structure-solution process into an optimization problem and have allowed its automation. The SOLVE software has been used to solve MAD data sets with as many as 52 selenium sites in the asymmetric unit. The automated structure-solution process developed is a major step towards the fully automated structure-determination, model-building and refinement procedure which is needed for genomic scale structure determinations. PMID:10089316

  5. Automated structure solution with the PHENIX suite

    SciTech Connect

    Terwilliger, Thomas C; Zwart, Peter H; Afonine, Pavel V; Grosse - Kunstleve, Ralf W

    2008-01-01

    Significant time and effort are often required to solve and complete a macromolecular crystal structure. The development of automated computational methods for the analysis, solution, and completion of crystallographic structures has the potential to produce minimally biased models in a short time without the need for manual intervention. The PHENIX software suite is a highly automated system for macromolecular structure determination that can rapidly arrive at an initial partial model of a structure without significant human intervention, given moderate resolution, and good quality data. This achievement has been made possible by the development of new algorithms for structure determination, maximum-likelihood molecular replacement (PHASER), heavy-atom search (HySS), template- and pattern-based automated model-building (RESOLVE, TEXTAL), automated macromolecular refinement (phenix. refine), and iterative model-building, density modification and refinement that can operate at moderate resolution (RESOLVE, AutoBuild). These algorithms are based on a highly integrated and comprehensive set of crystallographic libraries that have been built and made available to the community. The algorithms are tightly linked and made easily accessible to users through the PHENIX Wizards and the PHENIX GUI.

  6. Automated Structure Solution with the PHENIX Suite

    SciTech Connect

    Zwart, Peter H.; Zwart, Peter H.; Afonine, Pavel; Grosse-Kunstleve, Ralf W.; Hung, Li-Wei; Ioerger, Tom R.; McCoy, A.J.; McKee, Eric; Moriarty, Nigel; Read, Randy J.; Sacchettini, James C.; Sauter, Nicholas K.; Storoni, L.C.; Terwilliger, Tomas C.; Adams, Paul D.

    2008-06-09

    Significant time and effort are often required to solve and complete a macromolecular crystal structure. The development of automated computational methods for the analysis, solution and completion of crystallographic structures has the potential to produce minimally biased models in a short time without the need for manual intervention. The PHENIX software suite is a highly automated system for macromolecular structure determination that can rapidly arrive at an initial partial model of a structure without significant human intervention, given moderate resolution and good quality data. This achievement has been made possible by the development of new algorithms for structure determination, maximum-likelihood molecular replacement (PHASER), heavy-atom search (HySS), template and pattern-based automated model-building (RESOLVE, TEXTAL), automated macromolecular refinement (phenix.refine), and iterative model-building, density modification and refinement that can operate at moderate resolution (RESOLVE, AutoBuild). These algorithms are based on a highly integrated and comprehensive set of crystallographic libraries that have been built and made available to the community. The algorithms are tightly linked and made easily accessible to users through the PHENIX Wizards and the PHENIX GUI.

  7. Resolving macromolecular structures from electron cryo-tomography data using subtomogram averaging in RELION.

    PubMed

    Bharat, Tanmay A M; Scheres, Sjors H W

    2016-11-01

    Electron cryo-tomography (cryo-ET) is a technique that is used to produce 3D pictures (tomograms) of complex objects such as asymmetric viruses, cellular organelles or whole cells from a series of tilted electron cryo-microscopy (cryo-EM) images. Averaging of macromolecular complexes found within tomograms is known as subtomogram averaging, and this technique allows structure determination of macromolecular complexes in situ. Subtomogram averaging is also gaining in popularity for the calculation of initial models for single-particle analysis. We describe herein a protocol for subtomogram averaging from cryo-ET data using the RELION software (http://www2.mrc-lmb.cam.ac.uk/relion). RELION was originally developed for cryo-EM single-particle analysis, and the subtomogram averaging approach presented in this protocol has been implemented in the existing workflow for single-particle analysis so that users may conveniently tap into existing capabilities of the RELION software. We describe how to calculate 3D models for the contrast transfer function (CTF) that describe the transfer of information in the imaging process, and we illustrate the results of classification and subtomogram averaging refinement for cryo-ET data of purified hepatitis B capsid particles and Saccharomyces cerevisiae 80S ribosomes. Using the steps described in this protocol, along with the troubleshooting and optimization guidelines, high-resolution maps can be obtained in which secondary structure elements are resolved subtomogram. PMID:27685097

  8. Resolving macromolecular structures from electron cryo-tomography data using subtomogram averaging in RELION.

    PubMed

    Bharat, Tanmay A M; Scheres, Sjors H W

    2016-11-01

    Electron cryo-tomography (cryo-ET) is a technique that is used to produce 3D pictures (tomograms) of complex objects such as asymmetric viruses, cellular organelles or whole cells from a series of tilted electron cryo-microscopy (cryo-EM) images. Averaging of macromolecular complexes found within tomograms is known as subtomogram averaging, and this technique allows structure determination of macromolecular complexes in situ. Subtomogram averaging is also gaining in popularity for the calculation of initial models for single-particle analysis. We describe herein a protocol for subtomogram averaging from cryo-ET data using the RELION software (http://www2.mrc-lmb.cam.ac.uk/relion). RELION was originally developed for cryo-EM single-particle analysis, and the subtomogram averaging approach presented in this protocol has been implemented in the existing workflow for single-particle analysis so that users may conveniently tap into existing capabilities of the RELION software. We describe how to calculate 3D models for the contrast transfer function (CTF) that describe the transfer of information in the imaging process, and we illustrate the results of classification and subtomogram averaging refinement for cryo-ET data of purified hepatitis B capsid particles and Saccharomyces cerevisiae 80S ribosomes. Using the steps described in this protocol, along with the troubleshooting and optimization guidelines, high-resolution maps can be obtained in which secondary structure elements are resolved subtomogram.

  9. PURY: a database of geometric restraints of hetero compounds for refinement in complexes with macromolecular structures.

    PubMed

    Andrejasic, Miha; Praaenikar, Jure; Turk, Dusan

    2008-11-01

    The number and variety of macromolecular structures in complex with ;hetero' ligands is growing. The need for rapid delivery of correct geometric parameters for their refinement, which is often crucial for understanding the biological relevance of the structure, is growing correspondingly. The current standard for describing protein structures is the Engh-Huber parameter set. It is an expert data set resulting from selection and analysis of the crystal structures gathered in the Cambridge Structural Database (CSD). Clearly, such a manual approach cannot be applied to the vast and ever-growing number of chemical compounds. Therefore, a database, named PURY, of geometric parameters of chemical compounds has been developed, together with a server that accesses it. PURY is a compilation of the whole CSD. It contains lists of atom classes and bonds connecting them, as well as angle, chirality, planarity and conformation parameters. The current compilation is based on CSD 5.28 and contains 1978 atom classes and 32,702 bonding, 237,068 angle, 201,860 dihedral and 64,193 improper geometric restraints. Analysis has confirmed that the restraints from the PURY database are suitable for use in macromolecular crystal structure refinement and should be of value to the crystallographic community. The database can be accessed through the web server http://pury.ijs.si/, which creates topology and parameter files from deposited coordinates in suitable forms for the refinement programs MAIN, CNS and REFMAC. In the near future, the server will move to the CSD website http://pury.ccdc.cam.ac.uk/.

  10. PURY: a database of geometric restraints of hetero compounds for refinement in complexes with macromolecular structures.

    PubMed

    Andrejasic, Miha; Praaenikar, Jure; Turk, Dusan

    2008-11-01

    The number and variety of macromolecular structures in complex with ;hetero' ligands is growing. The need for rapid delivery of correct geometric parameters for their refinement, which is often crucial for understanding the biological relevance of the structure, is growing correspondingly. The current standard for describing protein structures is the Engh-Huber parameter set. It is an expert data set resulting from selection and analysis of the crystal structures gathered in the Cambridge Structural Database (CSD). Clearly, such a manual approach cannot be applied to the vast and ever-growing number of chemical compounds. Therefore, a database, named PURY, of geometric parameters of chemical compounds has been developed, together with a server that accesses it. PURY is a compilation of the whole CSD. It contains lists of atom classes and bonds connecting them, as well as angle, chirality, planarity and conformation parameters. The current compilation is based on CSD 5.28 and contains 1978 atom classes and 32,702 bonding, 237,068 angle, 201,860 dihedral and 64,193 improper geometric restraints. Analysis has confirmed that the restraints from the PURY database are suitable for use in macromolecular crystal structure refinement and should be of value to the crystallographic community. The database can be accessed through the web server http://pury.ijs.si/, which creates topology and parameter files from deposited coordinates in suitable forms for the refinement programs MAIN, CNS and REFMAC. In the near future, the server will move to the CSD website http://pury.ccdc.cam.ac.uk/. PMID:19020347

  11. Function and dynamics of macromolecular complexes explored by integrative structural and computational biology.

    PubMed

    Purdy, Michael D; Bennett, Brad C; McIntire, William E; Khan, Ali K; Kasson, Peter M; Yeager, Mark

    2014-08-01

    Three vignettes exemplify the potential of combining EM and X-ray crystallographic data with molecular dynamics (MD) simulation to explore the architecture, dynamics and functional properties of multicomponent, macromolecular complexes. The first two describe how EM and X-ray crystallography were used to solve structures of the ribosome and the Arp2/3-actin complex, which enabled MD simulations that elucidated functional dynamics. The third describes how EM, X-ray crystallography, and microsecond MD simulations of a GPCR:G protein complex were used to explore transmembrane signaling by the β-adrenergic receptor. Recent technical advancements in EM, X-ray crystallography and computational simulation create unprecedented synergies for integrative structural biology to reveal new insights into heretofore intractable biological systems.

  12. Macromolecular conformation in solution. Study of carbonic anhydrase by the positron annihilation technique.

    PubMed Central

    Handel, E D; Graf, G; Glass, J C

    1980-01-01

    The structural features of carbonic anhydrase (carbonate hydro-lyase; EC 4.2.1.1) in aqueous solution were probed by the positron annihilation technique. The data obtained under varying conditions of temperature, pH, and enzyme concentration were interpreted in terms of the free volume model. The change of enzymic activity with temperature is accompanied by a change in free volume of the protein. Upon thermal denaturation an irreversible change in free volume of the molecule occurred. At low temperatures the protein-water interactions were investigated. These results are discussed in terms of current concepts of structure-function relationships in proteins. This study shows the sensitivity of the positron annihilation method toward the structure of proteins related to their overall conformation and to the nature of bound water. PMID:6789901

  13. Instrumentation on Multi-Scaled Scattering of Bio-Macromolecular Solutions

    PubMed Central

    Chu, Benjamin; Fang, Dufei; Mao, Yimin

    2015-01-01

    The design, construction and initial tests on a combined laser light scattering and synchrotron X-ray scattering instrument can cover studies of length scales from atomic sizes in Angstroms to microns and dynamics from microseconds to seconds are presented. In addition to static light scattering (SLS), dynamic light scattering (DLS), small angle X-ray scattering (SAXS) and wide angle X-ray diffraction (WAXD), the light scattering instrument is being developed to carry out studies in mildly turbid solutions, in the presence of multiple scattering. Three-dimensional photon cross correlation function (3D-PCCF) measurements have been introduced to couple with synchrotron X-ray scattering to study the structure, size and dynamics of macromolecules in solution. PMID:25946340

  14. Flexible torsion-angle noncrystallographic symmetry restraints for improved macromolecular structure refinement

    PubMed Central

    Headd, Jeffrey J.; Echols, Nathaniel; Afonine, Pavel V.; Moriarty, Nigel W.; Gildea, Richard J.; Adams, Paul D.

    2014-01-01

    One of the great challenges in refining macromolecular crystal structures is a low data-to-parameter ratio. Historically, knowledge from chemistry has been used to help to improve this ratio. When a macromolecule crystallizes with more than one copy in the asymmetric unit, the noncrystallographic symmetry relationships can be exploited to provide additional restraints when refining the working model. However, although globally similar, NCS-related chains often have local differences. To allow for local differences between NCS-related molecules, flexible torsion-based NCS restraints have been introduced, coupled with intelligent rotamer handling for protein chains, and are available in phenix.refine for refinement of models at all resolutions. PMID:24816103

  15. Analytical ultracentrifugation: A versatile tool for the characterisation of macromolecular complexes in solution.

    PubMed

    Patel, Trushar R; Winzor, Donald J; Scott, David J

    2016-02-15

    Analytical ultracentrifugation, an early technique developed for characterizing quantitatively the solution properties of macromolecules, remains a powerful aid to structural biologists in their quest to understand the formation of biologically important protein complexes at the molecular level. Treatment of the basic tenets of the sedimentation velocity and sedimentation equilibrium variants of analytical ultracentrifugation is followed by considerations of the roles that it, in conjunction with other physicochemical procedures, has played in resolving problems encountered in the delineation of complex formation for three biological systems - the cytoplasmic dynein complex, mitogen-activated protein kinase (ERK2) self-interaction, and the terminal catalytic complex in selenocysteine synthesis. PMID:26555086

  16. A graph theoretical approach for assessing bio-macromolecular complex structural stability.

    PubMed

    Del Carpio, Carlos Adriel; Iulian Florea, Mihai; Suzuki, Ai; Tsuboi, Hideyuki; Hatakeyama, Nozomu; Endou, Akira; Takaba, Hiromitsu; Ichiishi, Eiichiro; Miyamoto, Akira

    2009-11-01

    Fast and proper assessment of bio macro-molecular complex structural rigidity as a measure of structural stability can be useful in systematic studies to predict molecular function, and can also enable the design of rapid scoring functions to rank automatically generated bio-molecular complexes. Based on the graph theoretical approach of Jacobs et al. [Jacobs DJ, Rader AJ, Kuhn LA, Thorpe MF (2001) Protein flexibility predictions using graph theory. Proteins: Struct Funct Genet 44:150-165] for expressing molecular flexibility, we propose a new scheme to analyze the structural stability of bio-molecular complexes. This analysis is performed in terms of the identification in interacting subunits of clusters of flappy amino acids (those constituting regions of potential internal motion) that undergo an increase in rigidity at complex formation. Gains in structural rigidity of the interacting subunits upon bio-molecular complex formation can be evaluated by expansion of the network of intra-molecular inter-atomic interactions to include inter-molecular inter-atomic interaction terms. We propose two indices for quantifying this change: one local, which can express localized (at the amino acid level) structural rigidity, the other global to express overall structural stability for the complex. The new system is validated with a series of protein complex structures reported in the protein data bank. Finally, the indices are used as scoring coefficients to rank automatically generated protein complex decoys.

  17. Using support vector machines to improve elemental ion identification in macromolecular crystal structures

    SciTech Connect

    Morshed, Nader; Echols, Nathaniel; Adams, Paul D.

    2015-05-01

    A method to automatically identify possible elemental ions in X-ray crystal structures has been extended to use support vector machine (SVM) classifiers trained on selected structures in the PDB, with significantly improved sensitivity over manually encoded heuristics. In the process of macromolecular model building, crystallographers must examine electron density for isolated atoms and differentiate sites containing structured solvent molecules from those containing elemental ions. This task requires specific knowledge of metal-binding chemistry and scattering properties and is prone to error. A method has previously been described to identify ions based on manually chosen criteria for a number of elements. Here, the use of support vector machines (SVMs) to automatically classify isolated atoms as either solvent or one of various ions is described. Two data sets of protein crystal structures, one containing manually curated structures deposited with anomalous diffraction data and another with automatically filtered, high-resolution structures, were constructed. On the manually curated data set, an SVM classifier was able to distinguish calcium from manganese, zinc, iron and nickel, as well as all five of these ions from water molecules, with a high degree of accuracy. Additionally, SVMs trained on the automatically curated set of high-resolution structures were able to successfully classify most common elemental ions in an independent validation test set. This method is readily extensible to other elemental ions and can also be used in conjunction with previous methods based on a priori expectations of the chemical environment and X-ray scattering.

  18. Biological Macromolecular Structures Data from the RCSB Protein Data Bank (RCSB PDB)

    DOE Data Explorer

    The Research Collaboratory for Structural Bioinformatics (RCSB) is a non-profit consortium that works to improve understanding of the function of biological systems through the study of the 3-D structure of biological macromolecules. The RCSB PDB is one of three sites serving as deposition, data processing, and distribution sites of the Protein Data Bank Archive. Each site provides its own view of the primary data, thus providing a variety of tools and resources for the global community. RCSB is also the official keeper for the PDB archive, with sole access authority to the PDB archive directory structure and contents. The RCSB PDB Information Portal for Biological Macromolecular Structures offers online tools for search and retrieval, for visualizing structures, for depositing, validating, or downloading data, news and highlights, a discussion forum, and links to other areas of related research. The PDB archive is a repository of atomic coordinates and other information describing proteins and other important biological macromolecules. Structural biologists use methods such as X-ray crystallography, NMR spectroscopy, and cryo-electron microscopy to determine the location of each atom relative to each other in the molecule. They then deposit this information, which is then annotated and publicly released into the archive by the wwPDB. Results can be viewed as 3-D images or models.

  19. Macromolecular Crystallography for Synthetic Abiological Molecules: Combining xMDFF and PHENIX for Structure Determination of Cyanostar Macrocycles.

    PubMed

    Singharoy, Abhishek; Venkatakrishnan, Balasubramanian; Liu, Yun; Mayne, Christopher G; Lee, Semin; Chen, Chun-Hsing; Zlotnick, Adam; Schulten, Klaus; Flood, Amar H

    2015-07-15

    Crystal structure determination has long provided insight into structure and bonding of small molecules. When those same small molecules are designed to come together in multimolecular assemblies, such as in coordination cages, supramolecular architectures and organic-based frameworks, their crystallographic characteristics closely resemble biological macromolecules. This resemblance suggests that biomacromolecular refinement approaches be used for structure determination of abiological molecular complexes that arise in an aggregate state. Following this suggestion we investigated the crystal structure of a pentagonal macrocycle, cyanostar, by means of biological structure analysis methods and compared results to traditional small molecule methods. Cyanostar presents difficulties seen in supramolecular crystallography including whole molecule disorder and highly flexible solvent molecules sitting in macrocyclic and intermolecule void spaces. We used the force-field assisted refinement method, molecular dynamics flexible fitting algorithm for X-ray crystallography (xMDFF), along with tools from the macromolecular structure determination suite PHENIX. We found that a standard implementation of PHENIX, namely one without xMDFF, either fails to produce a solution by molecular replacement alone or produces an inaccurate structure when using generic geometry restraints, even at a very high diffraction data resolution of 0.84 Å. The problems disappear when taking advantage of xMDFF, which applies an optimized force field to realign molecular models during phasing by providing accurate restraints. The structure determination for this model system shows excellent agreement with the small-molecule methods. Therefore, the joint xMDFF-PHENIX refinement protocol provides a new strategy that uses macromolecule methods for structure determination of small molecules and their assemblies. PMID:26121416

  20. Using support vector machines to improve elemental ion identification in macromolecular crystal structures

    SciTech Connect

    Morshed, Nader; Echols, Nathaniel; Adams, Paul D.

    2015-04-25

    In the process of macromolecular model building, crystallographers must examine electron density for isolated atoms and differentiate sites containing structured solvent molecules from those containing elemental ions. This task requires specific knowledge of metal-binding chemistry and scattering properties and is prone to error. A method has previously been described to identify ions based on manually chosen criteria for a number of elements. Here, the use of support vector machines (SVMs) to automatically classify isolated atoms as either solvent or one of various ions is described. Two data sets of protein crystal structures, one containing manually curated structures deposited with anomalous diffraction data and another with automatically filtered, high-resolution structures, were constructed. On the manually curated data set, an SVM classifier was able to distinguish calcium from manganese, zinc, iron and nickel, as well as all five of these ions from water molecules, with a high degree of accuracy. Additionally, SVMs trained on the automatically curated set of high-resolution structures were able to successfully classify most common elemental ions in an independent validation test set. This method is readily extensible to other elemental ions and can also be used in conjunction with previous methods based on a priori expectations of the chemical environment and X-ray scattering.

  1. Using support vector machines to improve elemental ion identification in macromolecular crystal structures

    DOE PAGES

    Morshed, Nader; Echols, Nathaniel; Adams, Paul D.

    2015-04-25

    In the process of macromolecular model building, crystallographers must examine electron density for isolated atoms and differentiate sites containing structured solvent molecules from those containing elemental ions. This task requires specific knowledge of metal-binding chemistry and scattering properties and is prone to error. A method has previously been described to identify ions based on manually chosen criteria for a number of elements. Here, the use of support vector machines (SVMs) to automatically classify isolated atoms as either solvent or one of various ions is described. Two data sets of protein crystal structures, one containing manually curated structures deposited with anomalousmore » diffraction data and another with automatically filtered, high-resolution structures, were constructed. On the manually curated data set, an SVM classifier was able to distinguish calcium from manganese, zinc, iron and nickel, as well as all five of these ions from water molecules, with a high degree of accuracy. Additionally, SVMs trained on the automatically curated set of high-resolution structures were able to successfully classify most common elemental ions in an independent validation test set. This method is readily extensible to other elemental ions and can also be used in conjunction with previous methods based on a priori expectations of the chemical environment and X-ray scattering.« less

  2. Using support vector machines to improve elemental ion identification in macromolecular crystal structures

    PubMed Central

    Morshed, Nader; Echols, Nathaniel; Adams, Paul D.

    2015-01-01

    In the process of macromolecular model building, crystallographers must examine electron density for isolated atoms and differentiate sites containing structured solvent molecules from those containing elemental ions. This task requires specific knowledge of metal-binding chemistry and scattering properties and is prone to error. A method has previously been described to identify ions based on manually chosen criteria for a number of elements. Here, the use of support vector machines (SVMs) to automatically classify isolated atoms as either solvent or one of various ions is described. Two data sets of protein crystal structures, one containing manually curated structures deposited with anomalous diffraction data and another with automatically filtered, high-resolution structures, were constructed. On the manually curated data set, an SVM classifier was able to distinguish calcium from manganese, zinc, iron and nickel, as well as all five of these ions from water molecules, with a high degree of accuracy. Additionally, SVMs trained on the automatically curated set of high-resolution structures were able to successfully classify most common elemental ions in an independent validation test set. This method is readily extensible to other elemental ions and can also be used in conjunction with previous methods based on a priori expectations of the chemical environment and X-ray scattering. PMID:25945580

  3. Automated MAD and MIR structure solution

    SciTech Connect

    Terwilliger, Thomas C.; Berendzen, Joel

    1999-04-01

    A fully automated procedure for solving MIR and MAD structures has been developed using a scoring scheme to convert the structure-solution process into an optimization problem. Obtaining an electron-density map from X-ray diffraction data can be difficult and time-consuming even after the data have been collected, largely because MIR and MAD structure determinations currently require many subjective evaluations of the qualities of trial heavy-atom partial structures before a correct heavy-atom solution is obtained. A set of criteria for evaluating the quality of heavy-atom partial solutions in macromolecular crystallography have been developed. These have allowed the conversion of the crystal structure-solution process into an optimization problem and have allowed its automation. The SOLVE software has been used to solve MAD data sets with as many as 52 selenium sites in the asymmetric unit. The automated structure-solution process developed is a major step towards the fully automated structure-determination, model-building and refinement procedure which is needed for genomic scale structure determinations.

  4. The structural biology center at the APS: an integrated user facility for macromolecular crystallography

    NASA Astrophysics Data System (ADS)

    Rosenbaum, G.; Westbrook, E. M.

    1997-07-01

    The Structural Biology Center (SBC) has developed and operates a sector (undulator and bending magnet) of the APS as a user facility for macromolecular crystallography. Crystallographically determined structures of proteins, nucleic acids and their complexes with proteins, viruses, and complexes between macromolecules and small ligands have become of central importance in molecular and cellular biology. Major design goals were to make the extremely high brilliance of the APS available for brilliance limited studies, and to achieve a high throughput of less demanding studies, as well as optimization for MAS-phasing. Crystal samples will include extremely small crystals, crystals with large unit cells (viruses, ribosomes, etc.) and ensembles of closely similar crystal structures for drug design, protein engineering, etc. Data are recorded on a 3000×3000 pixel CCD-area detector (optionally on image plates). The x-ray optics of both beamlines has been designed to produce a highly demagnified image of the source in order to match the focal size with the sizes of the sample and the resolution element of the detector. Vertical focusing is achieved by a flat, cylindrically bent mirror. Horizontal focusing is achieved by sagitally bending the second crystal of the double crystal monochromator. Monochromatic fluxes of 1.3*1013 ph/s into focal sizes of 0.08 mm (horizontal)×0.04 mm (vertical) FWHM (flux density 3.5*1015 ph/s/mm2) have been recorded.

  5. MolProbity: all-atom structure validation for macromolecular crystallography

    SciTech Connect

    Chen, Vincent B.; Arendall, W. Bryan III; Headd, Jeffrey J.; Keedy, Daniel A.; Immormino, Robert M.; Kapral, Gary J.; Murray, Laura W.; Richardson, Jane S.; Richardson, David C.

    2010-01-01

    MolProbity structure validation will diagnose most local errors in macromolecular crystal structures and help to guide their correction. MolProbity is a structure-validation web service that provides broad-spectrum solidly based evaluation of model quality at both the global and local levels for both proteins and nucleic acids. It relies heavily on the power and sensitivity provided by optimized hydrogen placement and all-atom contact analysis, complemented by updated versions of covalent-geometry and torsion-angle criteria. Some of the local corrections can be performed automatically in MolProbity and all of the diagnostics are presented in chart and graphical forms that help guide manual rebuilding. X-ray crystallography provides a wealth of biologically important molecular data in the form of atomic three-dimensional structures of proteins, nucleic acids and increasingly large complexes in multiple forms and states. Advances in automation, in everything from crystallization to data collection to phasing to model building to refinement, have made solving a structure using crystallography easier than ever. However, despite these improvements, local errors that can affect biological interpretation are widespread at low resolution and even high-resolution structures nearly all contain at least a few local errors such as Ramachandran outliers, flipped branched protein side chains and incorrect sugar puckers. It is critical both for the crystallographer and for the end user that there are easy and reliable methods to diagnose and correct these sorts of errors in structures. MolProbity is the authors’ contribution to helping solve this problem and this article reviews its general capabilities, reports on recent enhancements and usage, and presents evidence that the resulting improvements are now beneficially affecting the global database.

  6. Solution structure of (+)-discodermolide.

    PubMed

    Smith, A B; LaMarche, M J; Falcone-Hindley, M

    2001-03-01

    [structure: see text]. The solution structure of (+)-discodermolide (1) has been determined via 1- and 2-D NMR techniques in conjunction with Monte Carlo conformational analysis. Taken together, the results demonstrate that in solution (+)-discodermolide occupies a helical conformation remarkably similar to the solid state conformation.

  7. Biomolecular interactions modulate macromolecular structure and dynamics in atomistic model of a bacterial cytoplasm

    PubMed Central

    Yu, Isseki; Mori, Takaharu; Ando, Tadashi; Harada, Ryuhei; Jung, Jaewoon; Sugita, Yuji; Feig, Michael

    2016-01-01

    Biological macromolecules function in highly crowded cellular environments. The structure and dynamics of proteins and nucleic acids are well characterized in vitro, but in vivo crowding effects remain unclear. Using molecular dynamics simulations of a comprehensive atomistic model cytoplasm we found that protein-protein interactions may destabilize native protein structures, whereas metabolite interactions may induce more compact states due to electrostatic screening. Protein-protein interactions also resulted in significant variations in reduced macromolecular diffusion under crowded conditions, while metabolites exhibited significant two-dimensional surface diffusion and altered protein-ligand binding that may reduce the effective concentration of metabolites and ligands in vivo. Metabolic enzymes showed weak non-specific association in cellular environments attributed to solvation and entropic effects. These effects are expected to have broad implications for the in vivo functioning of biomolecules. This work is a first step towards physically realistic in silico whole-cell models that connect molecular with cellular biology. DOI: http://dx.doi.org/10.7554/eLife.19274.001 PMID:27801646

  8. In situ macromolecular crystallography using microbeams

    PubMed Central

    Axford, Danny; Owen, Robin L.; Aishima, Jun; Foadi, James; Morgan, Ann W.; Robinson, James I.; Nettleship, Joanne E.; Owens, Raymond J.; Moraes, Isabel; Fry, Elizabeth E.; Grimes, Jonathan M.; Harlos, Karl; Kotecha, Abhay; Ren, Jingshan; Sutton, Geoff; Walter, Thomas S.; Stuart, David I.; Evans, Gwyndaf

    2012-01-01

    Despite significant progress in high-throughput methods in macromolecular crystallography, the production of diffraction-quality crystals remains a major bottleneck. By recording diffraction in situ from crystals in their crystallization plates at room temperature, a number of problems associated with crystal handling and cryoprotection can be side-stepped. Using a dedicated goniometer installed on the microfocus macromolecular crystallography beamline I24 at Diamond Light Source, crystals have been studied in situ with an intense and flexible microfocus beam, allowing weakly diffracting samples to be assessed without a manual crystal-handling step but with good signal to noise, despite the background scatter from the plate. A number of case studies are reported: the structure solution of bovine enterovirus 2, crystallization screening of membrane proteins and complexes, and structure solution from crystallization hits produced via a high-throughput pipeline. These demonstrate the potential for in situ data collection and structure solution with microbeams. PMID:22525757

  9. MACROMOLECULAR THERAPEUTICS

    PubMed Central

    Yang, Jiyuan; Kopeček, Jindřich

    2014-01-01

    This review covers water-soluble polymer-drug conjugates and macromolecules that possess biological activity without attached low molecular weight drugs. The main design principles of traditional and backbone degradable polymer-drug conjugates as well as the development of a new paradigm in nanomedicines – (low molecular weight) drug-free macromolecular therapeutics are discussed. To address the biological features of cancer, macromolecular therapeutics directed to stem/progenitor cells and the tumor microenvironment are deliberated. Finally, the future perspectives of the field are briefly debated. PMID:24747162

  10. Predictive Mechanical Characterization of Macro-Molecular Material Chemistry Structures of Cement Paste at Nano Scale - Two-phase Macro-Molecular Structures of Calcium Silicate Hydrate, Tri-Calcium Silicate, Di-Calcium Silicate and Calcium Hydroxide

    NASA Astrophysics Data System (ADS)

    Padilla Espinosa, Ingrid Marcela

    Concrete is a hierarchical composite material with a random structure over a wide range of length scales. At submicron length scale the main component of concrete is cement paste, formed by the reaction of Portland cement clinkers and water. Cement paste acts as a binding matrix for the other components and is responsible for the strength of concrete. Cement paste microstructure contains voids, hydrated and unhydrated cement phases. The main crystalline phases of unhydrated cement are tri-calcium silicate (C3S) and di-calcium silicate (C2S), and of hydrated cement are calcium silicate hydrate (CSH) and calcium hydroxide (CH). Although efforts have been made to comprehend the chemical and physical nature of cement paste, studies at molecular level have primarily been focused on individual components. Present research focuses on the development of a method to model, at molecular level, and analysis of the two-phase combination of hydrated and unhydrated phases of cement paste as macromolecular systems. Computational molecular modeling could help in understanding the influence of the phase interactions on the material properties, and mechanical performance of cement paste. Present work also strives to create a framework for molecular level models suitable for potential better comparisons with low length scale experimental methods, in which the sizes of the samples involve the mixture of different hydrated and unhydrated crystalline phases of cement paste. Two approaches based on two-phase cement paste macromolecular structures, one involving admixed molecular phases, and the second involving cluster of two molecular phases are investigated. The mechanical properties of two-phase macromolecular systems of cement paste consisting of key hydrated phase CSH and unhydrated phases C3S or C2S, as well as CSH with the second hydrated phase CH were calculated. It was found that these cement paste two-phase macromolecular systems predicted an isotropic material behavior. Also

  11. Free kick instead of cross-validation in maximum-likelihood refinement of macromolecular crystal structures

    SciTech Connect

    Pražnikar, Jure; Turk, Dušan

    2014-12-01

    The maximum-likelihood free-kick target, which calculates model error estimates from the work set and a randomly displaced model, proved superior in the accuracy and consistency of refinement of crystal structures compared with the maximum-likelihood cross-validation target, which calculates error estimates from the test set and the unperturbed model. The refinement of a molecular model is a computational procedure by which the atomic model is fitted to the diffraction data. The commonly used target in the refinement of macromolecular structures is the maximum-likelihood (ML) function, which relies on the assessment of model errors. The current ML functions rely on cross-validation. They utilize phase-error estimates that are calculated from a small fraction of diffraction data, called the test set, that are not used to fit the model. An approach has been developed that uses the work set to calculate the phase-error estimates in the ML refinement from simulating the model errors via the random displacement of atomic coordinates. It is called ML free-kick refinement as it uses the ML formulation of the target function and is based on the idea of freeing the model from the model bias imposed by the chemical energy restraints used in refinement. This approach for the calculation of error estimates is superior to the cross-validation approach: it reduces the phase error and increases the accuracy of molecular models, is more robust, provides clearer maps and may use a smaller portion of data for the test set for the calculation of R{sub free} or may leave it out completely.

  12. Novel 3D bio-macromolecular bilinear descriptors for protein science: Predicting protein structural classes.

    PubMed

    Marrero-Ponce, Yovani; Contreras-Torres, Ernesto; García-Jacas, César R; Barigye, Stephen J; Cubillán, Néstor; Alvarado, Ysaías J

    2015-06-01

    In the present study, we introduce novel 3D protein descriptors based on the bilinear algebraic form in the ℝ(n) space on the coulombic matrix. For the calculation of these descriptors, macromolecular vectors belonging to ℝ(n) space, whose components represent certain amino acid side-chain properties, were used as weighting schemes. Generalization approaches for the calculation of inter-amino acidic residue spatial distances based on Minkowski metrics are proposed. The simple- and double-stochastic schemes were defined as approaches to normalize the coulombic matrix. The local-fragment indices for both amino acid-types and amino acid-groups are presented in order to permit characterizing fragments of interest in proteins. On the other hand, with the objective of taking into account specific interactions among amino acids in global or local indices, geometric and topological cut-offs are defined. To assess the utility of global and local indices a classification model for the prediction of the major four protein structural classes, was built with the Linear Discriminant Analysis (LDA) technique. The developed LDA-model correctly classifies the 92.6% and 92.7% of the proteins on the training and test sets, respectively. The obtained model showed high values of the generalized square correlation coefficient (GC(2)) on both the training and test series. The statistical parameters derived from the internal and external validation procedures demonstrate the robustness, stability and the high predictive power of the proposed model. The performance of the LDA-model demonstrates the capability of the proposed indices not only to codify relevant biochemical information related to the structural classes of proteins, but also to yield suitable interpretability. It is anticipated that the current method will benefit the prediction of other protein attributes or functions. PMID:25843214

  13. Novel 3D bio-macromolecular bilinear descriptors for protein science: Predicting protein structural classes.

    PubMed

    Marrero-Ponce, Yovani; Contreras-Torres, Ernesto; García-Jacas, César R; Barigye, Stephen J; Cubillán, Néstor; Alvarado, Ysaías J

    2015-06-01

    In the present study, we introduce novel 3D protein descriptors based on the bilinear algebraic form in the ℝ(n) space on the coulombic matrix. For the calculation of these descriptors, macromolecular vectors belonging to ℝ(n) space, whose components represent certain amino acid side-chain properties, were used as weighting schemes. Generalization approaches for the calculation of inter-amino acidic residue spatial distances based on Minkowski metrics are proposed. The simple- and double-stochastic schemes were defined as approaches to normalize the coulombic matrix. The local-fragment indices for both amino acid-types and amino acid-groups are presented in order to permit characterizing fragments of interest in proteins. On the other hand, with the objective of taking into account specific interactions among amino acids in global or local indices, geometric and topological cut-offs are defined. To assess the utility of global and local indices a classification model for the prediction of the major four protein structural classes, was built with the Linear Discriminant Analysis (LDA) technique. The developed LDA-model correctly classifies the 92.6% and 92.7% of the proteins on the training and test sets, respectively. The obtained model showed high values of the generalized square correlation coefficient (GC(2)) on both the training and test series. The statistical parameters derived from the internal and external validation procedures demonstrate the robustness, stability and the high predictive power of the proposed model. The performance of the LDA-model demonstrates the capability of the proposed indices not only to codify relevant biochemical information related to the structural classes of proteins, but also to yield suitable interpretability. It is anticipated that the current method will benefit the prediction of other protein attributes or functions.

  14. Extra-column dispersion of macromolecular solutes in aqueous-phase size-exclusion chromatography.

    PubMed

    Grznárová, G; Polakovic, M; Acai, P; Görner, T

    2004-06-18

    A set of dextran standards was used to study the extra-column dispersion in conventional chromatographic equipment at a broad range of molecular weights, different mobile phase flow rates and connecting tube lengths and diameters. All known correlations for the tube dispersion at laminar flow, including those for short tubes, overestimated the values of the variance of the outlet concentration signal. The difference increased with the solute molecular weight and the flow rate. It was assumed that the discrepancy was due to the effect of natural convection invoked by the density differences of the injected dextran solutions and water. A suitable approximation of the relative band spreading was suggested in a form of a power function of the Reynolds and Schmidt numbers. A significant decrease of the dispersion was observed when the chromatography tubing was coiled into a circle. This decrease was successfully predicted combining the existing correlations for long coiled tubes and short straight tubes.

  15. Complexation of Statins with β-Cyclodextrin in Solutions of Small Molecular Additives and Macromolecular Colloids

    NASA Astrophysics Data System (ADS)

    Süle, András; Csempesz, Ferenc

    The solubility of lovastatin and simvastatin (inevitable drugs in the management of cardiovascular diseases) was studied by phase-solubility measurements in multicomponent colloidal and non-colloidal media. Complexation in aqueous solutions of the highly lipophilic statins with β-cyclodextrin (β-CD) in the absence and the presence of dissolved polyvinyl pyrrolidone, its monomeric compound, tartaric acid and urea, respectively, were investigated. For the characterization of the CD-statin inclusion complexes, stability constants for the associates have been calculated.

  16. Hydropyrolysis of insoluble carbonaceous matter in the Murchison meteorite: new insights into its macromolecular structure

    NASA Astrophysics Data System (ADS)

    Sephton, M. A.; Love, G. D.; Watson, J. S.; Verchovsky, A. B.; Wright, I. P.; Snape, C. E.; Gilmour, I.

    2004-03-01

    The major organic component of carbonaceous chondrites is a solvent-insoluble, high molecular weight macromolecular material that constitutes at least 70% of the total organic content in these meteorites. Analytical pyrolysis is often used to thermally decompose macromolecular organic matter in an inert atmosphere into lower molecular weight fragments that are more amenable to conventional organic analytical techniques. Hydropyrolysis refers to pyrolysis assisted by high hydrogen gas pressures and a dispersed catalytically-active molybdenum sulfide phase. Hydropyrolysis of meteorites has not been attempted previously although it is ideally suited to such studies due to its relatively high yields. Hydropyrolysis of the Murchison macromolecular material successfully releases significant amounts of high molecular weight PAH including phenanthrene, carbazole, fluoranthene, pyrene, chrysene, perylene, benzoperylene and coronene units with varying degrees of alklyation. Analysis of both the products and residue from hydropyrolysis reveals that the meteoritic organic network contains both labile (pyrolysable) and refractory (nonpyrolysable) fractions. Comparisons of hydropyrolysis yields of Murchison macromolecular materials with those from terrestrial coals indicate that the refractory component probably consists of a network dominated by at least five- or six-ring PAH units cross-linked together.

  17. NMR (Nuclear Magnetic Resonance) and macromolecular migration in a melt or in concentrated solutions

    NASA Technical Reports Server (NTRS)

    Addad, J. P. C.

    1983-01-01

    The purpose of this paper is to analyze the migration process of long polymer molecules in a melt or in concentrated solutions as it may be observed from the dynamics of the transverse magnetization of nuclear spins linked to these chains. The low frequency viscoelastic relaxation of polymer systems is known to be mainly controlled by the mechanism of dissociation of topological constraints excited on chains and which are called entanglements. This mechanism exhibits a strong dependence upon the chain molecular weight. These topological constraints also govern the diffusion process of polymer chains. So, the accurate description of the diffusion motion of a chain may be a convenient way to characterize disentanglement processes necessarily involved in any model proposed to explain viscoelastic effects.

  18. Hierarchical amplification of macromolecular helicity of dynamic helical poly(phenylacetylene)s composed of chiral and achiral phenylacetylenes in dilute solution, liquid crystal, and two-dimensional crystal.

    PubMed

    Ohsawa, Sousuke; Sakurai, Shin-ichiro; Nagai, Kanji; Banno, Motonori; Maeda, Katsuhiro; Kumaki, Jiro; Yashima, Eiji

    2011-01-12

    Optically active poly(phenylacetylene) copolymers consisting of optically active and achiral phenylacetylenes bearing L-alanine decyl esters (1L) and 2-aminoisobutylic acid decyl esters (Aib) as the pendant groups (poly(1L(m)-co-Aib(n))) with various compositions were synthesized by the copolymerization of the optically active 1L with achiral Aib using a rhodium catalyst, and their chiral amplification of the macromolecular helicity in a dilute solution, a lyotropic liquid crystalline (LC) state, and a two-dimensional (2D) crystal on the substrate was investigated by measuring the circular dichroism of the copolymers, mesoscopic cholesteric twist in the LC state (cholesteric helical pitch), and high-resolution atomic force microscopy (AFM) images of the self-assembled 2D helix-bundles of the copolymer chains. We found that the macromolecular helicity of poly(1L(m)-co-Aib(n))s could be hierarchically amplified in the order of the dilute solution, LC state, and 2D crystal. In sharp contrast, almost no chiral amplification of the macromolecular helicity was observed for the homopolymer mixtures of 1L and Aib in the LC state and 2D crystal on graphite. PMID:21141965

  19. Protein crystallography for non-crystallographers, or how to get the best (but not more) from published macromolecular structures

    PubMed Central

    Wlodawer, Alexander; Minor, Wladek; Dauter, Zbigniew; Jaskolski, Mariusz

    2015-01-01

    The number of macromolecular structures deposited in the Protein Data Bank now exceeds 45 000, with the vast majority determined using crystallographic methods. Thousands of studies describing such structures have been published in the scientific literature, and 14 Nobel prizes in chemistry or medicine have been awarded to protein crystallographers. As important as these structures are for understanding the processes that take place in living organisms and also for practical applications such as drug design, many non-crystallographers still have problems with critical evaluation of the structural literature data. This review attempts to provide a brief outline of technical aspects of crystallography and to explain the meaning of some parameters that should be evaluated by users of macromolecular structures in order to interpret, but not over-interpret, the information present in the coordinate files and in their description. A discussion of the extent of the information that can be gleaned from the coordinates of structures solved at different resolution, as well as problems and pitfalls encountered in structure determination and interpretation are also covered. PMID:18034855

  20. Sample preparation of biological macromolecular assemblies for the determination of high-resolution structures by cryo-electron microscopy.

    PubMed

    Stark, Holger; Chari, Ashwin

    2016-02-01

    Single particle cryo-EM has recently developed into a powerful tool to determine the 3D structure of macromolecular complexes at near-atomic resolution, which allows structural biologists to build atomic models of proteins. All technical aspects of cryo-EM technology have been considerably improved over the last two decades, including electron microscopic hardware, image processing software and the ever growing speed of computers. This leads to a more widespread use of the technique, and it can be anticipated that further automation of electron microscopes and image processing tools will soon fully shift the focus away from the technological aspects, onto biological questions that can be answered. In single particle cryo-EM, no crystals of a macromolecule are required. In contrast to X-ray crystallography, this significantly facilitates structure determination by cryo-EM. Nevertheless, a relatively high level of biochemical control is still essential to obtain high-resolution structures by cryo-EM, and it can be anticipated that the success of the cryo-EM technology goes hand in hand with further developments of sample purification and preparation techniques. This will allow routine high-resolution structure determination of the many macromolecular complexes of the cell that until now represent evasive targets for X-ray crystallographers. Here we discuss the various biochemical tools that are currently available and the existing sample purification and preparation techniques for cryo-EM grid preparation that are needed to obtain high-resolution images for structure determination. PMID:26671943

  1. Sample preparation of biological macromolecular assemblies for the determination of high-resolution structures by cryo-electron microscopy.

    PubMed

    Stark, Holger; Chari, Ashwin

    2016-02-01

    Single particle cryo-EM has recently developed into a powerful tool to determine the 3D structure of macromolecular complexes at near-atomic resolution, which allows structural biologists to build atomic models of proteins. All technical aspects of cryo-EM technology have been considerably improved over the last two decades, including electron microscopic hardware, image processing software and the ever growing speed of computers. This leads to a more widespread use of the technique, and it can be anticipated that further automation of electron microscopes and image processing tools will soon fully shift the focus away from the technological aspects, onto biological questions that can be answered. In single particle cryo-EM, no crystals of a macromolecule are required. In contrast to X-ray crystallography, this significantly facilitates structure determination by cryo-EM. Nevertheless, a relatively high level of biochemical control is still essential to obtain high-resolution structures by cryo-EM, and it can be anticipated that the success of the cryo-EM technology goes hand in hand with further developments of sample purification and preparation techniques. This will allow routine high-resolution structure determination of the many macromolecular complexes of the cell that until now represent evasive targets for X-ray crystallographers. Here we discuss the various biochemical tools that are currently available and the existing sample purification and preparation techniques for cryo-EM grid preparation that are needed to obtain high-resolution images for structure determination.

  2. Macromolecular Crystallization in Microgravity

    NASA Technical Reports Server (NTRS)

    Snell, Edward H.; Helliwell, John R.

    2004-01-01

    The key concepts that attracted crystal growers, macromolecular or solid state, to microgravity research is that density difference fluid flows and sedimentation of the growing crystals are greatly reduced. Thus, defects and flaws in the crystals can be reduced, even eliminated, and crystal volume can be increased. Macromolecular crystallography differs from the field of crystalline semiconductors. For the latter, crystals are harnessed for their electrical behaviors. A crystal of a biological macromolecule is used instead for diffraction experiments (X-ray or neutron) to determine the three-dimensional structure of the macromolecule. The better the internal order of the crystal of a biological macromolecule then the more molecular structure detail that can be extracted. This structural information that enables an understanding of how the molecule functions. This knowledge is changing the biological and chemical sciences with major potential in understanding disease pathologies. Macromolecular structural crystallography in general is a remarkable field where physics, biology, chemistry, and mathematics meet to enable insight to the basic fundamentals of life. In this review, we examine the use of microgravity as an environment to grow macromolecular crystals. We describe the crystallization procedures used on the ground, how the resulting crystals are studied and the knowledge obtained from those crystals. We address the features desired in an ordered crystal and the techniques used to evaluate those features in detail. We then introduce the microgravity environment, the techniques to access that environment, and the theory and evidence behind the use of microgravity for crystallization experiments. We describe how ground-based laboratory techniques have been adapted to microgravity flights and look at some of the methods used to analyze the resulting data. Several case studies illustrate the physical crystal quality improvements and the macromolecular structural

  3. Effects of macromolecular crowding on nuclear size.

    PubMed

    Rosania, G R; Swanson, J A

    1995-05-01

    The concentration of macromolecules inside cells is high, and the resultant crowding of cytoplasm can be expected to affect many interactions involving macromolecular assemblies. Here, we have examined the effect of solute size and concentration on nuclear volume in saponin-permeabilized macrophages. Nuclei swelled in the presence of small solutes and shrank reversibly in the presence of larger permeant solutes. Remarkably, the smallest solutes capable of shrinking the nucleus were not excluded by the pores in the nuclear envelope. Indeed, nuclei shrank in the presence of such solutes even after the nuclear envelope had been sheared mechanically or permeabilized with detergent. Nuclei extracted with 1% Triton X-100 shrank in the presence of very high concentrations of small solute molecules (30% w/v) as well as in lower concentrations of larger solutes. Consistent with a macromolecular crowding effect, changes in nuclear volume were dependent on solute size and not simply dependent on the colligative properties of solutes or the exclusion of solutes by the nuclear envelope. Solute size-dependent changes in nuclear volume were independent of the chemical nature of the solutes and of the activity of the ions in the buffer. Together, these observations indicate that high concentrations of macromolecules such as those found inside cells can influence the size of the nucleus by directly affecting nuclear structure.

  4. Teaching structure: student use of software tools for understanding macromolecular structure in an undergraduate biochemistry course.

    PubMed

    Jaswal, Sheila S; O'Hara, Patricia B; Williamson, Patrick L; Springer, Amy L

    2013-01-01

    Because understanding the structure of biological macromolecules is critical to understanding their function, students of biochemistry should become familiar not only with viewing, but also with generating and manipulating structural representations. We report a strategy from a one-semester undergraduate biochemistry course to integrate use of structural representation tools into both laboratory and homework activities. First, early in the course we introduce the use of readily available open-source software for visualizing protein structure, coincident with modules on amino acid and peptide bond properties. Second, we use these same software tools in lectures and incorporate images and other structure representations in homework tasks. Third, we require a capstone project in which teams of students examine a protein-nucleic acid complex and then use the software tools to illustrate for their classmates the salient features of the structure, relating how the structure helps explain biological function. To ensure engagement with a range of software and database features, we generated a detailed template file that can be used to explore any structure, and that guides students through specific applications of many of the software tools. In presentations, students demonstrate that they are successfully interpreting structural information, and using representations to illustrate particular points relevant to function. Thus, over the semester students integrate information about structural features of biological macromolecules into the larger discussion of the chemical basis of function. Together these assignments provide an accessible introduction to structural representation tools, allowing students to add these methods to their biochemical toolboxes early in their scientific development.

  5. Macromolecular structures probed by combining single-shot free-electron laser diffraction with synchrotron coherent X-ray imaging.

    PubMed

    Gallagher-Jones, Marcus; Bessho, Yoshitaka; Kim, Sunam; Park, Jaehyun; Kim, Sangsoo; Nam, Daewoong; Kim, Chan; Kim, Yoonhee; Noh, Do Young; Miyashita, Osamu; Tama, Florence; Joti, Yasumasa; Kameshima, Takashi; Hatsui, Takaki; Tono, Kensuke; Kohmura, Yoshiki; Yabashi, Makina; Hasnain, S Samar; Ishikawa, Tetsuya; Song, Changyong

    2014-01-01

    Nanostructures formed from biological macromolecular complexes utilizing the self-assembly properties of smaller building blocks such as DNA and RNA hold promise for many applications, including sensing and drug delivery. New tools are required for their structural characterization. Intense, femtosecond X-ray pulses from X-ray free-electron lasers enable single-shot imaging allowing for instantaneous views of nanostructures at ambient temperatures. When combined judiciously with synchrotron X-rays of a complimentary nature, suitable for observing steady-state features, it is possible to perform ab initio structural investigation. Here we demonstrate a successful combination of femtosecond X-ray single-shot diffraction with an X-ray free-electron laser and coherent diffraction imaging with synchrotron X-rays to provide an insight into the nanostructure formation of a biological macromolecular complex: RNA interference microsponges. This newly introduced multimodal analysis with coherent X-rays can be applied to unveil nano-scale structural motifs from functional nanomaterials or biological nanocomplexes, without requiring a priori knowledge.

  6. Macromolecular structures probed by combining single-shot free-electron laser diffraction with synchrotron coherent X-ray imaging.

    PubMed

    Gallagher-Jones, Marcus; Bessho, Yoshitaka; Kim, Sunam; Park, Jaehyun; Kim, Sangsoo; Nam, Daewoong; Kim, Chan; Kim, Yoonhee; Noh, Do Young; Miyashita, Osamu; Tama, Florence; Joti, Yasumasa; Kameshima, Takashi; Hatsui, Takaki; Tono, Kensuke; Kohmura, Yoshiki; Yabashi, Makina; Hasnain, S Samar; Ishikawa, Tetsuya; Song, Changyong

    2014-01-01

    Nanostructures formed from biological macromolecular complexes utilizing the self-assembly properties of smaller building blocks such as DNA and RNA hold promise for many applications, including sensing and drug delivery. New tools are required for their structural characterization. Intense, femtosecond X-ray pulses from X-ray free-electron lasers enable single-shot imaging allowing for instantaneous views of nanostructures at ambient temperatures. When combined judiciously with synchrotron X-rays of a complimentary nature, suitable for observing steady-state features, it is possible to perform ab initio structural investigation. Here we demonstrate a successful combination of femtosecond X-ray single-shot diffraction with an X-ray free-electron laser and coherent diffraction imaging with synchrotron X-rays to provide an insight into the nanostructure formation of a biological macromolecular complex: RNA interference microsponges. This newly introduced multimodal analysis with coherent X-rays can be applied to unveil nano-scale structural motifs from functional nanomaterials or biological nanocomplexes, without requiring a priori knowledge. PMID:24786694

  7. Teaching Structure: Student Use of Software Tools for Understanding Macromolecular Structure in an Undergraduate Biochemistry Course

    ERIC Educational Resources Information Center

    Jaswal, Sheila S.; O'Hara, Patricia B.; Williamson, Patrick L.; Springer, Amy L.

    2013-01-01

    Because understanding the structure of biological macromolecules is critical to understanding their function, students of biochemistry should become familiar not only with viewing, but also with generating and manipulating structural representations. We report a strategy from a one-semester undergraduate biochemistry course to integrate use of…

  8. Collagen macromolecular drug delivery systems

    SciTech Connect

    Gilbert, D.L.

    1988-01-01

    The objective of this study was to examine collagen for use as a macromolecular drug delivery system by determining the mechanism of release through a matrix. Collagen membranes varying in porosity, crosslinking density, structure and crosslinker were fabricated. Collagen characterized by infrared spectroscopy and solution viscosity was determined to be pure and native. The collagen membranes were determined to possess native vs. non-native quaternary structure and porous vs. dense aggregate membranes by electron microscopy. Collagen monolithic devices containing a model macromolecule (inulin) were fabricated. In vitro release rates were found to be linear with respect to t{sup {1/2}} and were affected by crosslinking density, crosslinker and structure. The biodegradation of the collagen matrix was also examined. In vivo biocompatibility, degradation and {sup 14}C-inulin release rates were evaluated subcutaneously in rats.

  9. Radiation damage and derivatization in macromolecular crystallography: a structure factor’s perspective

    PubMed Central

    Owen, Robin L.; Sherrell, Darren A.

    2016-01-01

    During, or even after, data collection the presence and effects of radiation damage in macromolecular crystallography may not always be immediately obvious. Despite this, radiation damage is almost always present, with site-specific damage occurring on very short time (dose) scales well before global damage becomes apparent. A result of both site-specific radiation damage and derivatization is a change in the relative intensity of reflections. The size and approximate rate of onset of X-ray-induced transformations is compared with the changes expected from derivatization, and strategies for minimizing radiation damage are discussed. PMID:26960125

  10. Single-step antibody-based affinity cryo-electron microscopy for imaging and structural analysis of macromolecular assemblies.

    PubMed

    Yu, Guimei; Vago, Frank; Zhang, Dongsheng; Snyder, Jonathan E; Yan, Rui; Zhang, Ci; Benjamin, Christopher; Jiang, Xi; Kuhn, Richard J; Serwer, Philip; Thompson, David H; Jiang, Wen

    2014-07-01

    Single particle cryo-electron microscopy (cryo-EM) is an emerging powerful tool for structural studies of macromolecular assemblies (i.e., protein complexes and viruses). Although single particle cryo-EM requires less concentrated and smaller amounts of samples than X-ray crystallography, it remains challenging to study specimens that are low-abundance, low-yield, or short-lived. The recent development of affinity grid techniques can potentially further extend single particle cryo-EM to these challenging samples by combining sample purification and cryo-EM grid preparation into a single step. Here we report a new design of affinity cryo-EM approach, cryo-SPIEM, that applies a traditional pathogen diagnosis tool Solid Phase Immune Electron Microscopy (SPIEM) to the single particle cryo-EM method. This approach provides an alternative, largely simplified and easier to use affinity grid that directly works with most native macromolecular complexes with established antibodies, and enables cryo-EM studies of native samples directly from cell cultures. In the present work, we extensively tested the feasibility of cryo-SPIEM with multiple samples including those of high or low molecular weight, macromolecules with low or high symmetry, His-tagged or native particles, and high- or low-yield macromolecules. Results for all these samples (non-purified His-tagged bacteriophage T7, His-tagged Escherichiacoli ribosomes, native Sindbis virus, and purified but low-concentration native Tulane virus) demonstrated the capability of cryo-SPIEM approach in specifically trapping and concentrating target particles on TEM grids with minimal view constraints for cryo-EM imaging and determination of 3D structures.

  11. O the Phase Refinement and Extension of Macromolecular Structures Using both Real and Reciprocal Space Approaches

    NASA Astrophysics Data System (ADS)

    Zhang, Kam Yong Jian

    Available from UMI in association with The British Library. By examining the solution to the phase problem of X-ray crystallography, it is established that the structure factor magnitudes and phases are linked through constraints on the electron density. There are real and reciprocal space approaches to the phase problem depending on the way the constraints on electron density are exploited. A constraint on the electron density--the correct density histogram--is added to the list of other constraints. A new density modification technique--histogram matching --was developed based on the matching of the density histogram to that of the correct one. Its application to 2Zn pig insulin successfully refined and extended the 1.9A MIR phases to 1.5A resolution. In order to obtain a molecular envelope with a detailed boundary, a molecular envelope refinement technique was designed which proved to be quite effective. A gradient technique of defining molecular boundary was also explored and was found to be better than the conventional convolution technique. The two dimensional histogram of density and gradient was examined. It was found that the matching of density histograms also matches that of the gradient histograms. The combination of Sayre's equation with solvent flattening and histogram matching led to a new phase refinement and extension technique--SQUASH. It proved to be a powerful technique by its successful refinement of 3.0A MIR phases of 2Zn pig insulin and subsequent extension to 2.0A resolution.

  12. Comprehensive objective maps of macromolecular conformations by quantitative SAXS analysis

    PubMed Central

    Hura, Greg L.; Budworth, Helen; Dyer, Kevin N.; Rambo, Robert P.; Hammel, Michal

    2013-01-01

    Comprehensive perspectives of macromolecular conformations are required to connect structure to biology. Here we present a small angle X-ray scattering (SAXS) Structural Similarity Map (SSM) and Volatility of Ratio (VR) metric providing comprehensive, quantitative and objective (superposition-independent) perspectives on solution state conformations. We validate VR and SSM utility on human MutSβ, a key ABC ATPase and chemotherapeutic target, by revealing MutSβ DNA sculpting and identifying multiple conformational states for biological activity. PMID:23624664

  13. Space-group and origin ambiguity in macromolecular structures with pseudo-symmetry and its treatment with the program Zanuda.

    PubMed

    Lebedev, Andrey A; Isupov, Michail N

    2014-09-01

    The presence of pseudo-symmetry in a macromolecular crystal and its interplay with twinning may lead to an incorrect space-group (SG) assignment. Moreover, if the pseudo-symmetry is very close to an exact crystallographic symmetry, the structure can be solved and partially refined in the wrong SG. Typically, in such incorrectly determined structures all or some of the pseudo-symmetry operations are, in effect, taken for crystallographic symmetry operations and vice versa. A mistake only becomes apparent when the R(free) ceases to decrease below 0.39 and further model rebuilding and refinement cannot improve the refinement statistics. If pseudo-symmetry includes pseudo-translation, the uncertainty in SG assignment may be associated with an incorrect choice of origin, as demonstrated by the series of examples provided here. The program Zanuda presented in this article was developed for the automation of SG validation. Zanuda runs a series of refinements in SGs compatible with the observed unit-cell parameters and chooses the model with the highest symmetry SG from a subset of models that have the best refinement statistics.

  14. Macromolecular Crowding Modulates Actomyosin Kinetics.

    PubMed

    Ge, Jinghua; Bouriyaphone, Sherry D; Serebrennikova, Tamara A; Astashkin, Andrei V; Nesmelov, Yuri E

    2016-07-12

    Actomyosin kinetics is usually studied in dilute solutions, which do not reflect conditions in the cytoplasm. In cells, myosin and actin work in a dense macromolecular environment. High concentrations of macromolecules dramatically reduce the amount of free space available for all solutes, which results in an effective increase of the solutes' chemical potential and protein stabilization. Moreover, in a crowded solution, the chemical potential depends on the size of the solute, with larger molecules experiencing a larger excluded volume than smaller ones. Therefore, since myosin interacts with two ligands of different sizes (actin and ATP), macromolecular crowding can modulate the kinetics of individual steps of the actomyosin ATPase cycle. To emulate the effect of crowding in cells, we studied actomyosin cycle reactions in the presence of a high-molecular-weight polymer, Ficoll70. We observed an increase in the maximum velocity of the actomyosin ATPase cycle, and our transient-kinetics experiments showed that virtually all individual steps of the actomyosin cycle were affected by the addition of Ficoll70. The observed effects of macromolecular crowding on the myosin-ligand interaction cannot be explained by the increase of a solute's chemical potential. A time-resolved Förster resonance energy transfer experiment confirmed that the myosin head assumes a more compact conformation in the presence of Ficoll70 than in a dilute solution. We conclude that the crowding-induced myosin conformational change plays a major role in the changed kinetics of actomyosin ATPase. PMID:27410745

  15. Statistical mechanics of macromolecular complexation

    NASA Astrophysics Data System (ADS)

    Nakamura, Issei

    The self-assembly of macromolecules through molecular association has attracted long-standing attention in soft-condensed matter physics. The hierarchical formation from small-scale building blocks into larger-scale complex structures often leads to very rich phase behavior controlled by various ambient conditions. The understanding and control of the phase behavior of self-assembling systems require detailed knowledge about the entropy and enthalpy contributions to the free energy of the system. However, this knowledge is limited at the present time because a comprehensive theoretical description of molecular association is still lacking. In this thesis, four tales of achievements in developing theories of macromolecular complexation are presented. (1) We begin with an analytically solvable model of the self-assembly of rigid macromolecules with surface adsorption. A generic understanding of the driving force and the role of entropy is obtained from the exact solutions. (2) We move on to further development of the theory in order to study the complexation between polymers and ionic molecules. The extension of the first model to chain-like molecules is performed using a well-established method in polymer physics, the self-consistent field theory (SCFT) of polymers. We also discuss gelation in this system within the scope of mean-filed approximations. (3) Then, a ladder-like polymer-polymer complexation is studied. Unconventional phase diagrams are predicted from the modified SCFT, indicating a large effect of variations in entropy due to the complexation on bulk properties. (4) Finally, the kinetic aspect of macromolecular binding reactions is discussed.

  16. Facilitating best practices in collecting anomalous scattering data for de novo structure solution at the ESRF Structural Biology Beamlines

    PubMed Central

    de Sanctis, Daniele; Oscarsson, Marcus; Popov, Alexander; Svensson, Olof; Leonard, Gordon

    2016-01-01

    The constant evolution of synchrotron structural biology beamlines, the viability of screening protein crystals for a wide range of heavy-atom derivatives, the advent of efficient protein labelling and the availability of automatic data-processing and structure-solution pipelines have combined to make de novo structure solution in macromolecular crystallography a less arduous task. Nevertheless, the collection of diffraction data of sufficient quality for experimental phasing is still a difficult and crucial step. Here, some examples of good data-collection practice for projects requiring experimental phasing are presented and recent developments at the ESRF Structural Biology beamlines that have facilitated these are illustrated. PMID:26960128

  17. Facilitating best practices in collecting anomalous scattering data for de novo structure solution at the ESRF Structural Biology Beamlines.

    PubMed

    de Sanctis, Daniele; Oscarsson, Marcus; Popov, Alexander; Svensson, Olof; Leonard, Gordon

    2016-03-01

    The constant evolution of synchrotron structural biology beamlines, the viability of screening protein crystals for a wide range of heavy-atom derivatives, the advent of efficient protein labelling and the availability of automatic data-processing and structure-solution pipelines have combined to make de novo structure solution in macromolecular crystallography a less arduous task. Nevertheless, the collection of diffraction data of sufficient quality for experimental phasing is still a difficult and crucial step. Here, some examples of good data-collection practice for projects requiring experimental phasing are presented and recent developments at the ESRF Structural Biology beamlines that have facilitated these are illustrated.

  18. Pathological Crystallography: Case Studies of Several Unusual Macromolecular Crystals

    SciTech Connect

    Dauter,Z.; Botos, I.; LaRonde-LeBlanc, N.; Wlodawer, A.

    2005-01-01

    Although macromolecular crystallography is rapidly becoming largely routine owing to advances in methods of data collection, structure solution and refinement, difficult cases are still common. To remind structural biologists about the kinds of crystallographic difficulties that might be encountered, case studies of several successfully completed structure determinations that utilized less than perfect crystals are discussed here. The structure of the proteolytic domain of Archaeoglobus fulgidus Lon was solved with crystals that contained superimposed orthorhombic and monoclinic lattices, a case not previously described for proteins. Another hexagonal crystal form of this protein exhibited an unusually high degree of non-isomorphism. Crystals of A. fulgidus Rio1 kinase exhibited both pseudosymmetry and twinning. Ways of identifying the observed phenomena and approaches to solving and refining macromolecular structures when only less than perfect crystals are available are discussed here.

  19. Three dimensional electron microscopy and in silico tools for macromolecular structure determination

    PubMed Central

    Borkotoky, Subhomoi; Meena, Chetan Kumar; Khan, Mohammad Wahab; Murali, Ayaluru

    2013-01-01

    Recently, structural biology witnessed a major tool - electron microscopy - in solving the structures of macromolecules in addition to the conventional techniques, X-ray crystallography and nuclear magnetic resonance (NMR). Three dimensional transmission electron microscopy (3DTEM) is one of the most sophisticated techniques for structure determination of molecular machines. Known to give the 3-dimensional structures in its native form with literally no upper limit on size of the macromolecule, this tool does not need the crystallization of the protein. Combining the 3DTEM data with in silico tools, one can have better refined structure of a desired complex. In this review we are discussing about the recent advancements in three dimensional electron microscopy and tools associated with it. PMID:27092033

  20. Structural Studies on Membrane Proteins and Biological Macromolecular Assemblies in Japan

    NASA Astrophysics Data System (ADS)

    Tsukihara, Tomitake

    Structural studies on membrane proteins have been performed at atomic level by both three-dimensional X-ray crystallography and two-dimensional electron crystallography in Japan as in Europe and Unites States. More than 13 membrane protein structures were elucidate by X-ray method in our country, and seven membrane protein structures were determined by cryo-electron microscopic method developed by Fujiyoshi of Kyoto University. Extensive crystallographic studies on calcium pump and cytochrome c oxidase elucidated their functional mechanisms at atomic level. Structure and switching mechanism of a flagellum were studied by X-ray and electron microscopic methods. Vault structure exhibiting D39 symmetry was determined by X-ray method.

  1. Teaching macromolecular modeling.

    PubMed

    Harvey, S C; Tan, R K

    1992-12-01

    Training newcomers to the field of macromolecular modeling is as difficult as is training beginners in x-ray crystallography, nuclear magnetic resonance, or other methods in structural biology. In one or two lectures, the most that can be conveyed is a general sense of the relationship between modeling and other structural methods. If a full semester is available, then students can be taught how molecular structures are built, manipulated, refined, and analyzed on a computer. Here we describe a one-semester modeling course that combines lectures, discussions, and a laboratory using a commercial modeling package. In the laboratory, students carry out prescribed exercises that are coordinated to the lectures, and they complete a term project on a modeling problem of their choice. The goal is to give students an understanding of what kinds of problems can be attacked by molecular modeling methods and which problems are beyond the current capabilities of those methods.

  2. Automated refinement of macromolecular structures at low resolution using prior information

    PubMed Central

    Kovalevskiy, Oleg; Nicholls, Robert A.; Murshudov, Garib N.

    2016-01-01

    Since the ratio of the number of observations to adjustable parameters is small at low resolution, it is necessary to use complementary information for the analysis of such data. ProSMART is a program that can generate restraints for macromolecules using homologous structures, as well as generic restraints for the stabilization of secondary structures. These restraints are used by REFMAC5 to stabilize the refinement of an atomic model. However, the optimal refinement protocol varies from case to case, and it is not always obvious how to select appropriate homologous structure(s), or other sources of prior information, for restraint generation. After running extensive tests on a large data set of low-resolution models, the best-performing refinement protocols and strategies for the selection of homologous structures have been identified. These strategies and protocols have been implemented in the Low-Resolution Structure Refinement (LORESTR) pipeline. The pipeline performs auto-detection of twinning and selects the optimal scaling method and solvent parameters. LORESTR can either use user-supplied homologous structures, or run an automated BLAST search and download homologues from the PDB. The pipeline executes multiple model-refinement instances using different parameters in order to find the best protocol. Tests show that the automated pipeline improves R factors, geometry and Ramachandran statistics for 94% of the low-resolution cases from the PDB included in the test set. PMID:27710936

  3. Designs for the self-assembly of open and closed macromolecular structures and a molecular switch using DNA methyltransferases to order proteins on nucleic acid scaffolds

    NASA Astrophysics Data System (ADS)

    Smith, Steven S.

    2002-06-01

    The methyltransferase-directed addressing of fusion proteins to DNA scaffolds offers an approach to the construction of protein/nucleic acid biostructures with potential in a variety of applications. The technology is currently only limited by the yield of high occupancy structures. However, current evidence shows that DNA scaffolds that contain three or four targeted proteins can be reliably constructed. This permits a variety of macromolecular designs, several of which are given in this paper. Designs for open and closed two-dimensional and three-dimensional assemblies and a design for a molecular switch are discussed. The closed two-dimensional assembly takes the form of a square, and could find application as a component of other systems including a macromolecular rotaxane. The closed three-dimensional system takes the form of a trigonal bipyramid and could find application as a macromolecular carcerand. The molecular switch could find application as a peptide biosensor. Guidelines for the construction and structural verification of these designs are reported.

  4. Avoidable errors in deposited macromolecular structures: an impediment to efficient data mining

    PubMed Central

    Dauter, Zbigniew; Wlodawer, Alexander; Minor, Wladek; Jaskolski, Mariusz; Rupp, Bernhard

    2014-01-01

    Whereas the vast majority of the more than 85 000 crystal structures of macromolecules currently deposited in the Protein Data Bank are of high quality, some suffer from a variety of imperfections. Although this fact has been pointed out in the past, it is still worth periodic updates so that the metadata obtained by global analysis of the available crystal structures, as well as the utilization of the individual structures for tasks such as drug design, should be based on only the most reliable data. Here, selected abnormal deposited structures have been analysed based on the Bayesian reasoning that the correctness of a model must be judged against both the primary evidence as well as prior knowledge. These structures, as well as information gained from the corresponding publications (if available), have emphasized some of the most prevalent types of common problems. The errors are often perfect illustrations of the nature of human cognition, which is frequently influenced by preconceptions that may lead to fanciful results in the absence of proper validation. Common errors can be traced to negligence and a lack of rigorous verification of the models against electron density, creation of non-parsimonious models, generation of improbable numbers, application of incorrect symmetry, illogical presentation of the results, or violation of the rules of chemistry and physics. Paying more attention to such problems, not only in the final validation stages but during the structure-determination process as well, is necessary not only in order to maintain the highest possible quality of the structural repositories and databases but most of all to provide a solid basis for subsequent studies, including large-scale data-mining projects. For many scientists PDB deposition is a rather infrequent event, so the need for proper training and supervision is emphasized, as well as the need for constant alertness of reason and critical judgment as absolutely necessary safeguarding

  5. Avoidable errors in deposited macromolecular structures: an impediment to efficient data mining.

    PubMed

    Dauter, Zbigniew; Wlodawer, Alexander; Minor, Wladek; Jaskolski, Mariusz; Rupp, Bernhard

    2014-05-01

    Whereas the vast majority of the more than 85 000 crystal structures of macromolecules currently deposited in the Protein Data Bank are of high quality, some suffer from a variety of imperfections. Although this fact has been pointed out in the past, it is still worth periodic updates so that the metadata obtained by global analysis of the available crystal structures, as well as the utilization of the individual structures for tasks such as drug design, should be based on only the most reliable data. Here, selected abnormal deposited structures have been analysed based on the Bayesian reasoning that the correctness of a model must be judged against both the primary evidence as well as prior knowledge. These structures, as well as information gained from the corresponding publications (if available), have emphasized some of the most prevalent types of common problems. The errors are often perfect illustrations of the nature of human cognition, which is frequently influenced by preconceptions that may lead to fanciful results in the absence of proper validation. Common errors can be traced to negligence and a lack of rigorous verification of the models against electron density, creation of non-parsimonious models, generation of improbable numbers, application of incorrect symmetry, illogical presentation of the results, or violation of the rules of chemistry and physics. Paying more attention to such problems, not only in the final validation stages but during the structure-determination process as well, is necessary not only in order to maintain the highest possible quality of the structural repositories and databases but most of all to provide a solid basis for subsequent studies, including large-scale data-mining projects. For many scientists PDB deposition is a rather infrequent event, so the need for proper training and supervision is emphasized, as well as the need for constant alertness of reason and critical judgment as absolutely necessary safeguarding

  6. Cooperative macromolecular device revealed by meta-analysis of static and time-resolved structures

    PubMed Central

    Ren, Zhong; Šrajer, Vukica; Knapp, James E.; Royer, William E.

    2012-01-01

    Here we present a meta-analysis of a large collection of static structures of a protein in the Protein Data Bank in order to extract the progression of structural events during protein function. We apply this strategy to the homodimeric hemoglobin HbI from Scapharca inaequivalvis. We derive a simple dynamic model describing how binding of the first ligand in one of the two chemically identical subunits facilitates a second binding event in the other partner subunit. The results of our ultrafast time-resolved crystallographic studies support this model. We demonstrate that HbI functions like a homodimeric mechanical device, such as pliers or scissors. Ligand-induced motion originating in one subunit is transmitted to the other via conserved pivot points, where the E and F′ helices from two partner subunits are “bolted” together to form a stable dimer interface permitting slight relative rotation but preventing sliding. PMID:22171006

  7. Cooperative macromolecular device revealed by meta-analysis of static and time-resolved structures

    SciTech Connect

    Ren, Zhong; Šrajer, Vukica; Knapp, James E.; Royer, Jr., William E.

    2013-04-08

    Here we present a meta-analysis of a large collection of static structures of a protein in the Protein Data Bank in order to extract the progression of structural events during protein function. We apply this strategy to the homodimeric hemoglobin HbI from Scapharca inaequivalvis. We derive a simple dynamic model describing how binding of the first ligand in one of the two chemically identical subunits facilitates a second binding event in the other partner subunit. The results of our ultrafast time-resolved crystallographic studies support this model. We demonstrate that HbI functions like a homodimeric mechanical device, such as pliers or scissors. Ligand-induced motion originating in one subunit is transmitted to the other via conserved pivot points, where the E and F' helices from two partner subunits are 'bolted' together to form a stable dimer interface permitting slight relative rotation but preventing sliding.

  8. Macromolecular recognition: Structural aspects of the origin of the genetic system

    NASA Technical Reports Server (NTRS)

    Rein, Robert; Sokalski, W. Andrzej; Barak, Dov; Luo, Ning; Zielinski, Theresa Julia; Shibata, Masayuki

    1991-01-01

    Theoretical simulation of prebiotic chemical processes is an invaluable tool for probing the phenomenon of the evolution of life. Using computational and modeling techniques and guided by analogies from present day systems, we seek to understand the emergence of the genetic apparatus, enzymatic catalysis and protein synthesis under prebiotic conditions. Modeling of the ancestral aminoacyl-tRNA-synthetases (aRS) may provide important clues to the emergence of the genetic code and the protein synthetic machinery. The minimal structural requirements for the catalysis of tRNA aminoacylation are being explored. A formation of an aminoacyl adenylate was studied in the framework of ab initio molecular orbital theory. The role of individual residues in the vicinity of the TyrRS active site was examined, and the effect of all possible amino acids substitutions near the active site was examined. A formation of aminoacyl tRNA was studied by the molecular modeling system SYBYL with the high resolution crystallographic structures of the present day tRNA, aRS's complexes. The ultimate goal is to propose a simple RNA segment that is small enough to be build in the primordial chemical environment but maintains the specificity and catalytic activity of the contemporary RNA enzyme. To understand the mechanism of ribozyme catalyzed reactions, ab initio and semi-empirical (ZINDO) programs were used to investigate the reaction path of transphosphorylation. A special emphasis was placed on the possible catalytic and structural roles played by the coordinated magnesium cation. Both the inline and adjacent mechanisms of transphosphorylation were studied. The structural characteristics of the target helices, particularly a possible role for the G-T pair, is also studied by a molecular dynamics (MD) simulation technique.

  9. NaOH treatment of chitosan films: Impact on macromolecular structure and film properties.

    PubMed

    Takara, E A; Marchese, J; Ochoa, N A

    2015-11-01

    In this paper, we examine the significance of treatment with NaOH on chitosan (CH) film structure to obtain biodegradable materials for several applications. In order to determine the structure of the films, an analysis based on SEM, FTIR spectroscopy and X-ray diffraction data was performed. In addition, the consequences of this treatment were evaluated by swelling index measurements and mechanical testing. As result of FTIR and X-ray analysis, three effects were identified: the deprotonation and phosphate extraction, which allowed new hydrogen bonds to form, and a higher CH deacetylation. These studies also revealed that two hydrated and anhydrous polymorphs were present in the CH-NaOH films. Moreover, the new hydrogen bond and the reduction of N-acetyl groups produced films with a more compact and disordered structure, reducing their swelling characteristics and increasing their brittleness. The introduction of a mild NaOH treatment is a versatile tool to obtain chitosan films with interesting and tunable properties.

  10. Macromolecular recognition: Structural aspects of the origin of the genetic system

    NASA Technical Reports Server (NTRS)

    Rein, Robert; Barak, Dov; Luo, Ning; Zielinski, Theresa Julia; Shibata, Masayuki

    1991-01-01

    Theoretical simulation of prebiotic chemical processes is an invaluable tool for probing the phenomenon of evolution of life. Using computational and modeling techniques and guided by analogies from present day systems we, seek to understand the emergence of genetic apparatus, enzymatic catalysis and protein synthesis under prebiotic conditions. In one possible scenario, the RNA enzymatic reaction plays a key role in the emergence of the self-replicating and offers a clue to the onset of enzymatic catalysis prior to the existence of the protein biosynthetic machinery. Our ultimate goal is to propose a simple RNA segment which contains the specificity and catalytic activity of the contemporary RNA enzyme and which could emerge in a primordial chemical environment. To understand the mechanism of ribozyme catalyzed reactions, ab initio and semi-empirical (ZINDO) programs were used to investigate the reaction path of transphosphorylation. A special emphasis was placed on the possible catalytic and structural roles played by the coordinated magnesium cation. Both the inline and adjacent mechanisms of transphosphorylation have been studied. Another important aspect of this reaction is the identity of the functional groups which are essential for the acid base catalysis. The structural characteristics of the target helices, particularly a possible role of G center dot T pair, is under examination by molecular dynamics (MD) simulation technique. Modeling of the ancestral aminoacyl-tRNA synthetases (aRS) may provide important clues to the emergence of the genetic code and the protein synthetic machinery. Assuming that the catalytic function evolved before the elements of specific recognition of a particular amino acid, we are exploring the minimal structural requirements for the catalysis of tRNA aminoacylation. The molecular modeling system SYBYL was used for this study based on the high resolution crystallographic structures of the present day tyrosyl-adenylate:tyrRS and

  11. Effects of macromolecular crowding on the structural stability of human α-lactalbumin.

    PubMed

    Zhang, De-Lin; Wu, Ling-Jia; Chen, Jie; Liang, Yi

    2012-08-01

    The folding of protein, an important process for protein to fulfill normal functions, takes place in crowded physiological environments. α-Lactalbumin, as a model system for protein-folding studies, has been used extensively because it can form stable molten globule states under a range of conditions. Here we report that the crowding agents Ficoll 70, dextran 70, and polyethylene glycol (PEG) 2000 have different effects on the structural stability of human α-lactalbumin (HLA) represented by the transition to a molten globule state: dextran 70 dramatically enhances the thermal stability of Ca(2+)-depleted HLA (apo-HLA) and Ficoll 70 enhances the thermal stability of apo-HLA to some extent, while PEG 2000 significantly decreases the thermal stability of apo-HLA. Ficoll 70 and dextran 70 have no obvious effects on trypsin degradation of apo-HLA but PEG 2000 accelerates apo-HLA degradation by trypsin and destabilizes the native conformation of apo-HLA. Furthermore, no interaction is observed between apo-HLA and Ficoll 70 or dextran 70, but a weak, non-specific interaction between the apo form of the protein and PEG 2000 is detected, and such a weak, non-specific interaction could overcome the excluded-volume effect of PEG 2000. Our data are consistent with the results of protein stability studies in cells and suggest that stabilizing excluded-volume effects of crowding agents can be ameliorated by non-specific interactions between proteins and crowders. PMID:22735492

  12. Emerging applications of small angle solution scattering in structural biology

    PubMed Central

    Chaudhuri, Barnali N

    2015-01-01

    Small angle solution X-ray and neutron scattering recently resurfaced as powerful tools to address an array of biological problems including folding, intrinsic disorder, conformational transitions, macromolecular crowding, and self or hetero-assembling of biomacromolecules. In addition, small angle solution scattering complements crystallography, nuclear magnetic resonance spectroscopy, and other structural methods to aid in the structure determinations of multidomain or multicomponent proteins or nucleoprotein assemblies. Neutron scattering with hydrogen/deuterium contrast variation, or X-ray scattering with sucrose contrast variation to a certain extent, is a convenient tool for characterizing the organizations of two-component systems such as a nucleoprotein or a lipid-protein assembly. Time-resolved small and wide-angle solution scattering to study biological processes in real time, and the use of localized heavy-atom labeling and anomalous solution scattering for applications as FRET-like molecular rulers, are amongst promising newer developments. Despite the challenges in data analysis and interpretation, these X-ray/neutron solution scattering based approaches hold great promise for understanding a wide variety of complex processes prevalent in the biological milieu. PMID:25516491

  13. Harvesting and cryo-cooling crystals of membrane proteins grown in lipidic mesophases for structure determination by macromolecular crystallography.

    PubMed

    Li, Dianfan; Boland, Coilín; Aragao, David; Walsh, Kilian; Caffrey, Martin

    2012-09-02

    An important route to understanding how proteins function at a mechanistic level is to have the structure of the target protein available, ideally at atomic resolution. Presently, there is only one way to capture such information as applied to integral membrane proteins (Figure 1), and the complexes they form, and that method is macromolecular X-ray crystallography (MX). To do MX diffraction quality crystals are needed which, in the case of membrane proteins, do not form readily. A method for crystallizing membrane proteins that involves the use of lipidic mesophases, specifically the cubic and sponge phases(1-5), has gained considerable attention of late due to the successes it has had in the G protein-coupled receptor field(6-21) (www.mpdb.tcd.ie). However, the method, henceforth referred to as the in meso or lipidic cubic phase method, comes with its own technical challenges. These arise, in part, due to the generally viscous and sticky nature of the lipidic mesophase in which the crystals, which are often micro-crystals, grow. Manipulating crystals becomes difficult as a result and particularly so during harvesting(22,23). Problems arise too at the step that precedes harvesting which requires that the glass sandwich plates in which the crystals grow (Figure 2)(24,25) are opened to expose the mesophase bolus, and the crystals therein, for harvesting, cryo-cooling and eventual X-ray diffraction data collection. The cubic and sponge mesophase variants (Figure 3) from which crystals must be harvested have profoundly different rheologies(4,26). The cubic phase is viscous and sticky akin to a thick toothpaste. By contrast, the sponge phase is more fluid with a distinct tendency to flow. Accordingly, different approaches for opening crystallization wells containing crystals growing in the cubic and the sponge phase are called for as indeed different methods are required for harvesting crystals from the two mesophase types. Protocols for doing just that have been

  14. A predictive aggregate transport model for microfiltration of combined macromolecular solutions and poly-disperse suspensions: model development.

    PubMed

    Baruah, Gautam Lal; Belfort, Georges

    2003-01-01

    A methodology, called the aggregate transport model, is presented that can a priori predict both the pressure-independent permeation flux and yield of target species for the microfiltration of poly-disperse solutions. The model captures the phenomenon of critical shear rate. Beyond the critical shear rate (expressed as a ratio of shear rate to permeation flux), the transmission of proteins drops sharply as a result of cake classification. The widely reported benefits of operating at uniform transmembrane pressure and constant wall concentration follow from this method. The methodology is general in nature and can be used predictively to obtain an optimal balance between flux and yield of target species during the microfiltration of many commercial poly-disperse suspensions. In the accompanying paper we test this model for microfiltration of transgenic whole goat milk.

  15. A Model for Macromolecular Crystallization

    NASA Technical Reports Server (NTRS)

    Pusey, Marc L.; Whitaker, Ann F. (Technical Monitor)

    2001-01-01

    Macromolecular crystallization is a complex process. involving a system which typically has 5 or more components (macromolecule, water, buffer + counter ion, and precipitant). Whereas small molecules have only several well defined contacts in the crystal lattice, macromolecules generally have 10's or even 100's of contacts between molecules. These can range from hydrogen bonds (direct or water-mediated), through van der Waals, hydrophobic, salt bridges, and ion-mediated contacts. The latter interactions are stronger and require some specificity in the molecular alignment, while the others are weaker, more prevalent, and more promiscuous, i.e., can often be readily broken and reformed between other sites. Formation of a consistent, ordered, 3D structure may be impossible in the absence of any or presence of too many strong interactions. Further complicating the process is the inherent structural asymmetry of monomeric single chain macromolecules. The process of crystal nucleation and growth involves the ordered assembly of growth units into a defined 3D lattice. We suggest that for many macromolecules, particularly those that are monomeric, this involves a preliminary solution-phase assembly process into a growth unit having some symmetry prior to addition to the lattice, recapitulating the initial stages of the nucleation process. If this model is correct then fluids and crystal growth models assuming a strictly monodisperse nutrient solution need to be revised. Experimental evidence, based upon face growth rate, AFM, and fluorescence energy transfer data, for a postulated model of the nucleation of tetragonal lysozyme crystals and how it transitions into crystal growth will be presented.

  16. Visualizing Macromolecular Complexes with In Situ Liquid Scanning Transmission Electron Microscopy

    SciTech Connect

    Evans, James E.; Jungjohann, K. L.; Wong, Peony C. K.; Chiu, Po-Lin; Dutrow, Gavin H.; Arslan, Ilke; Browning, Nigel D.

    2012-11-01

    A central focus of biological research is understanding the structure/function relationship of macromolecular protein complexes. Yet conventional transmission electron microscopy techniques are limited to static observations. Here we present the first direct images of purified macromolecular protein complexes using in situ liquid scanning transmission electron microscopy. Our results establish the capability of this technique for visualizing the interface between biology and nanotechnology with high fidelity while also probing the interactions of biomolecules within solution. This method represents an important advancement towards allowing future high-resolution observations of biological processes and conformational dynamics in real-time.

  17. Visualizing macromolecular complexes with in situ liquid scanning transmission electron microscopy.

    PubMed

    Evans, James E; Jungjohann, Katherine L; Wong, Peony C K; Chiu, Po-Lin; Dutrow, Gavin H; Arslan, Ilke; Browning, Nigel D

    2012-11-01

    A central focus of biological research is understanding the structure/function relationship of macromolecular protein complexes. Yet conventional transmission electron microscopy techniques are limited to static observations. Here we present the first direct images of purified macromolecular protein complexes using in situ liquid scanning transmission electron microscopy. Our results establish the capability of this technique for visualizing the interface between biology and nanotechnology with high fidelity while also probing the interactions of biomolecules within solution. This method represents an important advancement towards allowing future high-resolution observations of biological processes and conformational dynamics in real-time.

  18. Myochrysine Solution Structure and Reactivity

    PubMed Central

    Jones, William B.; Zhao, Zheng; Dorsey, John G.; Tepperman, Katherine

    1994-01-01

    We have determined the framework structure of Myochrysine (disodium gold(I)thiomalate) in the solid state and extremely concentrated aqueous solution, previously. It consists of an open chain polymer with linear gold coordination to two thiolates from the thiomalic acid moieties which bridge between pairs of gold atoms providing an Au-S-Au angle of 95°. The question remained: was this structure relevant to the dilute solutions of drugs administered and the still lower concentrations of gold found in the bodies of patients (typically 1 ppm Au in blood and urine or 5 μM in Au). We have provided an answer to that question using extended X-ray absorption spectroscopy (EXAFS) and capillary zone electrophoresis (CZE). EXAFS studies confirm that the polymeric structure with two sulfur atoms per gold atom persists from molar concentrations down to millimolar concentrations. CZE is able to separate and detect Myochrysine at millimolar levels. More importantly, at micromolar levels Myochrysine solutions exhibit identical CZE behavior to that measured at millimolar levels. Thus, aqueous solutions of the drug remain oligomeric at concentrations commensurate with those found in patient blood and urine. The reactivity of Myochrysine with cyanide, a species especially prevalent in smoking patients, was explored using CZE. Cyanide freely replaces thiomalic acid to form [Au(CN)2]- and thiomalic acid via a mixed ligand intermediate. The overall apparent equilibrium constant (Kapp) for the reaction is 6×10-4M-1. Further reaction of [Au(CN)2]- with a large excess of L, where L is cysteine, N-acetylcysteine, or glutathione, shows that these amino acids readily replace cyanide to form [AuL2]-. These species are thus potential metabolites and could possibly be active forms of gold in vivo. That all of these species are readily separated and quantified using CZE demonstrates that capillary electrophoresis is an accessible and powerful tool to add to those used for the study of gold

  19. What Macromolecular Crowding Can Do to a Protein

    PubMed Central

    Kuznetsova, Irina M.; Turoverov, Konstantin K.; Uversky, Vladimir N.

    2014-01-01

    The intracellular environment represents an extremely crowded milieu, with a limited amount of free water and an almost complete lack of unoccupied space. Obviously, slightly salted aqueous solutions containing low concentrations of a biomolecule of interest are too simplistic to mimic the “real life” situation, where the biomolecule of interest scrambles and wades through the tightly packed crowd. In laboratory practice, such macromolecular crowding is typically mimicked by concentrated solutions of various polymers that serve as model “crowding agents”. Studies under these conditions revealed that macromolecular crowding might affect protein structure, folding, shape, conformational stability, binding of small molecules, enzymatic activity, protein-protein interactions, protein-nucleic acid interactions, and pathological aggregation. The goal of this review is to systematically analyze currently available experimental data on the variety of effects of macromolecular crowding on a protein molecule. The review covers more than 320 papers and therefore represents one of the most comprehensive compendia of the current knowledge in this exciting area. PMID:25514413

  20. Crystal structure of Jararacussin-I: the highly negatively charged catalytic interface contributes to macromolecular selectivity in snake venom thrombin-like enzymes.

    PubMed

    Ullah, A; Souza, T A C B; Zanphorlin, L M; Mariutti, R B; Santana, V S; Murakami, M T; Arni, R K

    2013-01-01

    Snake venom serine proteinases (SVSPs) are hemostatically active toxins that perturb the maintenance and regulation of both the blood coagulation cascade and fibrinolytic feedback system at specific points, and hence, are widely used as tools in pharmacological and clinical diagnosis. The crystal structure of a thrombin-like enzyme (TLE) from Bothrops jararacussu venom (Jararacussin-I) was determined at 2.48 Å resolution. This is the first crystal structure of a TLE and allows structural comparisons with both the Agkistrodon contortrix contortrix Protein C Activator and the Trimeresurus stejnegeri plasminogen activator. Despite the highly conserved overall fold, significant differences in the amino acid compositions and three-dimensional conformations of the loops surrounding the active site significantly alter the molecular topography and charge distribution profile of the catalytic interface. In contrast to other SVSPs, the catalytic interface of Jararacussin-I is highly negatively charged, which contributes to its unique macromolecular selectivity. PMID:23139169

  1. Transition state structures in solution

    SciTech Connect

    Bertran, J.; Lluch, J. M.; Gonzalez-Lafont, A.; Dillet, V.; Perez, V.

    1995-04-05

    In the present paper the location of transition state structures for reactions in solution has been studied. Continuum model calculations have been carried out on the Friedel-Crafts alkylation reaction and a proton transfer through a water molecule between two oxygen atoms in formic acid. In this model the separation between the chemical system and the solvent has been introduced. On the other hand, the discrete Monte Carlo methodology has also been used to simulate the solvent effect on dissociative electron transfer processes. In this model, the hypothesis of separability is not assumed. Finally, the validity of both approaches is discussed.

  2. Transition state structures in solution

    NASA Astrophysics Data System (ADS)

    Bertrán, J.; Lluch, J. M.; Gonzàlez-Lafont, A.; Dillet, V.; Pérez, V.

    1995-04-01

    In the present paper the location of transition state structures for reactions in solution has been studied. Continuum model calculations have been carried out on the Friedel-Crafts alkylation reaction and a proton transfer through a water molecule between two oxygen atoms in formic acid. In this model the separation between the chemical system and the solvent has been introduced. On the other hand, the discrete Monte Carlo methodology has also been used to simulate the solvent effect on dissociative electron transfer processes. In this model, the hypothesis of separability is not assumed. Finally, the validity of both approaches is discussed.

  3. Myochrysine solution structure and reactivity.

    PubMed

    Elder, R C; Jones, W B; Zhao, Z; Dorsey, J G; Tepperman, K

    1994-01-01

    We have determined the framework structure of Myochrysine (disodium gold(I)thiomalate) in the solid state and extremely concentrated aqueous solution, previously. It consists of an open chain polymer with linear gold coordination to two thiolates from the thiomalic acid moieties which bridge between pairs of gold atoms providing an Au-S-Au angle of 95 degrees . The question remained: was this structure relevant to the dilute solutions of drugs administered and the still lower concentrations of gold found in the bodies of patients (typically 1 ppm Au in blood and urine or 5 muM in Au). We have provided an answer to that question using extended X-ray absorption spectroscopy (EXAFS) and capillary zone electrophoresis (CZE). EXAFS studies confirm that the polymeric structure with two sulfur atoms per gold atom persists from molar concentrations down to millimolar concentrations. CZE is able to separate and detect Myochrysine at millimolar levels. More importantly, at micromolar levels Myochrysine solutions exhibit identical CZE behavior to that measured at millimolar levels. Thus, aqueous solutions of the drug remain oligomeric at concentrations commensurate with those found in patient blood and urine.The reactivity of Myochrysine with cyanide, a species especially prevalent in smoking patients, was explored using CZE. Cyanide freely replaces thiomalic acid to form [Au(CN)(2)](-) and thiomalic acid via a mixed ligand intermediate. The overall apparent equilibrium constant (K(app)) for the reaction is 6x10(-4)M(-1). Further reaction of [Au(CN)(2)](-) with a large excess of L, where L is cysteine, N-acetylcysteine, or glutathione, shows that these amino acids readily replace cyanide to form [AuL(2)](-). These species are thus potential metabolites and could possibly be active forms of gold in vivo. That all of these species are readily separated and quantified using CZE demonstrates that capillary electrophoresis is an accessible and powerful tool to add to those used

  4. Macromolecular self-assembly and nanotechnology in China.

    PubMed

    Xu, Huaping; Chen, Daoyong; Wang, Shu; Zhou, Yongfeng; Sun, Junqi; Zhang, Wenke; Zhang, Xi

    2013-10-13

    Macromolecular self-assembly refers to the assembly of synthetic polymers, biomacromolecules and supra-molecular polymers. Through macromolecular self-assembly, the fabrication of ordered structures at different scales, the control of the dynamic assembly process and the integrations of advanced functions can be realized. Macromolecular self-assembly and nanotechnology research in China has developed rapidly, from the early periods of follow-up at low to high level and progress into a stage of innovation and creation. This review selects some representative progresses achieved recently, aiming to reflect the current status of macromolecular self-assembly and nanotechnology research in China.

  5. Structure of supersaturated zincate solutions

    SciTech Connect

    Dmitrenko, V.E.; Balyakina, N.N.; Baulov, V.I.; Kotov, A.V.; Zubov, M.S.

    1985-09-01

    During the discharge of chemical power sources with zinc electrodes, supersaturated zincate solution (SZS) is formed from which zinc oxide or hydroxide precipitates as a function of time. The deposit detracts from the functioning of these power sources. In view of the model suggested for the structure of SZS, it is expected that a stabilizing effect would be exerted on SZS by compounds having proton-donating groups which do not give off the protons in the strongly alkaline medium and are not discharged in this medium. For a check of this, the authors chose to use xylitol and molasses in their experiments. The SZS were produced with a mock-up silver-zinc battery using the procedure previously described.

  6. Continuous mutual improvement of macromolecular structure models in the PDB and of X-ray crystallographic software: the dual role of deposited experimental data

    SciTech Connect

    Terwilliger, Thomas C.; Bricogne, Gerard

    2014-10-01

    Macromolecular structures deposited in the PDB can and should be continually reinterpreted and improved on the basis of their accompanying experimental X-ray data, exploiting the steady progress in methods and software that the deposition of such data into the PDB on a massive scale has made possible. Accurate crystal structures of macromolecules are of high importance in the biological and biomedical fields. Models of crystal structures in the Protein Data Bank (PDB) are in general of very high quality as deposited. However, methods for obtaining the best model of a macromolecular structure from a given set of experimental X-ray data continue to progress at a rapid pace, making it possible to improve most PDB entries after their deposition by re-analyzing the original deposited data with more recent software. This possibility represents a very significant departure from the situation that prevailed when the PDB was created, when it was envisioned as a cumulative repository of static contents. A radical paradigm shift for the PDB is therefore proposed, away from the static archive model towards a much more dynamic body of continuously improving results in symbiosis with continuously improving methods and software. These simultaneous improvements in methods and final results are made possible by the current deposition of processed crystallographic data (structure-factor amplitudes) and will be supported further by the deposition of raw data (diffraction images). It is argued that it is both desirable and feasible to carry out small-scale and large-scale efforts to make this paradigm shift a reality. Small-scale efforts would focus on optimizing structures that are of interest to specific investigators. Large-scale efforts would undertake a systematic re-optimization of all of the structures in the PDB, or alternatively the redetermination of groups of structures that are either related to or focused on specific questions. All of the resulting structures should be

  7. Macromolecular architectures for organic photovoltaics.

    PubMed

    Popere, Bhooshan C; Della Pelle, Andrea M; Poe, Ambata; Thayumanavan, S

    2012-03-28

    Research in the field of organic photovoltaics has gained considerable momentum in the last two decades owing to the need for developing low-cost and efficient energy harvesting systems. Elegant molecular architectures have been designed, synthesized and employed as active materials for photovoltaic devices thereby leading to a better molecular structure-device property relationship understanding. In this perspective, we outline new macromolecular scaffolds that have been designed within the purview of each of the three fundamental processes involving light harvesting, charge separation and charge transport.

  8. Non-ideality of aqueous solutions of polyethylene glycol: consequences for its use as a macromolecular crystallizing agent in vapor-diffusion experiments.

    PubMed

    Arakali, S V; Luft, J R; DeTitta, G T

    1995-09-01

    Microisopiestic measurements of the concentrations of polyethylene glycol (PEG 8000) paired with the salts sodium chloride, ammonium sulfate and magnesium sulfate heptahydrate have been made in a sitting-drop arrangement with PEG in the droplet and salt in the reservoir. Resulting graphs of the concentrations of PEG and salt that are equivalent with respect to the vapor pressure of water are non-linear, do not intersect their origins, and demonstrate that relatively low (mM) salt concentrations are equivalent to relatively high PEG concentrations. The consequences of each of these observations for macromolecular crystallization by the vapor-diffusion technique when PEG is employed as the crystallizing agent are discussed.

  9. Solution Accounts for Structural Damping

    NASA Technical Reports Server (NTRS)

    Roussos, L. A.; Hyer, M. W.; Thornton, E. A.

    1982-01-01

    New analytical technique determines dynamic response of damped structures dominated by internal structural damping mechanisms. Though structural damping is often negligible compared with damping due to air friction and friction in joints, structural damping can be of major importance in structures having heavy damping treatments or in outer-space structures. Finite-element model includes nonlinear, nonviscous internal damping.

  10. Macromolecular engineering by atom transfer radical polymerization.

    PubMed

    Matyjaszewski, Krzysztof; Tsarevsky, Nicolay V

    2014-05-01

    This Perspective presents recent advances in macromolecular engineering enabled by ATRP. They include the fundamental mechanistic and synthetic features of ATRP with emphasis on various catalytic/initiation systems that use parts-per-million concentrations of Cu catalysts and can be run in environmentally friendly media, e.g., water. The roles of the major components of ATRP--monomers, initiators, catalysts, and various additives--are explained, and their reactivity and structure are correlated. The effects of media and external stimuli on polymerization rates and control are presented. Some examples of precisely controlled elements of macromolecular architecture, such as chain uniformity, composition, topology, and functionality, are discussed. Syntheses of polymers with complex architecture, various hybrids, and bioconjugates are illustrated. Examples of current and forthcoming applications of ATRP are covered. Future challenges and perspectives for macromolecular engineering by ATRP are discussed.

  11. Continuous mutual improvement of macromolecular structure models in the PDB and of X-ray crystallographic software: the dual role of deposited experimental data.

    PubMed

    Terwilliger, Thomas C; Bricogne, Gerard

    2014-10-01

    Accurate crystal structures of macromolecules are of high importance in the biological and biomedical fields. Models of crystal structures in the Protein Data Bank (PDB) are in general of very high quality as deposited. However, methods for obtaining the best model of a macromolecular structure from a given set of experimental X-ray data continue to progress at a rapid pace, making it possible to improve most PDB entries after their deposition by re-analyzing the original deposited data with more recent software. This possibility represents a very significant departure from the situation that prevailed when the PDB was created, when it was envisioned as a cumulative repository of static contents. A radical paradigm shift for the PDB is therefore proposed, away from the static archive model towards a much more dynamic body of continuously improving results in symbiosis with continuously improving methods and software. These simultaneous improvements in methods and final results are made possible by the current deposition of processed crystallographic data (structure-factor amplitudes) and will be supported further by the deposition of raw data (diffraction images). It is argued that it is both desirable and feasible to carry out small-scale and large-scale efforts to make this paradigm shift a reality. Small-scale efforts would focus on optimizing structures that are of interest to specific investigators. Large-scale efforts would undertake a systematic re-optimization of all of the structures in the PDB, or alternatively the redetermination of groups of structures that are either related to or focused on specific questions. All of the resulting structures should be made generally available, along with the precursor entries, with various views of the structures being made available depending on the types of questions that users are interested in answering.

  12. Continuous mutual improvement of macromolecular structure models in the PDB and of X-ray crystallographic software: the dual role of deposited experimental data

    PubMed Central

    Terwilliger, Thomas C.; Bricogne, Gerard

    2014-01-01

    Accurate crystal structures of macromolecules are of high importance in the biological and biomedical fields. Models of crystal structures in the Protein Data Bank (PDB) are in general of very high quality as deposited. However, methods for obtaining the best model of a macromolecular structure from a given set of experimental X-ray data continue to progress at a rapid pace, making it possible to improve most PDB entries after their deposition by re-analyzing the original deposited data with more recent software. This possibility represents a very significant departure from the situation that prevailed when the PDB was created, when it was envisioned as a cumulative repository of static contents. A radical paradigm shift for the PDB is therefore proposed, away from the static archive model towards a much more dynamic body of continuously improving results in symbiosis with continuously improving methods and software. These simultaneous improvements in methods and final results are made possible by the current deposition of processed crystallographic data (structure-factor amplitudes) and will be supported further by the deposition of raw data (diffraction images). It is argued that it is both desirable and feasible to carry out small-scale and large-scale efforts to make this paradigm shift a reality. Small-scale efforts would focus on optimizing structures that are of interest to specific investigators. Large-scale efforts would undertake a systematic re-optimization of all of the structures in the PDB, or alternatively the redetermination of groups of structures that are either related to or focused on specific questions. All of the resulting structures should be made generally available, along with the precursor entries, with various views of the structures being made available depending on the types of questions that users are interested in answering. PMID:25286839

  13. Continuous mutual improvement of macromolecular structure models in the PDB and of X-ray crystallographic software: The dual role of deposited experimental data

    DOE PAGES

    Terwilliger, Thomas C.; Bricogne, Gerard

    2014-09-30

    Accurate crystal structures of macromolecules are of high importance in the biological and biomedical fields. Models of crystal structures in the Protein Data Bank (PDB) are in general of very high quality as deposited. However, methods for obtaining the best model of a macromolecular structure from a given set of experimental X-ray data continue to progress at a rapid pace, making it possible to improve most PDB entries after their deposition by re-analyzing the original deposited data with more recent software. This possibility represents a very significant departure from the situation that prevailed when the PDB was created, when itmore » was envisioned as a cumulative repository of static contents. A radical paradigm shift for the PDB is therefore proposed, away from the static archive model towards a much more dynamic body of continuously improving results in symbiosis with continuously improving methods and software. These simultaneous improvements in methods and final results are made possible by the current deposition of processed crystallographic data (structure-factor amplitudes) and will be supported further by the deposition of raw data (diffraction images). It is argued that it is both desirable and feasible to carry out small-scale and large-scale efforts to make this paradigm shift a reality. Small-scale efforts would focus on optimizing structures that are of interest to specific investigators. Large-scale efforts would undertake a systematic re-optimization of all of the structures in the PDB, or alternatively the redetermination of groups of structures that are either related to or focused on specific questions. All of the resulting structures should be made generally available, along with the precursor entries, with various views of the structures being made available depending on the types of questions that users are interested in answering.« less

  14. Continuous mutual improvement of macromolecular structure models in the PDB and of X-ray crystallographic software: The dual role of deposited experimental data

    SciTech Connect

    Terwilliger, Thomas C.; Bricogne, Gerard

    2014-09-30

    Accurate crystal structures of macromolecules are of high importance in the biological and biomedical fields. Models of crystal structures in the Protein Data Bank (PDB) are in general of very high quality as deposited. However, methods for obtaining the best model of a macromolecular structure from a given set of experimental X-ray data continue to progress at a rapid pace, making it possible to improve most PDB entries after their deposition by re-analyzing the original deposited data with more recent software. This possibility represents a very significant departure from the situation that prevailed when the PDB was created, when it was envisioned as a cumulative repository of static contents. A radical paradigm shift for the PDB is therefore proposed, away from the static archive model towards a much more dynamic body of continuously improving results in symbiosis with continuously improving methods and software. These simultaneous improvements in methods and final results are made possible by the current deposition of processed crystallographic data (structure-factor amplitudes) and will be supported further by the deposition of raw data (diffraction images). It is argued that it is both desirable and feasible to carry out small-scale and large-scale efforts to make this paradigm shift a reality. Small-scale efforts would focus on optimizing structures that are of interest to specific investigators. Large-scale efforts would undertake a systematic re-optimization of all of the structures in the PDB, or alternatively the redetermination of groups of structures that are either related to or focused on specific questions. All of the resulting structures should be made generally available, along with the precursor entries, with various views of the structures being made available depending on the types of questions that users are interested in answering.

  15. Using NMR to Determine Protein Structure in Solution

    NASA Astrophysics Data System (ADS)

    Cavagnero, Silvia

    2003-02-01

    Nuclear magnetic resonance (NMR) is a marvelous spectroscopic technique that chemists, physicists, and biochemists routinely employ for their research around the world. This year half of the Nobel Prize for chemistry went to Kurt Wüthrich, who was recognized for the development of NMR-based techniques that lead to the structure determination of biomolecules in solution. In addition to implementing novel pulse sequences and software packages, Wüthrich also applied his methods to several biological systems of key importance to human health. These include the prion protein, which is heavily involved in the spongiform encephalopathy (best known as 'mad cow disease'), which recently caused numerous human deaths, particularly in the UK, due to ingestion of contaminated meat. Transverse relaxation optimized spectroscopy (TROSY) is the most intriguing new NMR method recently developed by Wüthrich and coworkers. This and other closely related pulse sequences promise to play a pivotal role in the extension of NMR to the conformational analysis of very large (up to the megadalton range) macromolecules and macromolecular complexes. More exciting new developments are expected in the near future.

  16. Automated macromolecular crystallization screening

    DOEpatents

    Segelke, Brent W.; Rupp, Bernhard; Krupka, Heike I.

    2005-03-01

    An automated macromolecular crystallization screening system wherein a multiplicity of reagent mixes are produced. A multiplicity of analysis plates is produced utilizing the reagent mixes combined with a sample. The analysis plates are incubated to promote growth of crystals. Images of the crystals are made. The images are analyzed with regard to suitability of the crystals for analysis by x-ray crystallography. A design of reagent mixes is produced based upon the expected suitability of the crystals for analysis by x-ray crystallography. A second multiplicity of mixes of the reagent components is produced utilizing the design and a second multiplicity of reagent mixes is used for a second round of automated macromolecular crystallization screening. In one embodiment the multiplicity of reagent mixes are produced by a random selection of reagent components.

  17. Blueprinting macromolecular electronics.

    PubMed

    Palma, Carlos-Andres; Samorì, Paolo

    2011-06-01

    Recently, by mastering either top-down or bottom-up approaches, tailor-made macromolecular nano-objects with semiconducting properties have been fabricated. These engineered nanostructures for organic electronics are based on conjugated systems predominantly made up of sp²-hybridized carbon, such as graphene nanoribbons. Here, we describe developments in a selection of these nanofabrication techniques, which include graphene carving, stimulus-induced synthesis of conjugated polymers and surface-assisted synthesis. We also assess their potential to reproduce chemically and spatially precise molecular arrangements, that is, molecular blueprints. In a broad context, the engineering of a molecular blueprint represents the fabrication of an integrated all-organic macromolecular electronic circuit. In this Perspective, we suggest chemical routes, as well as convergent on-surface synthesis and microfabrication approaches, for the ultimate goal of bringing the field closer to technology.

  18. Practical macromolecular cryocrystallography

    SciTech Connect

    Pflugrath, J. W.

    2015-05-27

    Current methods, reagents and experimental hardware for successfully and reproducibly flash-cooling macromolecular crystals to cryogenic temperatures for X-ray diffraction data collection are reviewed. Cryocrystallography is an indispensable technique that is routinely used for single-crystal X-ray diffraction data collection at temperatures near 100 K, where radiation damage is mitigated. Modern procedures and tools to cryoprotect and rapidly cool macromolecular crystals with a significant solvent fraction to below the glass-transition phase of water are reviewed. Reagents and methods to help prevent the stresses that damage crystals when flash-cooling are described. A method of using isopentane to assess whether cryogenic temperatures have been preserved when dismounting screened crystals is also presented.

  19. Clustering procedures for the optimal selection of data sets from multiple crystals in macromolecular crystallography

    SciTech Connect

    Foadi, James; Aller, Pierre; Alguel, Yilmaz; Cameron, Alex; Axford, Danny; Owen, Robin L.; Armour, Wes; Waterman, David G.; Iwata, So; Evans, Gwyndaf

    2013-08-01

    A systematic approach to the scaling and merging of data from multiple crystals in macromolecular crystallography is introduced and explained. The availability of intense microbeam macromolecular crystallography beamlines at third-generation synchrotron sources has enabled data collection and structure solution from microcrystals of <10 µm in size. The increased likelihood of severe radiation damage where microcrystals or particularly sensitive crystals are used forces crystallographers to acquire large numbers of data sets from many crystals of the same protein structure. The associated analysis and merging of multi-crystal data is currently a manual and time-consuming step. Here, a computer program, BLEND, that has been written to assist with and automate many of the steps in this process is described. It is demonstrated how BLEND has successfully been used in the solution of a novel membrane protein.

  20. Magnetic macromolecular cross linked enzyme aggregates (CLEAs) of glucoamylase.

    PubMed

    Nadar, Shamraja S; Rathod, Virendra K

    2016-02-01

    This work illustrates the preparation of magnetic macromolecular glucoamylase CLEAs using dialdehydic pectin, as a cross linker instead of traditional glutaraldehyde. The effect of precipitators type and amount, cross linker concentration, cross linking time and amount of amino functionalized magnetic nanoparticles (AFMNs) on glucoamylase activity was studied. Glucoamylase magnetic macromolecular CLEAs prepared by precipitation in presence of AFMNs by ammonium sulfate were subsequently cross linked by dialdehydic pectin. After cross-linked by pectin, 95.4% activity recovery was achieved in magnetic macromolecular CLEAs, whereas in case of glutaraldehyde cross linker, 85.3% activity recovery was achieved. Magnetic macromolecular CLEAs showed 2.91 and 1.27 folds higher thermal stability as compared to free and magnetic glutaraldehyde CLEAs. In kinetics study, magnetic macromolecular CLEAs retained same Km values, whereas magnetic glutaraldehyde CLEAs showed higher Km value than free enzyme. The porous structure of magnetic macromolecular CLEAs was not only enhanced mass transfer toward macromolecular substrates, but also showed compression resistance for 5 consecutive cycles which was checked in terms of effectiveness factor. At the end, in reusability study; magnetic macromolecular CLEAs were retained 84% activity after 10(th) cycle without leaching of enzyme which is 22% higher than traditional magnetic CLEAs.

  1. In-vacuum long-wavelength macromolecular crystallography.

    PubMed

    Wagner, Armin; Duman, Ramona; Henderson, Keith; Mykhaylyk, Vitaliy

    2016-03-01

    Structure solution based on the weak anomalous signal from native (protein and DNA) crystals is increasingly being attempted as part of synchrotron experiments. Maximizing the measurable anomalous signal by collecting diffraction data at longer wavelengths presents a series of technical challenges caused by the increased absorption of X-rays and larger diffraction angles. A new beamline at Diamond Light Source has been built specifically for collecting data at wavelengths beyond the capability of other synchrotron macromolecular crystallography beamlines. Here, the theoretical considerations in support of the long-wavelength beamline are outlined and the in-vacuum design of the endstation is discussed, as well as other hardware features aimed at enhancing the accuracy of the diffraction data. The first commissioning results, representing the first in-vacuum protein structure solution, demonstrate the promising potential of the beamline.

  2. In-vacuum long-wavelength macromolecular crystallography

    PubMed Central

    Wagner, Armin; Duman, Ramona; Henderson, Keith; Mykhaylyk, Vitaliy

    2016-01-01

    Structure solution based on the weak anomalous signal from native (protein and DNA) crystals is increasingly being attempted as part of synchrotron experiments. Maximizing the measurable anomalous signal by collecting diffraction data at longer wavelengths presents a series of technical challenges caused by the increased absorption of X-rays and larger diffraction angles. A new beamline at Diamond Light Source has been built specifically for collecting data at wavelengths beyond the capability of other synchrotron macromolecular crystallography beamlines. Here, the theoretical considerations in support of the long-wavelength beamline are outlined and the in-vacuum design of the endstation is discussed, as well as other hardware features aimed at enhancing the accuracy of the diffraction data. The first commissioning results, representing the first in-vacuum protein structure solution, demonstrate the promising potential of the beamline. PMID:26960130

  3. The Structure of Plant Cell Walls: I. The Macromolecular Components of the Walls of Suspension-cultured Sycamore Cells with a Detailed Analysis of the Pectic Polysaccharides.

    PubMed

    Talmadge, K W; Keegstra, K; Bauer, W D; Albersheim, P

    1973-01-01

    This is the first in a series of papers dealing with the structure of cell walls isolated from suspension-cultured sycamore cells (Acer pseudoplatanus). These studies have been made possible by the availability of purified hydrolytic enzymes and by recent improvements in the techniques of methylation analysis. These techniques have permitted us to identify and quantitate the macromolecular components of sycamore cell walls. These walls are composed of 10% arabinan, 2% 3,6-linked arabinogalactan, 23% cellulose, 9% oligo-arabinosides (attached to hydroxyproline), 8% 4-linked galactan, 10% hydroxyproline-rich protein, 16% rhamnogalacturonan, and 21% xyloglucan.The structures of the pectic polymers (the neutral arabinan, the neutral galactan, and the acidic rhamnogalacturonan) were obtained, in part, by methylation analysis of fragments of these polymers which were released from the sycamore walls by the action of a highly purified endopolygalacturonase. The data suggest a branched arabinan and a linear 4-linked galactan occurring as side chains on the rhamnogalacturonan. Small amounts or pieces of a xyloglucan, the wall hemicellulose, appear to be covalently linked to some of the galactan chains. Thus, the galactan appears to serve as a bridge between the xyloglucan and rhamnogalacturonan components of the wall.The rhamnogalacturonan consists of an alpha-(1 --> 4)-linked galacturonan chain which is interspersed with 2-linked rhamnosyl residues. The rhamnosyl residues are not randomly distributed in the chain but probably occur in units of rhamnosyl- (1 --> 4)-galacturonosyl- (1 --> 2)-rhamnosyl. This sequence appears to alternate with a homogalacturonan sequence containing approximately 8 residues of 4-linked galacturonic acid. About half of the rhamnosyl residues are branched, having a substituent attached to carbon 4. This is likely to be the site of attachment of the 4-linked galactan.The hydroxyprolyl oligo-arabinosides of the hydroxyproline-rich glycoprotein

  4. Practical macromolecular cryocrystallography

    PubMed Central

    Pflugrath, J. W.

    2015-01-01

    Cryocrystallography is an indispensable technique that is routinely used for single-crystal X-ray diffraction data collection at temperatures near 100 K, where radiation damage is mitigated. Modern procedures and tools to cryoprotect and rapidly cool macromolecular crystals with a significant solvent fraction to below the glass-transition phase of water are reviewed. Reagents and methods to help prevent the stresses that damage crystals when flash-cooling are described. A method of using isopentane to assess whether cryogenic temperatures have been preserved when dismounting screened crystals is also presented. PMID:26057787

  5. Microgravity and Macromolecular Crystallography

    NASA Technical Reports Server (NTRS)

    Kundrot, Craig E.; Judge, Russell A.; Pusey, Marc L.; Snell, Edward H.; Rose, M. Franklin (Technical Monitor)

    2000-01-01

    Macromolecular crystal growth has been seen as an ideal experiment to make use of the reduced acceleration environment provided by an orbiting spacecraft. The experiments are small, simply operated and have a high potential scientific and economic impact. In this review we examine the theoretical reasons why microgravity should be a beneficial environment for crystal growth and survey the history of experiments on the Space Shuttle Orbiter, on unmanned spacecraft, and on the Mir space station. Finally we outline the direction for optimizing the future use of orbiting platforms.

  6. Macromolecular crowding in the Escherichia coli periplasm maintains alpha-synuclein disorder.

    PubMed

    McNulty, Brian C; Young, Gregory B; Pielak, Gary J

    2006-02-01

    The natively disordered protein alpha-synuclein is the primary component of Lewy bodies, the cellular hallmark of Parkinson's disease. Most studies of this protein are performed in dilute solution, but its biologically relevant role is performed in the crowded environment inside cells. We addressed the effects of macromolecular crowding on alpha-synuclein by combining NMR data acquired in living Escherichia coli with in vitro NMR data. The crowded environment in the E.coli periplasm prevents a conformational change that is detected at 35 degrees C in dilute solution. This change is associated with an increase in hydrodynamic radius and the formation of secondary structure in the N-terminal 100 amino acid residues. By preventing this temperature-induced conformational change, crowding in the E.coli periplasm stabilizes the disordered monomer. We obtain the same stabilization in vitro upon crowding alpha-synuclein with 300 g/l of bovine serum albumin, indicating that crowding alone is sufficient to stabilize the disordered, monomeric protein. Two disease-associated variants (A30P and A53T) behave in the same way in both dilute solution and in the E.coli periplasm. These data reveal the importance of approaching the effects of macromolecular crowding on a case-by-case basis. Additionally, our work shows that discrete structured protein conformations may not be achieved by alpha-synuclein inside cells, implicating the commonly overlooked aspect of macromolecular crowding as a possible factor in the etiology of Parkinson's disease.

  7. Local Kinetic Measures of Macromolecular Structure Reveal Partitioning Among Multiple Parallel Pathways from the Earliest Steps in the Folding of a Large RNA Molecule

    SciTech Connect

    Laederach,A.; Shcherbakova, I.; Liang, M.; Brenowitz, M.; Altman, R.

    2006-01-01

    At the heart of the RNA folding problem is the number, structures, and relationships among the intermediates that populate the folding pathways of most large RNA molecules. Unique insight into the structural dynamics of these intermediates can be gleaned from the time-dependent changes in local probes of macromolecular conformation (e.g. reports on individual nucleotide solvent accessibility offered by hydroxyl radical ({center_dot}OH) footprinting). Local measures distributed around a macromolecule individually illuminate the ensemble of separate changes that constitute a folding reaction. Folding pathway reconstruction from a multitude of these individual measures is daunting due to the combinatorial explosion of possible kinetic models as the number of independent local measures increases. Fortunately, clustering of time progress curves sufficiently reduces the dimensionality of the data so as to make reconstruction computationally tractable. The most likely folding topology and intermediates can then be identified by exhaustively enumerating all possible kinetic models on a super-computer grid. The folding pathways and measures of the relative flux through them were determined for Mg{sup 2+} and Na{sup +}-mediated folding of the Tetrahymena thermophila group I intron using this combined experimental and computational approach. The flux during Mg{sup 2+}-mediated folding is divided among numerous parallel pathways. In contrast, the flux during the Na{sup +}-mediated reaction is predominantly restricted through three pathways, one of which is without detectable passage through intermediates. Under both conditions, the folding reaction is highly parallel with no single pathway accounting for more than 50% of the molecular flux. This suggests that RNA folding is non-sequential under a variety of different experimental conditions even at the earliest stages of folding. This study provides a template for the systematic analysis of the time-evolution of RNA structure

  8. Functional Sub-states by High-pressure Macromolecular Crystallography.

    PubMed

    Dhaussy, Anne-Claire; Girard, Eric

    2015-01-01

    At the molecular level, high-pressure perturbation is of particular interest for biological studies as it allows trapping conformational substates. Moreover, within the context of high-pressure adaptation of deep-sea organisms, it allows to decipher the molecular determinants of piezophily. To provide an accurate description of structural changes produced by pressure in a macromolecular system, developments have been made to adapt macromolecular crystallography to high-pressure studies. The present chapter is an overview of results obtained so far using high-pressure macromolecular techniques, from nucleic acids to virus capsid through monomeric as well as multimeric proteins.

  9. Macromolecular Crystallization in Microfluidics for the International Space Station

    NASA Technical Reports Server (NTRS)

    Monaco, Lisa A.; Spearing, Scott

    2003-01-01

    At NASA's Marshall Space Flight Center, the Iterative Biological Crystallization (IBC) project has begun development on scientific hardware for macromolecular crystallization on the International Space Station (ISS). Currently ISS crystallization research is limited to solution recipes that were prepared on the ground prior to launch. The proposed hardware will conduct solution mixing and dispensing on board the ISS, be fully automated, and have imaging functions via remote commanding from the ground. Utilizing microfluidic technology, IBC will allow for on orbit iterations. The microfluidics LabChip(R) devices that have been developed, along with Caliper Technologies, will greatly benefit researchers by allowing for precise fluid handling of nano/pico liter sized volumes. IBC will maximize the amount of science return by utilizing the microfluidic approach and be a valuable tool to structural biologists investigating medically relevant projects.

  10. ACHESYM: an algorithm and server for standardized placement of macromolecular models in the unit cell

    PubMed Central

    Kowiel, Marcin; Jaskolski, Mariusz; Dauter, Zbigniew

    2014-01-01

    Despite the existence of numerous useful conventions in structural crystallography, for example for the choice of the asymmetric part of the unit cell or of reciprocal space, surprisingly no standards are in use for the placement of the molecular model in the unit cell, often leading to inconsistencies or confusion. A conceptual solution for this problem has been proposed for macromolecular crystal structures based on the idea of the anti-Cheshire unit cell. Here, a program and server (called ACHESYM; http://achesym.ibch.poznan.pl) are presented for the practical implementation of this concept. In addition, the first task of ACHESYM is to find an optimal (compact) macromolecular assembly if more than one polymer chain exists. ACHESYM processes PDB (atomic parameters and TLS matrices) and mmCIF (diffraction data) input files to produce a new coordinate set and to reindex the reflections and modify their phases, if necessary. PMID:25478846

  11. Growth and Dissolution of Macromolecular Markov Chains

    NASA Astrophysics Data System (ADS)

    Gaspard, Pierre

    2016-07-01

    The kinetics and thermodynamics of free living copolymerization are studied for processes with rates depending on k monomeric units of the macromolecular chain behind the unit that is attached or detached. In this case, the sequence of monomeric units in the growing copolymer is a kth-order Markov chain. In the regime of steady growth, the statistical properties of the sequence are determined analytically in terms of the attachment and detachment rates. In this way, the mean growth velocity as well as the thermodynamic entropy production and the sequence disorder can be calculated systematically. These different properties are also investigated in the regime of depolymerization where the macromolecular chain is dissolved by the surrounding solution. In this regime, the entropy production is shown to satisfy Landauer's principle.

  12. Cell-Free Protein Expression under Macromolecular Crowding Conditions

    PubMed Central

    Ge, Xumeng; Luo, Dan; Xu, Jianfeng

    2011-01-01

    Background Cell-free protein expression (CFPE) comprised of in vitro transcription and translation is currently manipulated in relatively dilute solutions, in which the macromolecular crowding effects present in living cells are largely ignored. This may not only affect the efficiency of protein synthesis in vitro, but also limit our understanding of the functions and interactions of biomolecules involved in this fundamental biological process. Methodology/Principal Findings Using cell-free synthesis of Renilla luciferase in wheat germ extract as a model system, we investigated the CFPE under macromolecular crowding environments emulated with three different crowding agents: PEG-8000, Ficoll-70 and Ficoll-400, which vary in chemical properties and molecular size. We found that transcription was substantially enhanced in the macromolecular crowding solutions; up to 4-fold increase in the mRNA production was detected in the presence of 20% (w/v) of Ficoll-70. In contrast, translation was generally inhibited by the addition of each of the three crowding agents. This might be due to PEG-induced protein precipitation and non-specific binding of translation factors to Ficoll molecules. We further explored a two-stage CFPE in which transcription and translation was carried out under high then low macromolecular crowding conditions, respectively. It produced 2.2-fold higher protein yield than the coupled CFPE control. The macromolecular crowding effects on CFPE were subsequently confirmed by cell-free synthesis of an approximately two-fold larger protein, Firefly luciferase, under macromolecular crowding environments. Conclusions/Significance Three macromolecular crowding agents used in this research had opposite effects on transcription and translation. The results of this study should aid researchers in their choice of macromolecular crowding agents and shows that two-stage CFPE is more efficient than coupled CFPE. PMID:22174874

  13. Refined solution structure of human profilin I.

    PubMed Central

    Metzler, W. J.; Farmer, B. T.; Constantine, K. L.; Friedrichs, M. S.; Lavoie, T.; Mueller, L.

    1995-01-01

    Profilin is a ubiquitous eukaryotic protein that binds to both cytosolic actin and the phospholipid phosphatidylinositol-4,5-bisphosphate. These dual competitive binding capabilities of profilin suggest that profilin serves as a link between the phosphatidyl inositol cycle and actin polymerization, and thus profilin may be an essential component in the signaling pathway leading to cytoskeletal rearrangement. The refined three-dimensional solution structure of human profilin I has been determined using multidimensional heteronuclear NMR spectroscopy. Twenty structures were selected to represent the solution conformational ensemble. This ensemble of structures has root-mean-square distance deviations from the mean structure of 0.58 A for the backbone atoms and 0.98 A for all non-hydrogen atoms. Comparison of the solution structure of human profilin to the crystal structure of bovine profilin reveals that, although profilin adopts essentially identical conformations in both states, the solution structure is more compact than the crystal structure. Interestingly, the regions that show the most structural diversity are located at or near the actin-binding site of profilin. We suggest that structural differences are reflective of dynamical properties of profilin that facilitate favorable interactions with actin. The global folding pattern of human profilin also closely resembles that of Acanthamoeba profilin I, reflective of the 22% sequence identity and approximately 45% sequence similarity between these two proteins. PMID:7795529

  14. Computing the origin and evolution of the ribosome from its structure — Uncovering processes of macromolecular accretion benefiting synthetic biology

    PubMed Central

    Caetano-Anollés, Gustavo; Caetano-Anollés, Derek

    2015-01-01

    Accretion occurs pervasively in nature at widely different timeframes. The process also manifests in the evolution of macromolecules. Here we review recent computational and structural biology studies of evolutionary accretion that make use of the ideographic (historical, retrodictive) and nomothetic (universal, predictive) scientific frameworks. Computational studies uncover explicit timelines of accretion of structural parts in molecular repertoires and molecules. Phylogenetic trees of protein structural domains and proteomes and their molecular functions were built from a genomic census of millions of encoded proteins and associated terminal Gene Ontology terms. Trees reveal a ‘metabolic-first’ origin of proteins, the late development of translation, and a patchwork distribution of proteins in biological networks mediated by molecular recruitment. Similarly, the natural history of ancient RNA molecules inferred from trees of molecular substructures built from a census of molecular features shows patchwork-like accretion patterns. Ideographic analyses of ribosomal history uncover the early appearance of structures supporting mRNA decoding and tRNA translocation, the coevolution of ribosomal proteins and RNA, and a first evolutionary transition that brings ribosomal subunits together into a processive protein biosynthetic complex. Nomothetic structural biology studies of tertiary interactions and ancient insertions in rRNA complement these findings, once concentric layering assumptions are removed. Patterns of coaxial helical stacking reveal a frustrated dynamics of outward and inward ribosomal growth possibly mediated by structural grafting. The early rise of the ribosomal ‘turnstile’ suggests an evolutionary transition in natural biological computation. Results make explicit the need to understand processes of molecular growth and information transfer of macromolecules. PMID:27096056

  15. Significance of wall structure, macromolecular composition, and surface polymers to the survival and transport of Cryptosporidium parvum Oocysts

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The structure and composition of the oocyst wall are primary factors determining the survival of Cryptosporidium parvum oocysts outside the host. An external polymer matrix (glycocalyx) may mediate interactions with environmental surfaces and, thus, affect the transport of oocysts in water, soil, an...

  16. Complex structures – smart solutions

    PubMed Central

    2011-01-01

    The siliceous skeletal elements of the sponges, the spicules, represent one of the very few examples from where the molecule toolkit required for the formation of an extracellular mineral-based skeleton, has been elucidated. The distinguished feature of the inorganic matrix, the bio-silica, is its enzymatic synthesis mediated by silicatein. Ortho-silicate undergoes in the presence of silicatein a polycondensation reaction and forms bio-silica under release of reaction water. The protein silicatein aggregates non-covalently to larger filaments, a process that is stabilized by the silicatein-associated protein, silintaphin-1. These structured clusters form the axial filament that is located in the center of the spicules, the axial canal. Surprisingly it has now been found that the initial axial orientation, in which the spicules grow, is guided by cell processes through evagination. The approximately two µm wide cell extensions release silicatein that forms the first organic axial filament, which then synthesizes the inner core of the siliceous spicule rods. In parallel, the radial growth of the spicules is controlled by a telescopic arrangement of organic layers, into which bio-silica and ortho-silicate are deposited. Hence, the formation of a mature siliceous spicule is completed by a centrifugal accretion of bio-silica mediated by the silicatein in the axial filament, and a centripetal bio-silica deposition catalyzed by the extra-spicular silicatein. Finally this contribution highlights that for the ultimate determination of the spicule shapes, their species-specific morphologies, bio-silica hardens during a process which removes reaction water. The data presented can also provide new blueprints for the fabrication of novel biomaterials for biomedical applications.  PMID:22446527

  17. Macromolecular crystal growing system

    NASA Technical Reports Server (NTRS)

    Snyder, Robert S. (Inventor); Herren, Blair J. (Inventor); Carter, Daniel C. (Inventor); Yost, Vaughn H. (Inventor); Bugg, Charles E. (Inventor); Delucas, Lawrence J. (Inventor); Suddath, Fred L. (Inventor)

    1991-01-01

    A macromolecular crystal growing system especially designed for growing crystals in the low gravity of space as well as the gravity of earth includes at least one tray assembly, a carrier assembly which receives the tray, and a refrigeration-incubation module in which the carrier assembly is received. The tray assembly includes a plurality of sealed chambers with a plastic syringe and a plug means for the double tip of the syringe provided therein. Ganging mechanisms operate the syringes and plugs simultaneously in a precise and smooth operation. Preferably, the tray assemblies are mounted on ball bearing slides for smooth operation in inserting and removing the tray assemblies into the carrier assembly. The plugging mechanism also includes a loading control mechanism. A mechanism for leaving a syringe unplugged is also provided.

  18. Workshop on algorithms for macromolecular modeling. Final project report, June 1, 1994--May 31, 1995

    SciTech Connect

    Leimkuhler, B.; Hermans, J.; Skeel, R.D.

    1995-07-01

    A workshop was held on algorithms and parallel implementations for macromolecular dynamics, protein folding, and structural refinement. This document contains abstracts and brief reports from that workshop.

  19. RapiData: a practical course in macromolecular X-ray diffraction data measurement and structure solving at the NSLS.

    PubMed

    Sweet, R M; Soares, A

    2010-10-01

    RapiData provides two days of high-level lectures, then two more of experimental work on several beamlines of the National Synchrotron Light Source, for about 50 students. Students are invited to bring their own research projects for measurement, and about half of them do. The students frequently solve half a dozen structures during the course. Tutorials by the lecturers run throughout the data-collection period. The crystal-preparation laboratory is popular for tutorials and practice, and often there is a beamline available for practice. This article provides details about the organization of the course and tells some of the reasons for its success. PMID:21695040

  20. RapiData: a Practical Course in Macromolecular X-ray Diffraction Data Measurement and Structure Solving at the NSLS

    SciTech Connect

    Sweet, R.; Soares, A

    2010-01-01

    RapiData provides two days of high-level lectures, then two more of experimental work on several beamlines of the National Synchrotron Light Source, for about 50 students. Students are invited to bring their own research projects for measurement, and about half of them do. The students frequently solve half a dozen structures during the course. Tutorials by the lecturers run throughout the data-collection period. The crystal-preparation laboratory is popular for tutorials and practice, and often there is a beamline available for practice. This article provides details about the organization of the course and tells some of the reasons for its success.

  1. Mechanisms, kinetics, impurities and defects: consequences in macromolecular crystallization

    PubMed Central

    McPherson, Alexander; Kuznetsov, Yurii G.

    2014-01-01

    The nucleation and growth of protein, nucleic acid and virus crystals from solution are functions of underlying kinetic and thermodynamic parameters that govern the process, and these are all supersaturation-dependent. While the mechanisms of macromolecular crystal growth are essentially the same as for conventional crystals, the underlying parameters are vastly different, in some cases orders of magnitude lower, and this produces very different crystallization processes. Numerous physical features of macromolecular crystals are of serious interest to X-ray diffractionists; the resolution limit and mosaicity, for example, reflect the degree of molecular and lattice order. The defect structure of crystals has an impact on their response to flash-cooling, and terminal crystal size is dependent on impurity absorption and incorporation. The variety and extent of these issues are further unique to crystals of biological macromolecules. All of these features are amenable to study using atomic force microscopy, which provides direct images at the nanoscale level. Some of those images are presented here. PMID:24699728

  2. Role of Macromolecular Structure in the Ultrafast Energy and Electron Transfer Dynamics of a Light-Harvesting Polymer.

    PubMed

    Morseth, Zachary A; Pho, Toan V; Gilligan, Alexander T; Dillon, Robert J; Schanze, Kirk S; Reynolds, John R; Papanikolas, John M

    2016-08-18

    Ultrafast energy and electron transfer (EnT and ET, respectively) are characterized in a light-harvesting assembly based on a π-conjugated polymer (poly(fluorene)) functionalized with broadly absorbing pendant organic isoindigo (iI) chromophores using a combination of femtosecond transient absorption spectroscopy and large-scale computer simulation. Photoexcitation of the π-conjugated polymer leads to near-unity quenching of the excitation through a combination of EnT and ET to the iI pendants. The excited pendants formed by EnT rapidly relax within 30 ps, whereas recombination of the charge-separated state formed following ET occurs within 1200 ps. A computer model of the excited-state processes is developed by combining all-atom molecular dynamics simulations, which provides a molecular-level view of the assembly structure, with a kinetic model that accounts for the multiple excited-state quenching pathways. Direct comparison of the simulations with experimental data reveals that the underlying structure has a dramatic effect on the partitioning between EnT and ET in the polymer assembly, where the distance and orientation of the pendants in relation to the backbone serve to direct the dominant quenching pathway. PMID:27433946

  3. Fractionation and characterization of soy β-conglycinin-dextran conjugates via macromolecular crowding environment and dry heating.

    PubMed

    Weng, Jingyi; Qi, Junru; Yin, Shouwei; Wang, Jinmei; Guo, Jian; Feng, Jilu; Liu, Qianru; Zhu, Jianhua; Yang, Xiaoquan

    2016-04-01

    Conjugates of β-conglycinin and dextran were prepared by heating in solution under macromolecular crowding environment and dry-heating methods, and then fractionated by solubility at pH 4.8 and pH 6.5 and characterized. The results showed that the degree of glycation of the conjugates extracted from pH 4.8 were higher than the conjugates extracted from pH 6.5. Corresponding to the higher degree of glycation, it was supposed that the β-conglycinin of groups 4.8 of macromolecular crowding environment was completely surrounded by the dextran molecular while that of groups 6.5 were encircled partially with a lower degree of glycation. Compared to β-conglycinin, groups 4.8 demonstrated a decreasing surface hydrophobicity and sulfhydryl group content while groups 6.5 increased. The secondary structure of β-conglycinin soluble at pH 4.8 after conjugating under macromolecular crowding environment tended to stretch out and the highly ordered structure turn to random structures. The differences between the extraction of pH 4.8 and pH 6.5 conjugated by dry-heating methods were not as remarkable as the difference between the extraction conjugated by macromolecular crowding environment.

  4. Exosites determine macromolecular substrate recognition by prothrombinase.

    PubMed

    Krishnaswamy, S; Betz, A

    1997-10-01

    The prothrombinase complex, composed of factor Xa and factor Va assembled on a membrane surface, catalyzes the proteolytic formation of thrombin during blood coagulation. The molecular basis for the macromolecular substrate specificity of prothrombinase is poorly understood. By kinetic studies of prethrombin 2 cleavage by prothrombinase in the presence or absence of fragment 1.2, we show that occupation of the active site of the catalyst by inhibitors or alternate peptidyl substrates does not alter the affinity for prethrombin 2. Productive recognition of the macromolecular substrate therefore results from an initial interaction at enzymic sites (exosites) distinct from the active site, which largely determines substrate affinity. This interaction at exosites is evident even in the absence of activation peptide domains responsible for mediating the binding of the substrate to membranes or factor Va. Interactions at the active site with structures surrounding the scissile bond then precede bond cleavage and product release. The second binding step, which appears unfavorable, does not affect substrate affinity but contributes to the maximum catalytic rate. Therefore, binding specificity of prothrombinase for the macromolecular substrate is determined by exosites on the enzyme. We show that competitive inhibition of prethrombin 2 cleavage can be accomplished by interfering with the exosite binding step without obscuring the active site of the enzyme. These findings suggest limitations to the common approach of inferring the basis of factor Xa specificity with active site mutants or the targeting the active site of factor Xa with reversible inhibitors for therapeutic purposes. The achievement of distinctive macromolecular substrate specificities through exosite interactions and modulation of maximum catalytic rate through binding steps may also underlie the reactions catalyzed by the other coagulation complexes containing trypsin-like enzymes. PMID:9315846

  5. Energy transfer in macromolecular arrays

    NASA Astrophysics Data System (ADS)

    Andrews, David L.; Jenkins, Robert D.

    2003-11-01

    Macromolecular systems comprised of many light-sensitive centres (the photosynthetic unit, dendrimers, and other highly symmetric multichromophore arrays) are important structures offering challenges to theoreticians and synthetic chemists alike. Here we outline novel photophysical interactions predicted and observed in such arrays. Using the tools of molecular quantum electrodynamics (QED) we present quantum amplitudes for a variety of higher-order resonance energy transfer (RET) schemes associated with well-known nonlinear optical effects such as two- and three-photon absorption. The initial analysis is extended to account for situations where the participant donor species are identical and exist in a highly symmetric environment, leading to the possible formation of excitons. It emerges from the QED theory that such excitons are closely associated with the higher-order RET processes. General results are interpreted by analyzing particular molecular architectures which offer interesting features such as rate enhancement or limitation and exciton pathway quenching. Applications in the areas of photosynthesis, molecular logic gates and low-intensity fluorescence energy transfer are predicted.

  6. A database of macromolecular motions.

    PubMed Central

    Gerstein, M; Krebs, W

    1998-01-01

    We describe a database of macromolecular motions meant to be of general use to the structural community. The database, which is accessible on the World Wide Web with an entry point at http://bioinfo.mbb.yale.edu/MolMovDB , attempts to systematize all instances of protein and nucleic acid movement for which there is at least some structural information. At present it contains >120 motions, most of which are of proteins. Protein motions are further classified hierarchically into a limited number of categories, first on the basis of size (distinguishing between fragment, domain and subunit motions) and then on the basis of packing. Our packing classification divides motions into various categories (shear, hinge, other) depending on whether or not they involve sliding over a continuously maintained and tightly packed interface. In addition, the database provides some indication about the evidence behind each motion (i.e. the type of experimental information or whether the motion is inferred based on structural similarity) and attempts to describe many aspects of a motion in terms of a standardized nomenclature (e.g. the maximum rotation, the residue selection of a fixed core, etc.). Currently, we use a standard relational design to implement the database. However, the complexity and heterogeneity of the information kept in the database makes it an ideal application for an object-relational approach, and we are moving it in this direction. Specifically, in terms of storing complex information, the database contains plausible representations for motion pathways, derived from restrained 3D interpolation between known endpoint conformations. These pathways can be viewed in a variety of movie formats, and the database is associated with a server that can automatically generate these movies from submitted coordinates. PMID:9722650

  7. Solution structure of pyoverdin GM-II.

    PubMed

    Mohn, G; Koehl, P; Budzikiewicz, H; Lefèvre, J F

    1994-03-15

    The three-dimensional structure in solution of ferri-pyoverdin GM-II isolated from the culture medium of Pseudomonas fluorescens was determined by application of NMR methods to the Ga3+ analogue. Distance geometry calculations were performed with FILMAN using interproton distances and coupling constants as constraints. Further conformational analysis was carried out by energy minimization with MM2 and AMBER. Back-calculation of the NOESY spectra shows that the resulting structures are in agreement with the experimental data. PMID:8130197

  8. The use of a mini-κ goniometer head in macromolecular crystallography diffraction experiments

    SciTech Connect

    Brockhauser, Sandor; Ravelli, Raimond B. G.; McCarthy, Andrew A.

    2013-07-01

    Hardware and software solutions for MX data-collection strategies using the EMBL/ESRF miniaturized multi-axis goniometer head are presented. Most macromolecular crystallography (MX) diffraction experiments at synchrotrons use a single-axis goniometer. This markedly contrasts with small-molecule crystallography, in which the majority of the diffraction data are collected using multi-axis goniometers. A novel miniaturized κ-goniometer head, the MK3, has been developed to allow macromolecular crystals to be aligned. It is available on the majority of the structural biology beamlines at the ESRF, as well as elsewhere. In addition, the Strategy for the Alignment of Crystals (STAC) software package has been developed to facilitate the use of the MK3 and other similar devices. Use of the MK3 and STAC is streamlined by their incorporation into online analysis tools such as EDNA. The current use of STAC and MK3 on the MX beamlines at the ESRF is discussed. It is shown that the alignment of macromolecular crystals can result in improved diffraction data quality compared with data obtained from randomly aligned crystals.

  9. Structure and dynamics of calmodulin in solution.

    PubMed Central

    Wriggers, W; Mehler, E; Pitici, F; Weinstein, H; Schulten, K

    1998-01-01

    To characterize the dynamic behavior of calmodulin in solution, we have carried out molecular dynamics (MD) simulations of the Ca2+-loaded structure. The crystal structure of calmodulin was placed in a solvent sphere of radius 44 A, and 6 Cl- and 22 Na+ ions were included to neutralize the system and to model a 150 mM salt concentration. The total number of atoms was 32,867. During the 3-ns simulation, the structure exhibits large conformational changes on the nanosecond time scale. The central alpha-helix, which has been shown to unwind locally upon binding of calmodulin to target proteins, bends and unwinds near residue Arg74. We interpret this result as a preparative step in the more extensive structural transition observed in the "flexible linker" region 74-82 of the central helix upon complex formation. The major structural change is a reorientation of the two Ca2+-binding domains with respect to each other and a rearrangement of alpha-helices in the N-terminus domain that makes the hydrophobic target peptide binding site more accessible. This structural rearrangement brings the domains to a more favorable position for target binding, poised to achieve the orientation observed in the complex of calmodulin with myosin light-chain kinase. Analysis of solvent structure reveals an inhomogeneity in the mobility of water in the vicinity of the protein, which is attributable to the hydrophobic effect exerted by calmodulin's binding sites for target peptides. PMID:9545028

  10. Folding dynamics of Trp-cage in the presence of chemical interference and macromolecular crowding. I

    NASA Astrophysics Data System (ADS)

    Samiotakis, Antonios; Cheung, Margaret S.

    2011-11-01

    Proteins fold and function in the crowded environment of the cell's interior. In the recent years it has been well established that the so-called "macromolecular crowding" effect enhances the folding stability of proteins by destabilizing their unfolded states for selected proteins. On the other hand, chemical and thermal denaturation is often used in experiments as a tool to destabilize a protein by populating the unfolded states when probing its folding landscape and thermodynamic properties. However, little is known about the complicated effects of these synergistic perturbations acting on the kinetic properties of proteins, particularly when large structural fluctuations, such as protein folding, have been involved. In this study, we have first investigated the folding mechanism of Trp-cage dependent on urea concentration by coarse-grained molecular simulations where the impact of urea is implemented into an energy function of the side chain and/or backbone interactions derived from the all-atomistic molecular dynamics simulations with urea through a Boltzmann inversion method. In urea solution, the folding rates of a model miniprotein Trp-cage decrease and the folded state slightly swells due to a lack of contact formation between side chains at the terminal regions. In addition, the equilibrium m-values of Trp-cage from the computer simulations are in agreement with experimental measurements. We have further investigated the combined effects of urea denaturation and macromolecular crowding on Trp-cage's folding mechanism where crowding agents are modeled as hard-spheres. The enhancement of folding rates of Trp-cage is most pronounced by macromolecular crowding effect when the extended conformations of Trp-cast dominate at high urea concentration. Our study makes quantitatively testable predictions on protein folding dynamics in a complex environment involving both chemical denaturation and macromolecular crowding effects.

  11. Organic sulphur in macromolecular sedimentary organic matter: I. Structure and origin of sulphur-containing moieties in kerogen, asphaltenes and coal as revealed by flash pyrolysis

    NASA Astrophysics Data System (ADS)

    Sinninghe Damsté, Jaap S.; Eglinton, Timothy I.; De Leeuw, Jan W.; Schenck, P. A.

    1989-04-01

    The distributions of sulphur-containing compounds generated by flash pyrolysis of macromolecular sedimentary organic matter (kerogen, coal, asphaltenes) were studied by gas chromatography in combination with Sselective flame photometric detection or mass spectrometry. The abundance of S-containing pyrolysis products in the pyrolysates relative to other products was highly variable depending on the sample but the types of products were generally similar, being mainly composed of "gaseous" compounds ( e.g., hydrogen sulphide) and low molecular weight alkylthiophenes and alkylbenzothiophenes. The distribution patterns of the alkylated thiophenes were dominated by a limited number of all theoretically possible isomers. The alkyl substitution patterns of the dominant isomers bear a strong similarity to those of the organic S compounds present in the GC-amenable fractions of bitumens and immature oils. Therefore, it is suggested that these S-containing pyrolysis products are formed by pyrolysis of related thiophenic and benzothiophenic moieties present in the macromolecular sedimentary substances. Specific examples include those with linear alkyl, iso and anteiso alkyl, isoprenoid alkyl and steroidal carbon skeletons. The presence of higher molecular weight alkylthiophenes and alkylbenzothiophenes with these same carbon skeletons in pyrolysates of S-rich kerogens provided further evidence for the presence of these S-containing moieties. It is likely that these moieties have been formed by abiogenic S incorporation into sedimentary organic matter during early diagenesis.

  12. Transmucosal macromolecular drug delivery.

    PubMed

    Prego, C; García, M; Torres, D; Alonso, M J

    2005-01-01

    Mucosal surfaces are the most common and convenient routes for delivering drugs to the body. However, macromolecular drugs such as peptides and proteins are unable to overcome the mucosal barriers and/or are degraded before reaching the blood stream. Among the approaches explored so far in order to optimize the transport of these macromolecules across mucosal barriers, the use of nanoparticulate carriers represents a challenging but promising strategy. The present paper aims to compare the characteristics and potential of nanostructures based on the mucoadhesive polysaccharide chitosan (CS). These are CS nanoparticles, CS-coated oil nanodroplets (nanocapsules) and CS-coated lipid nanoparticles. The characteristics and behavior of CS nanoparticles and CS-coated lipid nanoparticles already reported [A. Vila, A. Sanchez, M. Tobio, P. Calvo, M.J. Alonso, Design of biodegradable particles for protein delivery, J. Control. Rel. 78 (2002) 15-24; R. Fernandez-Urrusuno, P. Calvo, C. Remunan-Lopez, J.L. Vila-Jato, M.J. Alonso, Enhancement of nasal absorption of insulin using chitosan nanoparticles, Pharm. Res. 16 (1999) 1576-1581; M. Garcia-Fuentes, D. Torres, M.J. Alonso, New surface-modified lipid nanoparticles as delivery vehicles for salmon calcitonin (submitted for publication).] are compared with those of CS nanocapsules originally reported here. The three types of systems have a size in the nanometer range and a positive zeta potential that was attributed to the presence of CS on their surface. They showed an important capacity for the association of peptides such as insulin, salmon calcitonin and proteins, such as tetanus toxoid. Their mechanism of interaction with epithelia was investigated using the Caco-2 model cell line. The results showed that CS-coated systems caused a concentration-dependent reduction in the transepithelial resistance of the cell monolayer. Moreover, within the range of concentrations investigated, these systems were internalized in the

  13. Molecular Control of Macromolecular Properties

    NASA Astrophysics Data System (ADS)

    Holcombe, Thomas Wesley, III

    Molecular level control over macromolecules has been at the heart of human advancement, long before Hermann Staudinger coined the term Makromolekule. From the development of primitive pharmaceuticals to the advanced materials that sent Man into outer-space, We have been tinkering with God's paint since our inception. The work described herein primarily involves advances concerning poly-aromatic macromolecules for use in future electronic applications, particularly that of organic photovoltaics. There is a final chapter, however, that gives the reader a taste of how some molecular level changes can be directly visualized with modern microscopy techniques. Chapter 1 provides a very brief introduction to conjugated polymers and molecular level control over macromolecular properties. Chapters 2--4 introduces the concept of polymer substitution as a means by which to control and improve charge generation in organic photovoltaic devices. Chapters 5 and 6 show how these polymers can take on larger, defined structures, yet are still beholden to intrinsic molecular properties---such as regioregularity, a fancy word for the regularity of the position in which two aromatic rings are joined together. Chapter 7 re-examines the role of polymer substitution on photovoltaic performance, this time with an emphasis on homo-polymer packing rather than electron transfer at the donor/acceptor interface. Finally, Chapter 8 visualizes how controlling the environment about a single metal atom can lead directly to a cyclic polyolefin. Individually, these advances do not yield any breakthroughs noticeable to a general audience; collectively, they sit atop a mountain of human knowledge, waiting to provide a stepping stone for the next generation.

  14. Atomic-force-microscopy studies of phase separations in macromolecular systems

    NASA Astrophysics Data System (ADS)

    Kuznetsov, Yu. G.; Malkin, A. J.; McPherson, A.

    1998-09-01

    Atomic force microscopy (AFM) has been used to visualize events arising from the formation of intervening metastable phases at the surfaces of macromolecular crystals growing from solution. Crystals investigated were of the proteins canavalin, thaumatin, lipase, xylanase, and catalase, crystals of transfer RNA, and crystals of satellite tobacco mosaic virus. Two types of aggregates were observed. The first were small, linear and branched aggregates, perhaps fractile in structure. These were incorporated into growing crystals as impurities, and they produced defects of various kinds. The second aggregate form we infer to be liquid-protein droplets which were particularly evident in freshly mixed protein-precipitant solutions. Droplets, upon sedimentation, have two possible fates. In some cases they immediately restructured as crystalline multilayer stacks whose development was guided by and contiguous with the underlying lattice. These contributed to the ordered growth of the crystal by serving as sources of growth steps. In other cases, liquid-protein droplets formed distinct microcrystals, somehow discontinuous with the underlying lattice, and these were subsequently incorporated into the growing substrate crystal with the formation of defects. Scarring experiments with the AFM tip indicated that liquid-protein droplets with the potential to rapidly crystallize were a consequence of concentration instabilities near the crystal's surfaces. The AFM study suggests that phase separation and the appearance of aggregates having limited order is a common occurrence in supersaturated macromolecular solutions such as the protein-precipitant solutions used for crystallization.

  15. Structuring of polymer solutions upon solvent evaporation

    NASA Astrophysics Data System (ADS)

    Schaefer, C.; van der Schoot, P.; Michels, J. J.

    2015-02-01

    The morphology of solution-cast, phase-separated polymers becomes finer with increasing solvent evaporation rate. We address this observation theoretically for a model polymer where demixing is induced by steady solvent evaporation. In contrast to what is the case for a classical, thermal quench involving immiscible blends, the spinodal instability initially develops slowly and the associated length scale is not time invariant but decreases with time as t-1 /2. After a time lag, phase separation accelerates. Time lag and characteristic length exhibit power-law behavior as a function of the evaporation rate with exponents of -2 /3 and -1 /6 . Interestingly, at later stages the spinodal structure disappears completely while a second length scale develops. The associated structure coarsens but does not follow the usual Lifshitz-Slyozov-Wagner kinetics.

  16. Structure and aggregation in model tetramethylurea solutions

    SciTech Connect

    Gupta, Rini; Patey, G. N.

    2014-08-14

    The structure of model aqueous tetramethylurea (TMU) solutions is investigated employing large-scale (32 000, 64 000 particles) molecular dynamics simulations. Results are reported for TMU mole fractions, X{sub t}, ranging from infinite dilution up to 0.07, and for two temperatures, 300 and 330 K. Two existing force fields for TMU-water solutions are considered. These are the GROMOS 53A6 united-atom TMU model combined with SPC/E water [TMU(GROMOS-UA)/W(SPC/E)], and the more frequently employed AMBER03 all-atom force field for TMU combined with the TIP3P water model [TMU(AMBER-AA)/W(TIP3P)]. It is shown that TMU has a tendency towards aggregation for both models considered, but the tendency is significantly stronger for the [TMU(AMBER-AA)/W(TIP3P)] force field. For this model signs of aggregation are detected at X{sub t} = 0.005, aggregation is a well established feature of the solution at X{sub t} = 0.02, and the aggregates increase further in size with increasing concentration. This is in agreement with at least some experimental studies, which report signals of aggregation in the low concentration regime. The TMU aggregates exhibit little structure and are simply loosely ordered, TMU-rich regions of solution. The [TMU(GROMOS-UA)/W(SPC/E)] model shows strong signs of aggregation only at higher concentrations (X{sub t} ≳ 0.04), and the aggregates appear more loosely ordered, and less well-defined than those occurring in the [TMU(AMBER-AA)/W(TIP3P)] system. For both models, TMU aggregation increases when the temperature is increased from 300 to 330 K, consistent with an underlying entropy driven, hydrophobic interaction mechanism. At X{sub t} = 0.07, the extra-molecular correlation length expected for microheterogeneous solutions has become comparable with the size of the simulation cell for both models considered, indicating that even the systems simulated here are sufficiently large only at low concentrations.

  17. Probing the Macromolecular Organization of Cells by Electron Tomography

    PubMed Central

    Hoenger, Andreas; McIntosh, J. Richard

    2010-01-01

    Summary A major goal in cell biology is to understand the functional organization of macromolecular complexes in vivo. Electron microscopy is helping cell biologists to achieve this goal, thanks to its ability to resolve structural details in the nanometer range. While issues related to specimen preparation, imaging, and image interpretation make this approach to cell architecture difficult, recent improvements in methods, equipment, and software have facilitated the study of both important macromolecular complexes and comparatively large volumes from cellular specimens. Here, we describe recent progress in electron microscopy of cells and the ways in which the relevant methodologies are helping to elucidate cell architecture. PMID:19185480

  18. Solution structure of human sorting nexin 22.

    PubMed

    Song, Jikui; Zhao, Kate Qin; Newman, Carrie L Loushin; Vinarov, Dmitriy A; Markley, John L

    2007-05-01

    The sorting nexins (SNXs) constitute a large group of PX domain-containing proteins that play critical roles in protein trafficking. We report here the solution structure of human sorting nexin 22 (SNX22). Although SNX22 has <30% sequence identity with any PX domain protein of known structure, it was found to contain the alpha/beta fold and compact structural core characteristic of PX domains. Analysis of the backbone dynamics of SNX22 by NMR relaxation measurements revealed that the two walls of the ligand binding cleft undergo internal motions: on the picosecond timescale for the beta1/beta2 loop and on the micro- to millisecond timescale for the loop between the polyproline motif and helix alpha2. Regions of the SNX22 structure that differ from those of other PX domains include the loop connecting strands beta1 and beta2 and the loop connecting helices alpha1 and alpha2, which appear to be more mobile than corresponding loops in other known structures. The interaction of dibutanoyl-phosphatidylinositol-3-phosphate (dibutanoyl-PtdIns(3)P) with SNX22 was investigated by an NMR titration experiment, which identified the binding site in a basic cleft and indicated that ligand binding leads only to a local structural rearrangement as has been found with other PX domains. Because motions in the loops are damped out when dibutanoyl-PtdIns(3)P binds, entropic effects could contribute to the lower affinity of SNX22 for this ligand compared to other PX domains. PMID:17400918

  19. Photoinduced Electron Transfer Reactions for Macromolecular Syntheses.

    PubMed

    Dadashi-Silab, Sajjad; Doran, Sean; Yagci, Yusuf

    2016-09-14

    Photochemical reactions, particularly those involving photoinduced electron transfer processes, establish a substantial contribution to the modern synthetic chemistry, and the polymer community has been increasingly interested in exploiting and developing novel photochemical strategies. These reactions are efficiently utilized in almost every aspect of macromolecular architecture synthesis, involving initiation, control of the reaction kinetics and molecular structures, functionalization, and decoration, etc. Merging with polymerization techniques, photochemistry has opened up new intriguing and powerful avenues for macromolecular synthesis. Construction of various polymers with incredibly complex structures and specific control over the chain topology, as well as providing the opportunity to manipulate the reaction course through spatiotemporal control, are one of the unique abilities of such photochemical reactions. This review paper provides a comprehensive account of the fundamentals and applications of photoinduced electron transfer reactions in polymer synthesis. Besides traditional photopolymerization methods, namely free radical and cationic polymerizations, step-growth polymerizations involving electron transfer processes are included. In addition, controlled radical polymerization and "Click Chemistry" methods have significantly evolved over the last few decades allowing access to narrow molecular weight distributions, efficient regulation of the molecular weight and the monomer sequence and incredibly complex architectures, and polymer modifications and surface patterning are covered. Potential applications including synthesis of block and graft copolymers, polymer-metal nanocomposites, various hybrid materials and bioconjugates, and sequence defined polymers through photoinduced electron transfer reactions are also investigated in detail.

  20. Fluid Physics and Macromolecular Crystal Growth in Microgravity

    NASA Technical Reports Server (NTRS)

    Helliwell, John R.; Snell, Edward H.; Chayen, Naomi E.; Judge, Russell A.; Boggon, Titus J.; Pusey, M. L.; Rose, M. Franklin (Technical Monitor)

    2000-01-01

    The first protein crystallization experiment in microgravity was launched in April, 1981 and used Germany's Technologische Experimente unter Schwerelosigkeit (TEXUS 3) sounding rocket. The protein P-galactosidase (molecular weight 465Kda) was chosen as the sample with a liquid-liquid diffusion growth method. A sliding device brought the protein, buffer and salt solution into contact when microgravity was reached. The sounding rocket gave six minutes of microgravity time with a cine camera and schlieren optics used to monitor the experiment, a single growth cell. In microgravity a strictly laminar diffusion process was observed in contrast to the turbulent convection seen on the ground. Several single crystals, approx 100micron in length, were formed in the flight which were of inferior but of comparable visual quality to those grown on the ground over several days. A second experiment using the same protocol but with solutions cooled to -8C (kept liquid with glycerol antifreeze) again showed laminar diffusion. The science of macromolecular structural crystallography involves crystallization of the macromolecule followed by use of the crystal for X-ray diffraction experiments to determine the three dimensional structure of the macromolecule. Neutron protein crystallography is employed for elucidation of H/D exchange and for improved definition of the bound solvent (D20). The structural information enables an understanding of how the molecule functions with important potential for rational drug design, improved efficiency of industrial enzymes and agricultural chemical development. The removal of turbulent convection and sedimentation in microgravity, and the assumption that higher quality crystals will be produced, has given rise to the growing number of crystallization experiments now flown. Many experiments can be flown in a small volume with simple, largely automated, equipment - an ideal combination for a microgravity experiment. The term "protein crystal growth

  1. How can macromolecular crowding inhibit biological reactions? The enhanced formation of DNA nanoparticles

    PubMed Central

    Hou, Sen; Trochimczyk, Piotr; Sun, Lili; Wisniewska, Agnieszka; Kalwarczyk, Tomasz; Zhang, Xuzhu; Wielgus-Kutrowska, Beata; Bzowska, Agnieszka; Holyst, Robert

    2016-01-01

    In contrast to the already known effect that macromolecular crowding usually promotes biological reactions, solutions of PEG 6k at high concentrations stop the cleavage of DNA by HindIII enzyme, due to the formation of DNA nanoparticles. We characterized the DNA nanoparticles and probed the prerequisites for their formation using multiple techniques such as fluorescence correlation spectroscopy, dynamic light scattering, fluorescence analytical ultracentrifugation etc. In >25% PEG 6k solution, macromolecular crowding promotes the formation of DNA nanoparticles with dimensions of several hundreds of nanometers. The formation of DNA nanoparticles is a fast and reversible process. Both plasmid DNA (2686 bp) and double-stranded/single-stranded DNA fragment (66bp/nt) can form nanoparticles. We attribute the enhanced nanoparticle formation to the depletion effect of macromolecular crowding. This study presents our idea to enhance the formation of DNA nanoparticles by macromolecular crowding, providing the first step towards a final solution to efficient gene therapy. PMID:26903405

  2. Organoactinide chemistry: synthesis, structure, and solution dynamics

    SciTech Connect

    Brennan, J.G.

    1985-12-01

    This thesis considers three aspects of organoactinide chemistry. In chapter one, a bidentate phosphine ligand was used to kinetically stabilize complexes of the type Cp/sub 2/MX/sub 2/. Ligand redistribution processes are present throughout the synthetic work, as has often been observed in uranium cyclopentadienyl chemistry. The effects of covalent M-L bonding on the solution and solid state properties of U(III) coordination complexes are considered. In particular, the nature of the more subtle interaction between the metal and the neutral ligand are examined. Using relative basicity data obtained in solution, and solid state structural data (and supplemented by gas phase photoelectron measurements), it is demonstrated that the more electron rich U(III) centers engage in significant U ..-->.. L ..pi..-donation. Trivalent uranium is shown to be capable of acting either as a one- or two-electron reducing agent toward a wide variety of unsaturated organic and inorganic molecules, generating molecular classes unobtainable via traditional synthetic approaches, as well as offering an alternative synthetic approach to molecules accessible via metathesis reactions. Ligand redistribution processes are again observed, but given the information concerning ligand lability, this reactivity pattern is applied to the synthesis of pure materials inaccessible from redox chemistry. 214 refs., 33 figs., 10 tabs.

  3. Structure and Dynamics of Cellulose Molecular Solutions

    NASA Astrophysics Data System (ADS)

    Wang, Howard; Zhang, Xin; Tyagi, Madhusudan; Mao, Yimin; Briber, Robert

    Molecular dissolution of microcrystalline cellulose has been achieved through mixing with ionic liquid 1-Ethyl-3-methylimidazolium acetate (EMIMAc), and organic solvent dimethylformamide (DMF). The mechanism of cellulose dissolution in tertiary mixtures has been investigated by combining quasielastic and small angle neutron scattering (QENS and SANS). As SANS data show that cellulose chains take Gaussian-like conformations in homogenous solutions, which exhibit characteristics of having an upper critical solution temperature, the dynamic signals predominantly from EMIMAc molecules indicate strong association with cellulose in the dissolution state. The mean square displacement quantities support the observation of the stoichiometric 3:1 EMIMAc to cellulose unit molar ratio, which is a necessary criterion for the molecular dissolution of cellulose. Analyses of dynamics structure factors reveal the temperature dependence of a slow and a fast process for EMIMAc's bound to cellulose and in DMF, respectively, as well as a very fast process due possibly to the rotational motion of methyl groups, which persisted to near the absolute zero.

  4. Concentration Dependent Structure of Block Copolymer Solutions

    NASA Astrophysics Data System (ADS)

    Choi, Soohyung; Bates, Frank S.; Lodge, Timothy P.

    2015-03-01

    Addition of solvent molecules into block copolymer can induce additional interactions between the solvent and both blocks, and therefore expands the range of accessible self-assembled morphologies. In particular, the distribution of solvent molecules plays a key role in determining the microstructure and its characteristic domain spacing. In this study, concentration dependent structures formed by poly(styrene-b-ethylene-alt-propylene) (PS-PEP) solution in squalane are investigated using small-angle X-ray scattering. This reveals that squalane is essentially completely segregated into the PEP domains. In addition, the conformation of the PS block changes from stretched to nearly fully relaxed (i.e., Gaussian conformation) as amounts of squalane increases. NRF

  5. Structure of aqueous sodium perchlorate solutions.

    PubMed

    General, Ignacio J; Asciutto, Eliana K; Madura, Jeffry D

    2008-12-01

    Salt solutions have been the object of study of many scientists through history, but one of the most important findings came along when the Hofmeister series were discovered. Their importance arises from the fact that they influence the relative solubility of proteins, and solubility is directly related to one of today's holy grails: protein folding. In this work we characterize one of the more-destabilizing salts in the series, sodium perchlorate, by studying it as an aqueous solution at various concentrations ranging from 0.08 to 1.60 mol/L. Molecular dynamics simulations at room temperature permitted a detailed study of the organization of solvent and cosolvent, in terms of its radial distribution functions, along with the study of the structure of hydrogen bonds in the ions' solvation shells. We found that the distribution functions have some variations in their shape as concentration changes, but the position of their peaks is mostly unaffected. Regarding water, the most salient fact is the noticeable (although small) change in the second hydration shell and even beyond, especially for g(O(w)***O(w)), showing that the locality of salt effects should not be restricted to considerations of only the first solvation shell. The perturbation of the second shell also appears in the study of the HB network, where the difference between the number of HBs around a water molecule and around the Na(+) cation gets much smaller as one goes from the first to the second solvation shell, yet the difference is not negligible. Nevertheless, the effect of the ions past their first hydration shell is not enough to make a noticeable change in the global HB network. The Kirkwood-Buff theory of liquids was applied to our system, in order to calculate the activity derivative of the cosolvent. This coefficient, along with a previously calculated preferential binding, allowed us to establish that if a folded AP peptide is immersed in the studied solution, becoming the solute, then

  6. Macromolecular Materials and Engineering

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cover: The image shows electrospun fibers based on poly(lactic acid)/polyaniline blends with diameters from 90 to 1000 nm. The structural characteristics of the fibers are compared to cast films by scanning electron microscopy, small-angle X-ray scattering, differential scanning calorimetry, and ato...

  7. Multiscale macromolecular simulation: role of evolving ensembles.

    PubMed

    Singharoy, A; Joshi, H; Ortoleva, P J

    2012-10-22

    Multiscale analysis provides an algorithm for the efficient simulation of macromolecular assemblies. This algorithm involves the coevolution of a quasiequilibrium probability density of atomic configurations and the Langevin dynamics of spatial coarse-grained variables denoted order parameters (OPs) characterizing nanoscale system features. In practice, implementation of the probability density involves the generation of constant OP ensembles of atomic configurations. Such ensembles are used to construct thermal forces and diffusion factors that mediate the stochastic OP dynamics. Generation of all-atom ensembles at every Langevin time step is computationally expensive. Here, multiscale computation for macromolecular systems is made more efficient by a method that self-consistently folds in ensembles of all-atom configurations constructed in an earlier step, history, of the Langevin evolution. This procedure accounts for the temporal evolution of these ensembles, accurately providing thermal forces and diffusions. It is shown that efficiency and accuracy of the OP-based simulations is increased via the integration of this historical information. Accuracy improves with the square root of the number of historical timesteps included in the calculation. As a result, CPU usage can be decreased by a factor of 3-8 without loss of accuracy. The algorithm is implemented into our existing force-field based multiscale simulation platform and demonstrated via the structural dynamics of viral capsomers.

  8. Macromolecular Crystal Quality

    NASA Technical Reports Server (NTRS)

    Snell, Edward H.; Borgstahl, Gloria E. O.; Bellamy, Henry D.; Curreri, Peter A. (Technical Monitor)

    2001-01-01

    There are many ways of judging a good crystal. Which we use depends on the qualities we seek. For gemstones size, clarity and impurity levels (color) are paramount. For the semiconductor industry purity is probably the most important quality. For the structural crystallographer the primary desideratum is the somewhat more subtle concept of internal order. In this chapter we discuss the effect of internal order (or the lack of it) on the crystal's diffraction properties.

  9. Clustering procedures for the optimal selection of data sets from multiple crystals in macromolecular crystallography

    PubMed Central

    Foadi, James; Aller, Pierre; Alguel, Yilmaz; Cameron, Alex; Axford, Danny; Owen, Robin L.; Armour, Wes; Waterman, David G.; Iwata, So; Evans, Gwyndaf

    2013-01-01

    The availability of intense microbeam macromolecular crystallography beamlines at third-generation synchrotron sources has enabled data collection and structure solution from microcrystals of <10 µm in size. The increased likelihood of severe radiation damage where microcrystals or particularly sensitive crystals are used forces crystallographers to acquire large numbers of data sets from many crystals of the same protein structure. The associated analysis and merging of multi-crystal data is currently a manual and time-consuming step. Here, a computer program, BLEND, that has been written to assist with and automate many of the steps in this process is described. It is demonstrated how BLEND has successfully been used in the solution of a novel membrane protein. PMID:23897484

  10. Facile Preparation of a Macromolecular Benzophenone Photoinitiator

    NASA Astrophysics Data System (ADS)

    Huang, Qinghua; Gu, Lingling; Bai, Xiongxiong; Cheng, Chuanjie

    2014-08-01

    Photoinitiators play important roles in the preparation of photo-cured resins. Macromolecular as well as reactive photoinitiators have attracted much attention both in industry and in academia due to the disadvantages of conventional small molecular photoinitiators such as volatility and mobility. A macromolecular benzophenone photoinitiator was designed and efficiently synthesized in this study. Hydroxyl-containing Michler's ketone was firstly synthesized in 82% yield, followed by reacting with toluene di-isocyanate (TDI) to prepare polyurethanetype macromolecular benzophenone photoinitiator.

  11. An autonomous structural health monitoring solution

    NASA Astrophysics Data System (ADS)

    Featherston, Carol A.; Holford, Karen M.; Pullin, Rhys; Lees, Jonathan; Eaton, Mark; Pearson, Matthew

    2013-05-01

    Combining advanced sensor technologies, with optimised data acquisition and diagnostic and prognostic capability, structural health monitoring (SHM) systems provide real-time assessment of the integrity of bridges, buildings, aircraft, wind turbines, oil pipelines and ships, leading to improved safety and reliability and reduced inspection and maintenance costs. The implementation of power harvesting, using energy scavenged from ambient sources such as thermal gradients and sources of vibration in conjunction with wireless transmission enables truly autonomous systems, reducing the need for batteries and associated maintenance in often inaccessible locations, alongside bulky and expensive wiring looms. The design and implementation of such a system however presents numerous challenges. A suitable energy source or multiple sources capable of meeting the power requirements of the system, over the entire monitoring period, in a location close to the sensor must be identified. Efficient power management techniques must be used to condition the power and deliver it, as required, to enable appropriate measurements to be taken. Energy storage may be necessary, to match a continuously changing supply and demand for a range of different monitoring states including sleep, record and transmit. An appropriate monitoring technique, capable of detecting, locating and characterising damage and delivering reliable information, whilst minimising power consumption, must be selected. Finally a wireless protocol capable of transmitting the levels of information generated at the rate needed in the required operating environment must be chosen. This paper considers solutions to some of these challenges, and in particular examines SHM in the context of the aircraft environment.

  12. Macromolecular diffractive imaging using imperfect crystals

    PubMed Central

    Ayyer, Kartik; Yefanov, Oleksandr; Oberthür, Dominik; Roy-Chowdhury, Shatabdi; Galli, Lorenzo; Mariani, Valerio; Basu, Shibom; Coe, Jesse; Conrad, Chelsie E.; Fromme, Raimund; Schaffer, Alexander; Dörner, Katerina; James, Daniel; Kupitz, Christopher; Metz, Markus; Nelson, Garrett; Lourdu Xavier, Paulraj; Beyerlein, Kenneth R.; Schmidt, Marius; Sarrou, Iosifina; Spence, John C. H.; Weierstall, Uwe; White, Thomas A.; Yang, Jay-How; Zhao, Yun; Liang, Mengning; Aquila, Andrew; Hunter, Mark S.; Robinson, Joseph S.; Koglin, Jason E.; Boutet, Sébastien; Fromme, Petra; Barty, Anton; Chapman, Henry N.

    2016-01-01

    The three-dimensional structures of macromolecules and their complexes are predominantly elucidated by X-ray protein crystallography. A major limitation is access to high-quality crystals, to ensure X-ray diffraction extends to sufficiently large scattering angles and hence yields sufficiently high-resolution information that the crystal structure can be solved. The observation that crystals with shrunken unit-cell volumes and tighter macromolecular packing often produce higher-resolution Bragg peaks1,2 hints that crystallographic resolution for some macromolecules may be limited not by their heterogeneity but rather by a deviation of strict positional ordering of the crystalline lattice. Such displacements of molecules from the ideal lattice give rise to a continuous diffraction pattern, equal to the incoherent sum of diffraction from rigid single molecular complexes aligned along several discrete crystallographic orientations and hence with an increased information content3. Although such continuous diffraction patterns have long been observed—and are of interest as a source of information about the dynamics of proteins4 —they have not been used for structure determination. Here we show for crystals of the integral membrane protein complex photosystem II that lattice disorder increases the information content and the resolution of the diffraction pattern well beyond the 4.5 Å limit of measurable Bragg peaks, which allows us to directly phase5 the pattern. With the molecular envelope conventionally determined at 4.5 Å as a constraint, we then obtain a static image of the photosystem II dimer at 3.5 Å resolution. This result shows that continuous diffraction can be used to overcome long-supposed resolution limits of macromolecular crystallography, with a method that puts great value in commonly encountered imperfect crystals and opens up the possibility for model-free phasing6,7. PMID:26863980

  13. Macromolecular diffractive imaging using imperfect crystals.

    PubMed

    Ayyer, Kartik; Yefanov, Oleksandr M; Oberthür, Dominik; Roy-Chowdhury, Shatabdi; Galli, Lorenzo; Mariani, Valerio; Basu, Shibom; Coe, Jesse; Conrad, Chelsie E; Fromme, Raimund; Schaffer, Alexander; Dörner, Katerina; James, Daniel; Kupitz, Christopher; Metz, Markus; Nelson, Garrett; Xavier, Paulraj Lourdu; Beyerlein, Kenneth R; Schmidt, Marius; Sarrou, Iosifina; Spence, John C H; Weierstall, Uwe; White, Thomas A; Yang, Jay-How; Zhao, Yun; Liang, Mengning; Aquila, Andrew; Hunter, Mark S; Robinson, Joseph S; Koglin, Jason E; Boutet, Sébastien; Fromme, Petra; Barty, Anton; Chapman, Henry N

    2016-02-11

    The three-dimensional structures of macromolecules and their complexes are mainly elucidated by X-ray protein crystallography. A major limitation of this method is access to high-quality crystals, which is necessary to ensure X-ray diffraction extends to sufficiently large scattering angles and hence yields information of sufficiently high resolution with which to solve the crystal structure. The observation that crystals with reduced unit-cell volumes and tighter macromolecular packing often produce higher-resolution Bragg peaks suggests that crystallographic resolution for some macromolecules may be limited not by their heterogeneity, but by a deviation of strict positional ordering of the crystalline lattice. Such displacements of molecules from the ideal lattice give rise to a continuous diffraction pattern that is equal to the incoherent sum of diffraction from rigid individual molecular complexes aligned along several discrete crystallographic orientations and that, consequently, contains more information than Bragg peaks alone. Although such continuous diffraction patterns have long been observed--and are of interest as a source of information about the dynamics of proteins--they have not been used for structure determination. Here we show for crystals of the integral membrane protein complex photosystem II that lattice disorder increases the information content and the resolution of the diffraction pattern well beyond the 4.5-ångström limit of measurable Bragg peaks, which allows us to phase the pattern directly. Using the molecular envelope conventionally determined at 4.5 ångströms as a constraint, we obtain a static image of the photosystem II dimer at a resolution of 3.5 ångströms. This result shows that continuous diffraction can be used to overcome what have long been supposed to be the resolution limits of macromolecular crystallography, using a method that exploits commonly encountered imperfect crystals and enables model-free phasing.

  14. Macromolecular diffractive imaging using imperfect crystals

    NASA Astrophysics Data System (ADS)

    Ayyer, Kartik; Yefanov, Oleksandr M.; Oberthür, Dominik; Roy-Chowdhury, Shatabdi; Galli, Lorenzo; Mariani, Valerio; Basu, Shibom; Coe, Jesse; Conrad, Chelsie E.; Fromme, Raimund; Schaffer, Alexander; Dörner, Katerina; James, Daniel; Kupitz, Christopher; Metz, Markus; Nelson, Garrett; Xavier, Paulraj Lourdu; Beyerlein, Kenneth R.; Schmidt, Marius; Sarrou, Iosifina; Spence, John C. H.; Weierstall, Uwe; White, Thomas A.; Yang, Jay-How; Zhao, Yun; Liang, Mengning; Aquila, Andrew; Hunter, Mark S.; Robinson, Joseph S.; Koglin, Jason E.; Boutet, Sébastien; Fromme, Petra; Barty, Anton; Chapman, Henry N.

    2016-02-01

    The three-dimensional structures of macromolecules and their complexes are mainly elucidated by X-ray protein crystallography. A major limitation of this method is access to high-quality crystals, which is necessary to ensure X-ray diffraction extends to sufficiently large scattering angles and hence yields information of sufficiently high resolution with which to solve the crystal structure. The observation that crystals with reduced unit-cell volumes and tighter macromolecular packing often produce higher-resolution Bragg peaks suggests that crystallographic resolution for some macromolecules may be limited not by their heterogeneity, but by a deviation of strict positional ordering of the crystalline lattice. Such displacements of molecules from the ideal lattice give rise to a continuous diffraction pattern that is equal to the incoherent sum of diffraction from rigid individual molecular complexes aligned along several discrete crystallographic orientations and that, consequently, contains more information than Bragg peaks alone. Although such continuous diffraction patterns have long been observed—and are of interest as a source of information about the dynamics of proteins—they have not been used for structure determination. Here we show for crystals of the integral membrane protein complex photosystem II that lattice disorder increases the information content and the resolution of the diffraction pattern well beyond the 4.5-ångström limit of measurable Bragg peaks, which allows us to phase the pattern directly. Using the molecular envelope conventionally determined at 4.5 ångströms as a constraint, we obtain a static image of the photosystem II dimer at a resolution of 3.5 ångströms. This result shows that continuous diffraction can be used to overcome what have long been supposed to be the resolution limits of macromolecular crystallography, using a method that exploits commonly encountered imperfect crystals and enables model-free phasing.

  15. Macromolecular diffractive imaging using imperfect crystals.

    PubMed

    Ayyer, Kartik; Yefanov, Oleksandr M; Oberthür, Dominik; Roy-Chowdhury, Shatabdi; Galli, Lorenzo; Mariani, Valerio; Basu, Shibom; Coe, Jesse; Conrad, Chelsie E; Fromme, Raimund; Schaffer, Alexander; Dörner, Katerina; James, Daniel; Kupitz, Christopher; Metz, Markus; Nelson, Garrett; Xavier, Paulraj Lourdu; Beyerlein, Kenneth R; Schmidt, Marius; Sarrou, Iosifina; Spence, John C H; Weierstall, Uwe; White, Thomas A; Yang, Jay-How; Zhao, Yun; Liang, Mengning; Aquila, Andrew; Hunter, Mark S; Robinson, Joseph S; Koglin, Jason E; Boutet, Sébastien; Fromme, Petra; Barty, Anton; Chapman, Henry N

    2016-02-11

    The three-dimensional structures of macromolecules and their complexes are mainly elucidated by X-ray protein crystallography. A major limitation of this method is access to high-quality crystals, which is necessary to ensure X-ray diffraction extends to sufficiently large scattering angles and hence yields information of sufficiently high resolution with which to solve the crystal structure. The observation that crystals with reduced unit-cell volumes and tighter macromolecular packing often produce higher-resolution Bragg peaks suggests that crystallographic resolution for some macromolecules may be limited not by their heterogeneity, but by a deviation of strict positional ordering of the crystalline lattice. Such displacements of molecules from the ideal lattice give rise to a continuous diffraction pattern that is equal to the incoherent sum of diffraction from rigid individual molecular complexes aligned along several discrete crystallographic orientations and that, consequently, contains more information than Bragg peaks alone. Although such continuous diffraction patterns have long been observed--and are of interest as a source of information about the dynamics of proteins--they have not been used for structure determination. Here we show for crystals of the integral membrane protein complex photosystem II that lattice disorder increases the information content and the resolution of the diffraction pattern well beyond the 4.5-ångström limit of measurable Bragg peaks, which allows us to phase the pattern directly. Using the molecular envelope conventionally determined at 4.5 ångströms as a constraint, we obtain a static image of the photosystem II dimer at a resolution of 3.5 ångströms. This result shows that continuous diffraction can be used to overcome what have long been supposed to be the resolution limits of macromolecular crystallography, using a method that exploits commonly encountered imperfect crystals and enables model-free phasing. PMID

  16. Special quasirandom structures for perovskite solid solutions.

    PubMed

    Jiang, Zhijun; Nahas, Yousra; Xu, Bin; Prosandeev, Sergey; Wang, Dawei; Bellaiche, Laurent

    2016-11-30

    Special quasirandom structures (SQS) are presently generated for disordered (A'1-x [Formula: see text] x )BX3 and A(B'1-x [Formula: see text] x )X3 perovskite solid solutions, with x  =  1/2 as well as 1/3 and 2/3. These SQS configurations are obtained by imposing that the so-called Cowley parameters are as close to zero as possible for the three nearest neighboring shells. Moreover, these SQS configurations are slightly larger in size than those available in the literature for x  =  1/2, mostly because of the current capabilities of atomistic techniques. They are used here within effective Hamiltonian schemes to predict various properties, which are then compared to those associated with large random supercells, in a variety of compounds, namely (Ba1-x Sr x )TiO3, Pb(Zr1-x Ti x )O3, Pb(Sc0.5Nb0.5)O3, Ba(Zr1-x Ti x )O3, Pb(Mg1/3Nb2/3)O3 and (Bi1-x Nd x )FeO3. It is found that these SQS configurations can reproduce many properties of large random supercells of most of these disordered perovskite alloys, below some finite material-dependent temperature. Examples of these properties are electrical polarization, anti-phase and in-phase octahedral tiltings, antipolar motions, antiferromagnetism, strain, piezoelectric coefficients, dielectric response, specific heat and even the formation of polar nanoregions (PNRs) in some relaxors. Some limitations of these SQS configurations are also pointed out and explained. PMID:27661191

  17. Special quasirandom structures for perovskite solid solutions

    NASA Astrophysics Data System (ADS)

    Jiang, Zhijun; Nahas, Yousra; Xu, Bin; Prosandeev, Sergey; Wang, Dawei; Bellaiche, Laurent

    2016-11-01

    Special quasirandom structures (SQS) are presently generated for disordered (A‧1-x {{\\text{A}}\\prime \\prime} x )BX3 and A(B‧1-x {{\\text{B}}\\prime \\prime} x )X3 perovskite solid solutions, with x  =  1/2 as well as 1/3 and 2/3. These SQS configurations are obtained by imposing that the so-called Cowley parameters are as close to zero as possible for the three nearest neighboring shells. Moreover, these SQS configurations are slightly larger in size than those available in the literature for x  =  1/2, mostly because of the current capabilities of atomistic techniques. They are used here within effective Hamiltonian schemes to predict various properties, which are then compared to those associated with large random supercells, in a variety of compounds, namely (Ba1-x Sr x )TiO3, Pb(Zr1-x Ti x )O3, Pb(Sc0.5Nb0.5)O3, Ba(Zr1-x Ti x )O3, Pb(Mg1/3Nb2/3)O3 and (Bi1-x Nd x )FeO3. It is found that these SQS configurations can reproduce many properties of large random supercells of most of these disordered perovskite alloys, below some finite material-dependent temperature. Examples of these properties are electrical polarization, anti-phase and in-phase octahedral tiltings, antipolar motions, antiferromagnetism, strain, piezoelectric coefficients, dielectric response, specific heat and even the formation of polar nanoregions (PNRs) in some relaxors. Some limitations of these SQS configurations are also pointed out and explained.

  18. A strategy for dissecting the architectures of native macromolecular assemblies

    PubMed Central

    Shi, Yi; Pellarin, Riccardo; Fridy, Peter C.; Fernandez-Martinez, Javier; Thompson, Mary K.; Li, Yinyin; Wang, Qing Jun; Sali, Andrej; Rout, Michael P.; Chait, Brian T.

    2015-01-01

    Despite the central role of large multi-protein complexes in many biological processes, it remains challenging to elucidate their structures and particularly problematic to define the structures of native macromolecular assemblies, which are often of low abundance. Here, we present a strategy for isolating such complexes and for extracting distance restraints that allow the determination of their molecular architectures. The method was optimized to allow facile use of the extensive global resources of GFP-tagged transgenic cells and animals. PMID:26436480

  19. ProteoPlex: stability optimization of macromolecular complexes by sparse-matrix screening of chemical space.

    PubMed

    Chari, Ashwin; Haselbach, David; Kirves, Jan-Martin; Ohmer, Juergen; Paknia, Elham; Fischer, Niels; Ganichkin, Oleg; Möller, Vanessa; Frye, Jeremiah J; Petzold, Georg; Jarvis, Marc; Tietzel, Michael; Grimm, Clemens; Peters, Jan-Michael; Schulman, Brenda A; Tittmann, Kai; Markl, Jürgen; Fischer, Utz; Stark, Holger

    2015-09-01

    Molecular machines or macromolecular complexes are supramolecular assemblies of biomolecules with a variety of functions. Structure determination of these complexes in a purified state is often tedious owing to their compositional complexity and the associated relative structural instability. To improve the stability of macromolecular complexes in vitro, we present a generic method that optimizes the stability, homogeneity and solubility of macromolecular complexes by sparse-matrix screening of their thermal unfolding behavior in the presence of various buffers and small molecules. The method includes the automated analysis of thermal unfolding curves based on a biophysical unfolding model for complexes. We found that under stabilizing conditions, even large multicomponent complexes reveal an almost ideal two-state unfolding behavior. We envisage an improved biochemical understanding of purified macromolecules as well as a substantial boost in successful macromolecular complex structure determination by both X-ray crystallography and cryo-electron microscopy.

  20. ProteoPlex: stability optimization of macromolecular complexes by sparse-matrix screening of chemical space.

    PubMed

    Chari, Ashwin; Haselbach, David; Kirves, Jan-Martin; Ohmer, Juergen; Paknia, Elham; Fischer, Niels; Ganichkin, Oleg; Möller, Vanessa; Frye, Jeremiah J; Petzold, Georg; Jarvis, Marc; Tietzel, Michael; Grimm, Clemens; Peters, Jan-Michael; Schulman, Brenda A; Tittmann, Kai; Markl, Jürgen; Fischer, Utz; Stark, Holger

    2015-09-01

    Molecular machines or macromolecular complexes are supramolecular assemblies of biomolecules with a variety of functions. Structure determination of these complexes in a purified state is often tedious owing to their compositional complexity and the associated relative structural instability. To improve the stability of macromolecular complexes in vitro, we present a generic method that optimizes the stability, homogeneity and solubility of macromolecular complexes by sparse-matrix screening of their thermal unfolding behavior in the presence of various buffers and small molecules. The method includes the automated analysis of thermal unfolding curves based on a biophysical unfolding model for complexes. We found that under stabilizing conditions, even large multicomponent complexes reveal an almost ideal two-state unfolding behavior. We envisage an improved biochemical understanding of purified macromolecules as well as a substantial boost in successful macromolecular complex structure determination by both X-ray crystallography and cryo-electron microscopy. PMID:26237227

  1. Bridging the solution divide: comprehensive structural analyses of dynamic RNA, DNA, and protein assemblies by small angle X-ray scattering

    PubMed Central

    Rambo, Robert P.; Tainer, John A.

    2010-01-01

    Summary Small-Angle X-ray Scattering (SAXS) is changing how we perceive biological structures, because it reveals dynamic macromolecular conformations and assemblies in solution. SAXS information captures thermodynamic ensembles, enhances static structures detailed by high-resolution methods, uncovers commonalities among diverse macromolecules, and helps define biological mechanisms. SAXS-based experiments on RNA riboswitches and ribozymes and on DNA-protein complexes including DNA-PK and p53 discover flexibilities that better define structure-function relationships. Furthermore, SAXS results suggest conformational variation is a general functional feature of macromolecules. Thus, accurate structural analyses will require a comprehensive approach that assesses both flexibility, as seen by SAXS, and detail, as determined by X-ray crystallography and NMR. Here, we review recent SAXS computational tools, technologies, and applications to nucleic acids and related structures. PMID:20097063

  2. Macromolecular Crystal Growth by Means of Microfluidics

    NASA Technical Reports Server (NTRS)

    vanderWoerd, Mark; Ferree, Darren; Spearing, Scott; Monaco, Lisa; Molho, Josh; Spaid, Michael; Brasseur, Mike; Curreri, Peter A. (Technical Monitor)

    2002-01-01

    We have performed a feasibility study in which we show that chip-based, microfluidic (LabChip(TM)) technology is suitable for protein crystal growth. This technology allows for accurate and reliable dispensing and mixing of very small volumes while minimizing bubble formation in the crystallization mixture. The amount of (protein) solution remaining after completion of an experiment is minimal, which makes this technique efficient and attractive for use with proteins, which are difficult or expensive to obtain. The nature of LabChip(TM) technology renders it highly amenable to automation. Protein crystals obtained in our initial feasibility studies were of excellent quality as determined by X-ray diffraction. Subsequent to the feasibility study, we designed and produced the first LabChip(TM) device specifically for protein crystallization in batch mode. It can reliably dispense and mix from a range of solution constituents into two independent growth wells. We are currently testing this design to prove its efficacy for protein crystallization optimization experiments. In the near future we will expand our design to incorporate up to 10 growth wells per LabChip(TM) device. Upon completion, additional crystallization techniques such as vapor diffusion and liquid-liquid diffusion will be accommodated. Macromolecular crystallization using microfluidic technology is envisioned as a fully automated system, which will use the 'tele-science' concept of remote operation and will be developed into a research facility for the International Space Station as well as on the ground.

  3. Quantum chemistry of macromolecular shape

    NASA Astrophysics Data System (ADS)

    Mezey, Paul G.

    Some of the new developments in the quantum-chemical study of macromolecular shapes are reviewed, with special emphasis on the additive fuzzy electron density fragmentation methods and on the algebraic-topological shape group analysis of global and local shape features of fuzzy three-dimensional bodies of electron densities of macromolecules. Earlier applications of these methods to actual macromolecules are reviewed, including studies on the anticancer drug taxol, the proteins bovine insulin and HIV protease, and other macromolecules. The results of test calculations establishing the accuracy of these methods are also reviewed. The spherically weighted affine transformation technique is described and proposed for the deformation of electron densities approximating the changes occurring in small conformational displacements of atomic nuclei in macromolecules.

  4. Interfacial inhibition of macromolecular interactions: nature's paradigm for drug discovery.

    PubMed

    Pommier, Yves; Cherfils, Jacqueline

    2005-03-01

    One of nature's strategies for interfering with molecular interactions is to trap macromolecules in transition states with their partners in dead-end complexes that are unable to complete their biological function. This type of inhibition, which we refer to as "interfacial inhibition", is illustrated by two natural inhibitors, brefeldin A (BFA) and camptothecin (CPT), whose modes of action have been elucidated fully in structural studies. Interfacial inhibition occurs at the protein-protein interface in the case of BFA and at the protein-DNA interface in the case of CPT. In both systems, the drugs take advantage of transient structural and energetic conditions created by the macromolecular complex, which give rise to "hot-spots" for drug binding. In addition to these examples, several natural compounds such as forskolin, tubulin inhibitors and immunophilins target protein interfaces. We propose that interfacial inhibition is a paradigm for the discovery of drugs that interfere with macromolecular complexes.

  5. Temperature-dependent macromolecular X-ray crystallography

    SciTech Connect

    Weik, Martin Colletier, Jacques-Philippe

    2010-04-01

    The dynamical behaviour of crystalline macromolecules and their surrounding solvent as a function of cryo-temperature is reviewed. X-ray crystallography provides structural details of biological macromolecules. Whereas routine data are collected close to 100 K in order to mitigate radiation damage, more exotic temperature-controlled experiments in a broader temperature range from 15 K to room temperature can provide both dynamical and structural insights. Here, the dynamical behaviour of crystalline macromolecules and their surrounding solvent as a function of cryo-temperature is reviewed. Experimental strategies of kinetic crystallography are discussed that have allowed the generation and trapping of macromolecular intermediate states by combining reaction initiation in the crystalline state with appropriate temperature profiles. A particular focus is on recruiting X-ray-induced changes for reaction initiation, thus unveiling useful aspects of radiation damage, which otherwise has to be minimized in macromolecular crystallography.

  6. Temperature-dependent macromolecular X-ray crystallography

    PubMed Central

    Weik, Martin; Colletier, Jacques-Philippe

    2010-01-01

    X-ray crystallography provides structural details of biological macromolecules. Whereas routine data are collected close to 100 K in order to mitigate radiation damage, more exotic temperature-controlled experiments in a broader temperature range from 15 K to room temperature can provide both dynamical and structural insights. Here, the dynamical behaviour of crystalline macromolecules and their surrounding solvent as a function of cryo-temperature is reviewed. Experimental strategies of kinetic crystallography are discussed that have allowed the generation and trapping of macromolecular intermediate states by combining reaction initiation in the crystalline state with appropriate temperature profiles. A particular focus is on recruiting X-ray-induced changes for reaction initiation, thus unveiling useful aspects of radiation damage, which otherwise has to be minimized in macromolecular crystallography. PMID:20382997

  7. ‘Broken symmetries’ in macromolecular crystallography: phasing from unmerged data

    PubMed Central

    Schiltz, Marc; Bricogne, Gérard

    2010-01-01

    The space-group symmetry of a crystal structure imposes a point-group symmetry on its diffraction pattern, giving rise to so-called symmetry-equivalent reflections. Instances in macromolecular crystallography are discussed in which the sym­metry in reciprocal space is broken, i.e. where symmetry-related reflections are no longer equivalent. Such a situation occurs when the sample suffers from site-specific radiation damage during the X-ray measurements. Another example of broken symmetry arises from the polarization anisotropy of anomalous scattering. In these cases, the genuine intensity differences between symmetry-related reflections can be exploited to yield phase information in the structure-solution process. In this approach, the usual separation of the data merging and phasing steps is abandoned. The data are kept unmerged down to the Harker construction, where the symmetry-breaking effects are explicitly modelled and refined and become a source of supplementary phase information. PMID:20382998

  8. Expanding to fill the gap: a possible role for inert biopolymers in regulating the extent of the 'macromolecular crowding' effect.

    PubMed

    Hall, Damien; Dobson, Christopher M

    2006-05-15

    We discuss the potential for inert biopolymers existing in cells to play a role in regulating the macromolecular crowding effect via their ability to undergo shape changing structural transitions. We have explored this possibility by the use of theory and experiment. The theoretical component utilized Monte-Carlo based simulations to examine the folding of a hypothetical protein in a concentrated environment of hard spheres which are themselves capable of reversible expansion and contraction. The experimental component of the study involved examination of the effect of different sized crowding agents on the thermally induced denaturation of cytochrome c [in phosphate buffered saline solution containing 1.0M guanidinium hydrochloride at pH 7.0]. On the basis of our findings we suggest that in a crowded solution environment the presence of a non-reactive polymer capable of reversible expansion/contraction via folding and unfolding may alter the excluded volume component of the solution. This ability would confer on the non-reactive polymer a novel role in influencing other processes in solution affected by macromolecular crowding.

  9. WAXS studies of the structural diversity of hemoglobin in solution.

    SciTech Connect

    Makowski, L.; Bardhan, J.; Gore, D.; Lal, J.; Mandava, S.; Park, S.; Rodi, D. J.; Ho, N. T.; Ho, C.; Fischetti, R. F.

    2011-01-01

    Specific ligation states of hemoglobin are, when crystallized, capable of taking on multiple quaternary structures. The relationship between these structures, captured in crystal lattices, and hemoglobin structure in solution remains uncertain. Wide-angle X-ray solution scattering (WAXS) is a sensitive probe of protein structure in solution that can distinguish among similar structures and has the potential to contribute to these issues. We used WAXS to assess the relationships among the structures of human and bovine hemoglobins in different liganded forms in solution. WAXS data readily distinguished among the various forms of hemoglobins. WAXS patterns confirm some of the relationships among hemoglobin structures that have been defined through crystallography and NMR and extend others. For instance, methemoglobin A in solution is, as expected, nearly indistinguishable from HbCO A. Interestingly, for bovine hemoglobin, the differences between deoxy-Hb, methemoglobin and HbCO are smaller than the corresponding differences in human hemoglobin. WAXS data were also used to assess the spatial extent of structural fluctuations of various hemoglobins in solution. Dynamics has been implicated in allosteric control of hemoglobin, and increased dynamics has been associated with lowered oxygen affinity. Consistent with that notion, WAXS patterns indicate that deoxy-Hb A exhibits substantially larger structural fluctuations than HbCO A. Comparisons between the observed WAXS patterns and those predicted on the basis of atomic coordinate sets suggest that the structures of Hb in different liganded forms exhibit clear differences from known crystal structure.

  10. Protein folding, stability, and solvation structure in osmolyte solutions hydrophobicity

    NASA Astrophysics Data System (ADS)

    Montgomery Pettitt, B.

    2008-03-01

    The hydrophobic effect between solutes in aqueous solutions plays a central role in our understanding of recognition and folding of proteins and self assembly of lipids. Hydrophobicity induces nonideal solution behavior which plays a role in many aspects of biophysics. Work on the use of small biochemical compounds to crowd protein solutions indicates that a quantitative description of their non-ideal behavior is possible and straightforward. Here, we will show what the structural origin of this non-ideal solution behavior is from expression derived from a semi grand ensemble approach. We discuss the consequences of these findings regarding protein folding stability and solvation in crowded solutions through a structural analysis of the m-value or the change in free energy difference of a macromolecule in solution with respect to the concentration of a third component. This effect has recently been restudied and new mechanisms proposed for its origins in terms of transfer free energies and hydrophobicity.

  11. International summer school on macromolecular crystallographic computing. Final report

    SciTech Connect

    1998-08-01

    The School was the seventh in a series of International Union of Crystallography (IUCr) Crystallographic Symposia. The format of the School was formal lectures in the morning, tutorials in the afternoon, and software demonstrations and more lectures in the evening. The full program which left both the organizers and attendees exhausted, reflects the current state of excitement in the field of macromolecular structure determination using the technique of X-ray crystallography. The new and improved technologies and techniques described in these Proceedings are contributing to that growth and at the same time, as pointed out in the paper given by Sussman, creating challenges for the Protein Data Bank (PDB). As the School progressed, the authors were struck by the similarities to events which took place in small molecule crystallography beginning some 20 to 25 years ago. Growth then was fueled by the advent of new algorithms, affordable computer hardware, and good software. So it is today for macromolecular crystallography, but with the added bonus of the Internet which is changing how scientist conduct their research. Flack presented this view as part of his on-going contribution to how crystallographers use the Internet. After presentations discussing structures en masse they returned to the more traditional mode of presentation which parallels the determination of a single macromolecular structure: data collection -- phasing -- model building and visualization -- refinement.

  12. Controlled architecture for improved macromolecular memory within polymer networks.

    PubMed

    DiPasquale, Stephen A; Byrne, Mark E

    2016-08-01

    This brief review analyzes recent developments in the field of living/controlled polymerization and the potential of this technique for creating imprinted polymers with highly structured architecture with macromolecular memory. As a result, it is possible to engineer polymers at the molecular level with increased homogeneity relating to enhanced template binding and transport. Only recently has living/controlled polymerization been exploited to decrease heterogeneity and substantially improve the efficiency of the imprinting process for both highly and weakly crosslinked imprinted polymers. Living polymerization can be utilized to create imprinted networks that are vastly more efficient than similar polymers produced using conventional free radical polymerization, and these improvements increase the role that macromolecular memory can play in the design and engineering of new drug delivery and sensing platforms. PMID:27322505

  13. Biophysical Highlights from 54 Years of Macromolecular Crystallography

    PubMed Central

    Richardson, Jane S.; Richardson, David C.

    2014-01-01

    The United Nations has declared 2014 the International Year of Crystallography, and in commemoration, this review features a selection of 54 notable macromolecular crystal structures that have illuminated the field of biophysics in the 54 years since the first excitement of the myoglobin and hemoglobin structures in 1960. Chronological by publication of the earliest solved structure, each illustrated entry briefly describes key concepts or methods new at the time and key later work leveraged by knowledge of the three-dimensional atomic structure. PMID:24507592

  14. The solvent component of macromolecular crystals

    SciTech Connect

    Weichenberger, Christian X.; Kantardjieff, Katherine; Rupp, Bernhard

    2015-04-30

    On average, the mother liquor or solvent and its constituents occupy about 50% of a macromolecular crystal. Ordered as well as disordered solvent components need to be accurately accounted for in modelling and refinement, often with considerable complexity. The mother liquor from which a biomolecular crystal is grown will contain water, buffer molecules, native ligands and cofactors, crystallization precipitants and additives, various metal ions, and often small-molecule ligands or inhibitors. On average, about half the volume of a biomolecular crystal consists of this mother liquor, whose components form the disordered bulk solvent. Its scattering contributions can be exploited in initial phasing and must be included in crystal structure refinement as a bulk-solvent model. Concomitantly, distinct electron density originating from ordered solvent components must be correctly identified and represented as part of the atomic crystal structure model. Herein, are reviewed (i) probabilistic bulk-solvent content estimates, (ii) the use of bulk-solvent density modification in phase improvement, (iii) bulk-solvent models and refinement of bulk-solvent contributions and (iv) modelling and validation of ordered solvent constituents. A brief summary is provided of current tools for bulk-solvent analysis and refinement, as well as of modelling, refinement and analysis of ordered solvent components, including small-molecule ligands.

  15. Macromolecular structural changes in bituminous coals during extraction and solubilization. Quarterly technical progress report, 1 September 1981-1 December 1981

    SciTech Connect

    Peppas, N.A.

    1981-01-01

    Data are presented of the effect of coal pretreatment (extraction, flotation etc.) and porous structure on the apparent and effective swelling of several coals by four swelling agents. Analysis of the pore structure was achieved by mercury porosimetry and pyconometry. The effect of retained solvent in the pores is more prominent in the determination of the equilibrium coal volume fraction and the actual molecular weight between crosslinks, M/sub c/.

  16. Quantifying macromolecular conformational transition pathways

    NASA Astrophysics Data System (ADS)

    Seyler, Sean; Kumar, Avishek; Thorpe, Michael; Beckstein, Oliver

    2015-03-01

    Diverse classes of proteins function through large-scale conformational changes that are challenging for computer simulations. A range of fast path-sampling techniques have been used to generate transitions, but it has been difficult to compare paths from (and assess the relative strengths of) different methods. We introduce a comprehensive method (pathway similarity analysis, PSA) for quantitatively characterizing and comparing macromolecular pathways. The Hausdorff and Fréchet metrics (known from computational geometry) are used to quantify the degree of similarity between polygonal curves in configuration space. A strength of PSA is its use of the full information available from the 3 N-dimensional configuration space trajectory without requiring additional specific knowledge about the system. We compare a sample of eleven different methods for the closed-to-open transitions of the apo enzyme adenylate kinase (AdK) and also apply PSA to an ensemble of 400 AdK trajectories produced by dynamic importance sampling MD and the Geometrical Pathways algorithm. We discuss the method's potential to enhance our understanding of transition path sampling methods, validate them, and help guide future research toward deeper physical insights into conformational transitions.

  17. Complexation of Actinides in Solution: Thermodynamic Measurementsand Structural Characterization

    SciTech Connect

    Rao, L.

    2007-02-01

    This paper presents a brief introduction of the studies of actinide complexation in solution at Lawrence Berkeley National Laboratory. An integrated approach of thermodynamic measurements and structural characterization is taken to obtain fundamental understanding of actinide complexation in solution that is of importance in predicting the behavior of actinides in separation processes and environmental transport.

  18. SFCHECK: a unified set of procedures for evaluating the quality of macromolecular structure-factor data and their agreement with the atomic model.

    PubMed

    Vaguine, A A; Richelle, J; Wodak, S J

    1999-01-01

    In this paper we present SFCHECK, a stand-alone software package that features a unified set of procedures for evaluating the structure-factor data obtained from X-ray diffraction experiments and for assessing the agreement of the atomic coordinates with these data. The evaluation is performed completely automatically, and produces a concise PostScript pictorial output similar to that of PROCHECK [Laskowski, MacArthur, Moss & Thornton (1993). J. Appl. Cryst. 26, 283-291], greatly facilitating visual inspection of the results. The required inputs are the structure-factor amplitudes and the atomic coordinates. Having those, the program summarizes relevant information on the deposited structure factors and evaluates their quality using criteria such as data completeness, structure-factor uncertainty and the optical resolution computed from the Patterson origin peak. The dependence of various parameters on the nominal resolution (d spacing) is also given. To evaluate the global agreement of the atomic model with the experimental data, the program recomputes the R factor, the correlation coefficient between observed and calculated structure-factor amplitudes and Rfree (when appropriate). In addition, it gives several estimates of the average error in the atomic coordinates. The local agreement between the model and the electron-density map is evaluated on a per-residue basis, considering separately the macromolecule backbone and side-chain atoms, as well as solvent atoms and heterogroups. Among the criteria are the normalized average atomic displacement, the local density correlation coefficient and the polymer chain connectivity. The possibility of computing these criteria using the omit-map procedure is also provided. The described software should be a valuable tool in monitoring the refinement procedure and in assessing structures deposited in databases.

  19. The promise of macromolecular crystallization in microfluidic chips

    NASA Technical Reports Server (NTRS)

    van der Woerd, Mark; Ferree, Darren; Pusey, Marc

    2003-01-01

    Microfluidics, or lab-on-a-chip technology, is proving to be a powerful, rapid, and efficient approach to a wide variety of bioanalytical and microscale biopreparative needs. The low materials consumption, combined with the potential for packing a large number of experiments in a few cubic centimeters, makes it an attractive technique for both initial screening and subsequent optimization of macromolecular crystallization conditions. Screening operations, which require a macromolecule solution with a standard set of premixed solutions, are relatively straightforward and have been successfully demonstrated in a microfluidics platform. Optimization methods, in which crystallization solutions are independently formulated from a range of stock solutions, are considerably more complex and have yet to be demonstrated. To be competitive with either approach, a microfluidics system must offer ease of operation, be able to maintain a sealed environment over several weeks to months, and give ready access for the observation and harvesting of crystals as they are grown.

  20. Taking X-ray Diffraction to the Limit: Macromolecular Structures from Femtosecond X-ray Pulses and Diffraction Microscopy of Cells with Synchrotron Radiation

    SciTech Connect

    Chapman, H N; Miao, J; Kirz, J; Sayre, D; Hodgson, K O

    2003-10-01

    The methodology of X-ray crystallography has recently been successfully extended to the structure determination of non-crystalline specimens. The phase problem was solved by using the oversampling method, which takes advantage of ''continuous'' diffraction pattern from non-crystalline specimens. Here we review the principle of this newly developed technique and discuss the ongoing experiments of imaging non-periodic objects, like cells and cellular structures using coherent and bright X-rays from the 3rd generation synchrotron radiation. In the longer run, the technique may be applied to image single biomolecules by using the anticipated X-ray free electron lasers. Computer simulations have so far demonstrated two important steps: (1) by using an extremely intense femtosecond X-ray pulse, a diffraction pattern can be recorded from a macromolecule before radiation damage manifests itself, and (2) the phase information can be ab initio retrieved from a set of calculated noisy diffraction patterns of single protein molecules.

  1. IRIS explorer software for radial-depth cueing reovirus particles and other macromolecular structures determined by cryoelectron microscopy and image reconstruction.

    PubMed

    Spencer, S M; Sgro, J Y; Dryden, K A; Baker, T S; Nibert, M L

    1997-10-01

    Structures of biological macromolecules determined by transmission cryoelectron microscopy (cryo-TEM) and three-dimensional image reconstruction are often displayed as surface-shaded representations with depth cueing along the viewed direction (Z cueing). Depth cueing to indicate distance from the center of virus particles (radial-depth cueing, or R cueing) has also been used. We have found that a style of R cueing in which color is applied in smooth or discontinuous gradients using the IRIS Explorer software is an informative technique for displaying the structures of virus particles solved by cryo-TEM and image reconstruction. To develop and test these methods, we used existing cryo-TEM reconstructions of mammalian reovirus particles. The newly applied visualization techniques allowed us to discern several new structural features, including sites in the inner capsid through which the viral mRNAs may be extruded after they are synthesized by the reovirus transcriptase complexes. To demonstrate the broad utility of the methods, we also applied them to cryo-TEM reconstructions of human rhinovirus, native and swollen forms of cowpea chlorotic mottle virus, truncated core of pyruvate dehydrogenase complex from Saccharomyces cerevisiae, and flagellar filament of Salmonella typhimurium. We conclude that R cueing with color gradients is a useful tool for displaying virus particles and other macromolecules analyzed by cryo-TEM and image reconstruction.

  2. A Sco protein among the hypothetical proteins of Bacillus lehensis G1: Its 3D macromolecular structure and association with Cytochrome C Oxidase

    PubMed Central

    2014-01-01

    Background At least a quarter of any complete genome encodes for hypothetical proteins (HPs) which are largely non-similar to other known, well-characterized proteins. Predicting and solving their structures and functions is imperative to aid understanding of any given organism as a complete biological system. The present study highlights the primary effort to classify and cluster 1202 HPs of Bacillus lehensis G1 alkaliphile to serve as a platform to mine and select specific HP(s) to be studied further in greater detail. Results All HPs of B. lehensis G1 were grouped according to their predicted functions based on the presence of functional domains in their sequences. From the metal-binding group of HPs of the cluster, an HP termed Bleg1_2507 was discovered to contain a thioredoxin (Trx) domain and highly-conserved metal-binding ligands represented by Cys69, Cys73 and His159, similar to all prokaryotic and eukaryotic Sco proteins. The built 3D structure of Bleg1_2507 showed that it shared the βαβαββ core structure of Trx-like proteins as well as three flanking β-sheets, a 310 –helix at the N-terminus and a hairpin structure unique to Sco proteins. Docking simulations provided an interesting view of Bleg1_2507 in association with its putative cytochrome c oxidase subunit II (COXII) redox partner, Bleg1_2337, where the latter can be seen to hold its partner in an embrace, facilitated by hydrophobic and ionic interactions between the proteins. Although Bleg1_2507 shares relatively low sequence identity (47%) to BsSco, interestingly, the predicted metal-binding residues of Bleg1_2507 i.e. Cys-69, Cys-73 and His-159 were located at flexible active loops similar to other Sco proteins across biological taxa. This highlights structural conservation of Sco despite their various functions in prokaryotes and eukaryotes. Conclusions We propose that HP Bleg1_2507 is a Sco protein which is able to interact with COXII, its redox partner and therefore, may possess

  3. Viral capsomere structure, surface processes and growth kinetics in the crystallization of macromolecular crystals visualized by in situ atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Malkin, A. J.; Kuznetsov, Yu. G.; McPherson, A.

    2001-11-01

    In situ atomic force microscopy (AFM) was used to investigate surface evolution during the growth of single crystals of turnip yellow mosaic virus (TYMV), cucumber mosaic virus (CMV) and glucose isomerase. Growth of these crystals proceeded by two-dimensional (2D) nucleation. For glucose isomerase, from supersaturation dependencies of tangential step rates and critical step length, the kinetic coefficients of the steps and the surface free energy of the step edge were calculated for different crystallographic directions. The molecular structure of the step edges, the adsorption of individual virus particles and their aggregates, and the initial stages of formation of 2D nuclei on the surfaces of TYMV and CMV crystals were recorded. The surfaces of individual TYMV virions within crystals were visualized, and hexameric and pentameric capsomers of the T=3 capsids were clearly resolved. This, so far as we are aware, is the first direct visualization of the capsomere structure of a virus by AFM. In the course of recording the in situ development of the TYMV crystals, a profound restructuring of the surface arrangement was observed. This transformation was highly cooperative in nature, but the transitions were unambiguous and readily explicable in terms of an organized loss of classes of virus particles from specific lattice positions.

  4. Macromolecular crowding conditions enhance glycation and oxidation of whey proteins in ultrasound-induced Maillard reaction.

    PubMed

    Perusko, Marija; Al-Hanish, Ayah; Cirkovic Velickovic, Tanja; Stanic-Vucinic, Dragana

    2015-06-15

    High intensity ultrasound (HIUS) can promote Maillard reaction (MR). Macromolecular crowding conditions accelerate reactions and stabilise protein structure. The aim of this study was to investigate if combined application of ultrasound and macromolecular crowding can improve efficiency of MR. The presence of crowding agent (polyethylene glycol) significantly increased ultrasound-induced whey protein (WP) glycation by arabinose. An increase in glycation efficiency results only in slight change of WP structure. Macromolecular crowding intensifies oxidative modifications of WP, as well as formation of amyloid-like structures by enhancement of MR. Solubility at different pH, thermal stability and antioxidative capacity of glycated WP were increased, especially in the presence of crowding agent, compared to sonicated nonglycated proteins. The application of HIUS under crowding conditions can be a new approach for enhancement of reactions in general, enabling short processing time and mild conditions, while preserving protein structure and minimising protein aggregation.

  5. Macromolecular crowding conditions enhance glycation and oxidation of whey proteins in ultrasound-induced Maillard reaction.

    PubMed

    Perusko, Marija; Al-Hanish, Ayah; Cirkovic Velickovic, Tanja; Stanic-Vucinic, Dragana

    2015-06-15

    High intensity ultrasound (HIUS) can promote Maillard reaction (MR). Macromolecular crowding conditions accelerate reactions and stabilise protein structure. The aim of this study was to investigate if combined application of ultrasound and macromolecular crowding can improve efficiency of MR. The presence of crowding agent (polyethylene glycol) significantly increased ultrasound-induced whey protein (WP) glycation by arabinose. An increase in glycation efficiency results only in slight change of WP structure. Macromolecular crowding intensifies oxidative modifications of WP, as well as formation of amyloid-like structures by enhancement of MR. Solubility at different pH, thermal stability and antioxidative capacity of glycated WP were increased, especially in the presence of crowding agent, compared to sonicated nonglycated proteins. The application of HIUS under crowding conditions can be a new approach for enhancement of reactions in general, enabling short processing time and mild conditions, while preserving protein structure and minimising protein aggregation. PMID:25660883

  6. Building macromolecular assemblies by information-driven docking: introducing the HADDOCK multibody docking server.

    PubMed

    Karaca, Ezgi; Melquiond, Adrien S J; de Vries, Sjoerd J; Kastritis, Panagiotis L; Bonvin, Alexandre M J J

    2010-08-01

    Over the last years, large scale proteomics studies have generated a wealth of information of biomolecular complexes. Adding the structural dimension to the resulting interactomes represents a major challenge that classical structural experimental methods alone will have difficulties to confront. To meet this challenge, complementary modeling techniques such as docking are thus needed. Among the current docking methods, HADDOCK (High Ambiguity-Driven DOCKing) distinguishes itself from others by the use of experimental and/or bioinformatics data to drive the modeling process and has shown a strong performance in the critical assessment of prediction of interactions (CAPRI), a blind experiment for the prediction of interactions. Although most docking programs are limited to binary complexes, HADDOCK can deal with multiple molecules (up to six), a capability that will be required to build large macromolecular assemblies. We present here a novel web interface of HADDOCK that allows the user to dock up to six biomolecules simultaneously. This interface allows the inclusion of a large variety of both experimental and/or bioinformatics data and supports several types of cyclic and dihedral symmetries in the docking of multibody assemblies. The server was tested on a benchmark of six cases, containing five symmetric homo-oligomeric protein complexes and one symmetric protein-DNA complex. Our results reveal that, in the presence of either bioinformatics and/or experimental data, HADDOCK shows an excellent performance: in all cases, HADDOCK was able to generate good to high quality solutions and ranked them at the top, demonstrating its ability to model symmetric multicomponent assemblies. Docking methods can thus play an important role in adding the structural dimension to interactomes. However, although the current docking methodologies were successful for a vast range of cases, considering the variety and complexity of macromolecular assemblies, inclusion of some kind of

  7. Effects of Macromolecular Crowding on Genetic Networks

    PubMed Central

    Morelli, Marco J.; Allen, Rosalind J.; Rein ten Wolde, Pieter

    2011-01-01

    The intracellular environment is crowded with proteins, DNA, and other macromolecules. Under physiological conditions, macromolecular crowding can alter both molecular diffusion and the equilibria of bimolecular reactions and therefore is likely to have a significant effect on the function of biochemical networks. We propose a simple way to model the effects of macromolecular crowding on biochemical networks via an appropriate scaling of bimolecular association and dissociation rates. We use this approach, in combination with kinetic Monte Carlo simulations, to analyze the effects of crowding on a constitutively expressed gene, a repressed gene, and a model for the bacteriophage λ genetic switch, in the presence and absence of nonspecific binding of transcription factors to genomic DNA. Our results show that the effects of crowding are mainly caused by the shift of association-dissociation equilibria rather than the slowing down of protein diffusion, and that macromolecular crowding can have relevant and counterintuitive effects on biochemical network performance. PMID:22208186

  8. Ions in water: The microscopic structure of concentrated hydroxide solutions

    NASA Astrophysics Data System (ADS)

    Imberti, S.; Botti, A.; Bruni, F.; Cappa, G.; Ricci, M. A.; Soper, A. K.

    2005-05-01

    Neutron-diffraction data on aqueous solutions of hydroxides, at solute concentrations ranging from 1 solute per 12 water molecules to 1 solute per 3 water molecules, are analyzed by means of a Monte Carlo simulation (empirical potential structure refinement), in order to determine the hydration shell of the OH- in the presence of the smaller alkali metal ions. It is demonstrated that the symmetry argument between H+ and OH- cannot be used, at least in the liquid phase at such high concentrations, for determining the hydroxide hydration shell. Water molecules in the hydration shell of K+ orient their dipole moment at about 45° from the K+-water oxygen director, instead of radially as in the case of the Li+ and Na+ hydration shells. The K+-water oxygen radial distribution function shows a shallower first minimum compared to the other cation-water oxygen functions. The influence of the solutes on the water-water radial distribution functions is shown to have an effect on the water structure equivalent to an increase in the pressure of the water, depending on both ion concentration and ionic radius. The changes of the water structure in the presence of charged solutes and the differences among the hydration shells of the different cations are used to present a qualitative explanation of the observed cation mobility.

  9. Ions in water: the microscopic structure of concentrated hydroxide solutions.

    PubMed

    Imberti, S; Botti, A; Bruni, F; Cappa, G; Ricci, M A; Soper, A K

    2005-05-15

    Neutron-diffraction data on aqueous solutions of hydroxides, at solute concentrations ranging from 1 solute per 12 water molecules to 1 solute per 3 water molecules, are analyzed by means of a Monte Carlo simulation (empirical potential structure refinement), in order to determine the hydration shell of the OH- in the presence of the smaller alkali metal ions. It is demonstrated that the symmetry argument between H+ and OH- cannot be used, at least in the liquid phase at such high concentrations, for determining the hydroxide hydration shell. Water molecules in the hydration shell of K+ orient their dipole moment at about 45 degrees from the K+-water oxygen director, instead of radially as in the case of the Li+ and Na+ hydration shells. The K+-water oxygen radial distribution function shows a shallower first minimum compared to the other cation-water oxygen functions. The influence of the solutes on the water-water radial distribution functions is shown to have an effect on the water structure equivalent to an increase in the pressure of the water, depending on both ion concentration and ionic radius. The changes of the water structure in the presence of charged solutes and the differences among the hydration shells of the different cations are used to present a qualitative explanation of the observed cation mobility.

  10. Flexibility damps macromolecular crowding effects on protein folding dynamics: Application to the murine prion protein (121-231)

    NASA Astrophysics Data System (ADS)

    Bergasa-Caceres, Fernando; Rabitz, Herschel A.

    2014-01-01

    A model of protein folding kinetics is applied to study the combined effects of protein flexibility and macromolecular crowding on protein folding rate and stability. It is found that the increase in stability and folding rate promoted by macromolecular crowding is damped for proteins with highly flexible native structures. The model is applied to the folding dynamics of the murine prion protein (121-231). It is found that the high flexibility of the native isoform of the murine prion protein (121-231) reduces the effects of macromolecular crowding on its folding dynamics. The relevance of these findings for the pathogenic mechanism are discussed.

  11. Structural cluster analysis of chemical reactions in solution

    NASA Astrophysics Data System (ADS)

    Gallet, Grégoire A.; Pietrucci, Fabio

    2013-08-01

    We introduce a simple and general approach to the problem of clustering structures from atomic trajectories of chemical reactions in solution. By considering distance metrics which are invariant under permutation of identical atoms or molecules, we demonstrate that it is possible to automatically resolve as distinct structural clusters the configurations corresponding to reactants, products, and transition states, even in presence of atom-exchanges and of hundreds of solvent molecules. Our approach strongly simplifies the analysis of large trajectories and it opens the way to the construction of kinetic network models of activated processes in solution employing the available efficient schemes developed for proteins conformational ensembles.

  12. A local-optimization refinement algorithm in single particle analysis for macromolecular complex with multiple rigid modules.

    PubMed

    Shan, Hong; Wang, Zihao; Zhang, Fa; Xiong, Yong; Yin, Chang-Cheng; Sun, Fei

    2016-01-01

    Single particle analysis, which can be regarded as an average of signals from thousands or even millions of particle projections, is an efficient method to study the three-dimensional structures of biological macromolecules. An intrinsic assumption in single particle analysis is that all the analyzed particles must have identical composition and conformation. Thus specimen heterogeneity in either composition or conformation has raised great challenges for high-resolution analysis. For particles with multiple conformations, inaccurate alignments and orientation parameters will yield an averaged map with diminished resolution and smeared density. Besides extensive classification approaches, here based on the assumption that the macromolecular complex is made up of multiple rigid modules whose relative orientations and positions are in slight fluctuation around equilibriums, we propose a new method called as local optimization refinement to address this conformational heterogeneity for an improved resolution. The key idea is to optimize the orientation and shift parameters of each rigid module and then reconstruct their three-dimensional structures individually. Using simulated data of 80S/70S ribosomes with relative fluctuations between the large (60S/50S) and the small (40S/30S) subunits, we tested this algorithm and found that the resolutions of both subunits are significantly improved. Our method provides a proof-of-principle solution for high-resolution single particle analysis of macromolecular complexes with dynamic conformations.

  13. General analytical shakedown solution for structures with kinematic hardening materials

    NASA Astrophysics Data System (ADS)

    Guo, Baofeng; Zou, Zongyuan; Jin, Miao

    2016-04-01

    The effect of kinematic hardening behavior on the shakedown behaviors of structure has been investigated by performing shakedown analysis for some specific problems. The results obtained only show that the shakedown limit loads of structures with kinematic hardening model are larger than or equal to those with perfectly plastic model of the same initial yield stress. To further investigate the rules governing the different shakedown behaviors of kinematic hardening structures, the extended shakedown theorem for limited kinematic hardening is applied, the shakedown condition is then proposed, and a general analytical solution for the structural shakedown limit load is thus derived. The analytical shakedown limit loads for fully reversed cyclic loading and non-fully reversed cyclic loading are then given based on the general solution. The resulting analytical solution is applied to some specific problems: a hollow specimen subjected to tension and torsion, a flanged pipe subjected to pressure and axial force and a square plate with small central hole subjected to biaxial tension. The results obtained are compared with those in literatures, they are consistent with each other. Based on the resulting general analytical solution, rules governing the general effects of kinematic hardening behavior on the shakedown behavior of structure are clearly.

  14. Processes of ordered structure formation in polypeptide thin film solutions.

    SciTech Connect

    Botiz, I.; Schlaad, H.; Reiter, G.

    2010-06-17

    An experimental study is presented on the hierarchical assembly of {alpha}-helical block copolymers polystyrene-poly({gamma}-benzyl-L-glutamate) into anisotropic ordered structures. We transformed thin solid films into solutions through exposure to solvent vapor and studied the nucleation and growth of ordered three-dimensional structures in such solutions, with emphasis on the dependence of these processes on supersaturation with respect to the solubility limit. Interestingly, polymer solubility could be significantly influenced via variation of humidity in the surrounding gas phase. It is concluded that the interfacial tension between the ordered structures and the solution increased with humidity. The same effect was observed for other protic non-solvents in the surrounding gas phase and is attributed to a complexation of poly({gamma}-benzyl-L-glutamate) by protic non-solvent molecules (via hydrogen-bonding interactions). This change of polymer solubility was demonstrated to be reversible by addition or removal of small amounts of protic non-solvent in the surrounding gas phase. At a constant polymer concentration, ordered ellipsoidal structures could be dissolved by removing water or methanol present in the solution. Such structures formed once again when water or methanol was reintroduced via the vapor phase.

  15. Accounting for large amplitude protein deformation during in silico macromolecular docking.

    PubMed

    Bastard, Karine; Saladin, Adrien; Prévost, Chantal

    2011-02-22

    Rapid progress of theoretical methods and computer calculation resources has turned in silico methods into a conceivable tool to predict the 3D structure of macromolecular assemblages, starting from the structure of their separate elements. Still, some classes of complexes represent a real challenge for macromolecular docking methods. In these complexes, protein parts like loops or domains undergo large amplitude deformations upon association, thus remodeling the surface accessible to the partner protein or DNA. We discuss the problems linked with managing such rearrangements in docking methods and we review strategies that are presently being explored, as well as their limitations and success.

  16. Mixed Consolidation Solution for a Reinforced Concrete Structure

    NASA Astrophysics Data System (ADS)

    Lute, M.

    2016-06-01

    During the last years, reinforced concrete structures become subject for rehabilitation due to two factors: their long life span and large change in norms that leaded to a large increase of seismic loads in Eastern Europe. These lead to a necessity for rehabilitation of existing building stock in order to use them during their entire life span at the maximum potential. The present paper proposes a solution for rehabilitation for three reinforced concrete building of a hospital, that consumed a half of their life span and do not correspond anymore to present norms. The chosen solution is a combination between CFRP rehabilitation and increase of structural elements cross section in order to achieve the stiffness balance in the structure nodes that is required by present norms. As a further matter, correction in stiffness of local elements diminished the lateral drifts of the structure and improved the global seismic response of the building.

  17. Bringing macromolecular machinery to life using 3D animation.

    PubMed

    Iwasa, Janet H

    2015-04-01

    Over the past decade, there has been a rapid rise in the use of three-dimensional (3D) animation to depict molecular and cellular processes. Much of the growth in molecular animation has been in the educational arena, but increasingly, 3D animation software is finding its way into research laboratories. In this review, I will discuss a number of ways in which 3d animation software can play a valuable role in visualizing and communicating macromolecular structures and dynamics. I will also consider the challenges of using animation tools within the research sphere.

  18. Dealing with Multiple Solutions in Structural Vector Autoregressive Models.

    PubMed

    Beltz, Adriene M; Molenaar, Peter C M

    2016-01-01

    Structural vector autoregressive models (VARs) hold great potential for psychological science, particularly for time series data analysis. They capture the magnitude, direction of influence, and temporal (lagged and contemporaneous) nature of relations among variables. Unified structural equation modeling (uSEM) is an optimal structural VAR instantiation, according to large-scale simulation studies, and it is implemented within an SEM framework. However, little is known about the uniqueness of uSEM results. Thus, the goal of this study was to investigate whether multiple solutions result from uSEM analysis and, if so, to demonstrate ways to select an optimal solution. This was accomplished with two simulated data sets, an empirical data set concerning children's dyadic play, and modifications to the group iterative multiple model estimation (GIMME) program, which implements uSEMs with group- and individual-level relations in a data-driven manner. Results revealed multiple solutions when there were large contemporaneous relations among variables. Results also verified several ways to select the correct solution when the complete solution set was generated, such as the use of cross-validation, maximum standardized residuals, and information criteria. This work has immediate and direct implications for the analysis of time series data and for the inferences drawn from those data concerning human behavior.

  19. Enzymes as Green Catalysts for Precision Macromolecular Synthesis.

    PubMed

    Shoda, Shin-ichiro; Uyama, Hiroshi; Kadokawa, Jun-ichi; Kimura, Shunsaku; Kobayashi, Shiro

    2016-02-24

    The present article comprehensively reviews the macromolecular synthesis using enzymes as catalysts. Among the six main classes of enzymes, the three classes, oxidoreductases, transferases, and hydrolases, have been employed as catalysts for the in vitro macromolecular synthesis and modification reactions. Appropriate design of reaction including monomer and enzyme catalyst produces macromolecules with precisely controlled structure, similarly as in vivo enzymatic reactions. The reaction controls the product structure with respect to substrate selectivity, chemo-selectivity, regio-selectivity, stereoselectivity, and choro-selectivity. Oxidoreductases catalyze various oxidation polymerizations of aromatic compounds as well as vinyl polymerizations. Transferases are effective catalysts for producing polysaccharide having a variety of structure and polyesters. Hydrolases catalyzing the bond-cleaving of macromolecules in vivo, catalyze the reverse reaction for bond forming in vitro to give various polysaccharides and functionalized polyesters. The enzymatic polymerizations allowed the first in vitro synthesis of natural polysaccharides having complicated structures like cellulose, amylose, xylan, chitin, hyaluronan, and chondroitin. These polymerizations are "green" with several respects; nontoxicity of enzyme, high catalyst efficiency, selective reactions under mild conditions using green solvents and renewable starting materials, and producing minimal byproducts. Thus, the enzymatic polymerization is desirable for the environment and contributes to "green polymer chemistry" for maintaining sustainable society.

  20. Solution structure of At3g28950 from Arabidopsis thaliana

    PubMed Central

    de la Cruz, Norberto B.; Peterson, Francis C.; Volkman, Brian F.

    2009-01-01

    We determined the solution structure of At3g28950 from A. thaliana, a homolog of At5g39720, whose structure we solved earlier. The secondary structure of the 165-aa protein consists of a 5-strand anti-parallel β-barrel domain flanked by two α-helices and a 2-strand β-sheet; an additional free C-terminal α-helix extends into solution. Bioinformatic searches and analyses suggest that members of this growing set of structurally related proteins have been recruited to serve a wide variety of functions ranging from γ-glutamyl cyclotransferase activity, to participation in plant responses to chemical and biotic stimuli. Expression of a human homolog is elevated in bladder cancer tissues. Expression patterns for At3g28950 and its Arabidopsis paralogs suggest that each one evolved different a physiological role. The At3g28950 structure was solved as part of a structural genomics effort, and the results demonstrate how such a project can further understanding of genome evolution in addition to sequence-structure and structure-function relationships. PMID:18214976

  1. Extended structure of rat islet amyloid polypeptide in solution.

    PubMed

    Wei, Lei; Jiang, Ping; Manimekalai, Malathy Sony Subramanian; Hunke, Cornelia; Grüber, Gerhard; Pervushin, Konstantin; Mu, Yuguang

    2015-01-01

    The process of islet amyloid polypeptide (IAPP) formation and the prefibrillar oligomers are supposed to be one of the pathogenic agents causing pancreatic β-cell dysfunction. The human IAPP (hIAPP) aggregates easily and therefore, it is difficult to characterize its structural features by standard biophysical tools. The rat version of IAPP (rIAPP) that differs by six amino acids when compared with hIAPP, is not prone to aggregation and does not form amyloid fibrils. Similar to hIAPP it also demonstrates random-coiled nature in solution. The structural propensity of rIAPP has been studied as a hIAPP mimic in recent works. However, the overall shape of it in solution still remains elusive. Using small angle X-ray scattering (SAXS) measurements combined with nuclear magnetic resonance (NMR) and molecular dynamics simulations (MD) the solution structure of rIAPP was studied. An unambiguously extended structural model with a radius of gyration of 1.83 nm was determined from SAXS data. Consistent with previous studies, an overall random-coiled feature with residual helical propensity in the N-terminus was confirmed. Combined efforts are necessary to unambiguously resolve the structural features of intrinsic disordered proteins.

  2. Effects of cell turnover and leaky junctions on arterial macromolecular permeability - Relation to atherogenesis

    SciTech Connect

    Lin, Shingjong.

    1989-01-01

    To test the hypothesis that transiently open junctions around the endothelial cells undergoing turnover are the large pores through which macromolecules (with the size of albumin or larger) cross the endothelium in large arteries, experiments were performed on Sprague-Dawley, spontaneously hypertensive (SHR), and Wistar-Kyoto (WKY) rats under anesthesia. Endothelial macromolecular permeability was studied in both en face and cross-sectional preparations of the thoracic aorta by using fluorescence-labeled albumin and low density lipoprotein (LDL). Foci of macromolecular leakage in the aorta were visualized and mapped with fluorescence microscopy. Replicating endothelial cells were identified by {sup 3}H-thymidine autoradiography and hemotoxylin staining, and dead endothelia cells were detected by IgG immunocytochemistry. The occurrence of cell replication or death was correlated with macromolecular leakage in the aorta. Under the fluorescence microscope, endothelial macromolecular leakage was clearly identified as discrete spots or larger foci. Both the number density and the size of tracer leaky foci increased with time. A higher number density of leaky foci was found in branching areas. A high degree of correlation was found between macromolecular permeability and endothelial cell mitosis or death at the cellular level. A significantly higher density of tracer leaky foci was noted in SHR than in WKY, which was accompanied by greater frequencies of both endothelial cell mitosis and death. During cell turnover, aortic endothelial cells continuously undergo structural remodeling, including cell junctions. Poorly organized clefts associated with the remodeling provide the pathway through which macromolecules enter the arterial wall.

  3. Structure of graphene oxide membranes in solvents and solutions

    NASA Astrophysics Data System (ADS)

    Klechikov, Alexey; Yu, Junchun; Thomas, Diana; Sharifi, Tiva; Talyzin, Alexandr V.

    2015-09-01

    The change of distance between individual graphene oxide sheets due to swelling is the key parameter to explain and predict permeation of multilayered graphene oxide (GO) membranes by various solvents and solutions. In situ synchrotron X-ray diffraction study shows that swelling properties of GO membranes are distinctly different compared to precursor graphite oxide powder samples. Intercalation of liquid dioxolane, acetonitrile, acetone, and chloroform into the GO membrane structure occurs with maximum one monolayer insertion (Type I), in contrast with insertion of 2-3 layers of these solvents into the graphite oxide structure. However, the structure of GO membranes expands in liquid DMSO and DMF solvents similarly to precursor graphite oxide (Type II). It can be expected that Type II solvents will permeate GO membranes significantly faster compared to Type I solvents. The membranes are found to be stable in aqueous solutions of acidic and neutral salts, but dissolve slowly in some basic solutions of certain concentrations, e.g. in NaOH, NaHCO3 and LiF. Some larger organic molecules, alkylamines and alkylammonium cations are found to intercalate and expand the lattice of GO membranes significantly, e.g. up to ~35 Å in octadecylamine/methanol solution. Intercalation of solutes into the GO structure is one of the limiting factors for nano-filtration of certain molecules but it also allows modification of the inter-layer distance of GO membranes and tuning of their permeation properties. For example, GO membranes functionalized with alkylammonium cations are hydrophobized and they swell in non-polar solvents.The change of distance between individual graphene oxide sheets due to swelling is the key parameter to explain and predict permeation of multilayered graphene oxide (GO) membranes by various solvents and solutions. In situ synchrotron X-ray diffraction study shows that swelling properties of GO membranes are distinctly different compared to precursor graphite

  4. Development of solution techniques for nonlinear structural analysis

    NASA Technical Reports Server (NTRS)

    Vos, R. G.; Andrews, J. S.

    1974-01-01

    Nonlinear structural solution methods in the current research literature are classified according to order of the solution scheme, and it is shown that the analytical tools for these methods are uniformly derivable by perturbation techniques. A new perturbation formulation is developed for treating an arbitrary nonlinear material, in terms of a finite-difference generated stress-strain expansion. Nonlinear geometric effects are included in an explicit manner by appropriate definition of an applicable strain tensor. A new finite-element pilot computer program PANES (Program for Analysis of Nonlinear Equilibrium and Stability) is presented for treatment of problems involving material and geometric nonlinearities, as well as certain forms on nonconservative loading.

  5. Preparation of phenylboronate affinity rigid monolith with macromolecular porogen.

    PubMed

    Li, Xiang-Jie; Jia, Man; Zhao, Yong-Xin; Liu, Zhao-Sheng; Akber Aisa, Haji

    2016-03-18

    Boronate-affinity monolithic column was first prepared via polystyrene (PS) as porogen in this work. The monolithic polymer was synthetized using 4-vinylphenylboronic acid (4-VPBA) as functional monomer, ethylene glycol dimethacrylate (EDMA) as crosslinker monomer, and a mixture of PS solution in tetrahydrofuran, the linear macromolecular porogen, and toluene as porogen. Isoquercitrin (ISO) and hyperoside (HYP), isomer diol flavonoid glycosides, can be baseline separated on the poly(VPBA-co-EDMA) monolith. The effect of polymerization variables on the selectivity factor, e.g., the ratio of monomer to crosslinker (M/C), the amount of PS and the molecular weight of macromolecular porogen was investigated. The surface properties of the monolithic polymer were characterized by scanning electron microscopy and nitrogen adsorption. The best polymerization condition was the M/C ratio of 7:3, and the PS concentration of 40 mg/ml. The poly(VPBA-co-EDMA) polymer was also applied to extract cis-diol flavonoid glycosides from the crude extraction of cotton flower. After treated by poly(VPBA-co-EDMA) for solid phase extraction, high purity ISO and HYP (>99.96%) can be obtained with recovery of 83.7% and 78.6%, respectively. PMID:26896914

  6. Protein stabilization by macromolecular crowding through enthalpy rather than entropy.

    PubMed

    Senske, Michael; Törk, Lisa; Born, Benjamin; Havenith, Martina; Herrmann, Christian; Ebbinghaus, Simon

    2014-06-25

    The interior of the cell is a densely crowded environment in which protein stability is affected differently than in dilute solution. Macromolecular crowding is commonly understood in terms of an entropic volume exclusion effect based on hardcore repulsions among the macromolecules. We studied the thermal unfolding of ubiquitin in the presence of different cosolutes (glucose, dextran, poly(ethylene glycol), KCl, urea). Our results show that for a correct dissection of the cosolute-induced changes of the free energy into its enthalpic and entropic contributions, the temperature dependence of the heat capacity change needs to be explicitly taken into account. In contrast to the prediction by the excluded volume theory, we observed an enthalpic stabilization and an entropic destabilization for glucose, dextran, and poly(ethylene glycol). The enthalpic stabilization mechanism induced by the macromolecular crowder dextran was similar to the enthalpic stabilization mechanism of its monomeric building block glucose. In the case of poly(ethylene glycol), entropy is dominating over enthalpy leading to an overall destabilization. We propose a new model to classify cosolute effects in terms of their enthalpic contributions to protein stability.

  7. Preparation of phenylboronate affinity rigid monolith with macromolecular porogen.

    PubMed

    Li, Xiang-Jie; Jia, Man; Zhao, Yong-Xin; Liu, Zhao-Sheng; Akber Aisa, Haji

    2016-03-18

    Boronate-affinity monolithic column was first prepared via polystyrene (PS) as porogen in this work. The monolithic polymer was synthetized using 4-vinylphenylboronic acid (4-VPBA) as functional monomer, ethylene glycol dimethacrylate (EDMA) as crosslinker monomer, and a mixture of PS solution in tetrahydrofuran, the linear macromolecular porogen, and toluene as porogen. Isoquercitrin (ISO) and hyperoside (HYP), isomer diol flavonoid glycosides, can be baseline separated on the poly(VPBA-co-EDMA) monolith. The effect of polymerization variables on the selectivity factor, e.g., the ratio of monomer to crosslinker (M/C), the amount of PS and the molecular weight of macromolecular porogen was investigated. The surface properties of the monolithic polymer were characterized by scanning electron microscopy and nitrogen adsorption. The best polymerization condition was the M/C ratio of 7:3, and the PS concentration of 40 mg/ml. The poly(VPBA-co-EDMA) polymer was also applied to extract cis-diol flavonoid glycosides from the crude extraction of cotton flower. After treated by poly(VPBA-co-EDMA) for solid phase extraction, high purity ISO and HYP (>99.96%) can be obtained with recovery of 83.7% and 78.6%, respectively.

  8. Structure of graphene oxide membranes in solvents and solutions.

    PubMed

    Klechikov, Alexey; Yu, Junchun; Thomas, Diana; Sharifi, Tiva; Talyzin, Alexandr V

    2015-10-01

    The change of distance between individual graphene oxide sheets due to swelling is the key parameter to explain and predict permeation of multilayered graphene oxide (GO) membranes by various solvents and solutions. In situ synchrotron X-ray diffraction study shows that swelling properties of GO membranes are distinctly different compared to precursor graphite oxide powder samples. Intercalation of liquid dioxolane, acetonitrile, acetone, and chloroform into the GO membrane structure occurs with maximum one monolayer insertion (Type I), in contrast with insertion of 2-3 layers of these solvents into the graphite oxide structure. However, the structure of GO membranes expands in liquid DMSO and DMF solvents similarly to precursor graphite oxide (Type II). It can be expected that Type II solvents will permeate GO membranes significantly faster compared to Type I solvents. The membranes are found to be stable in aqueous solutions of acidic and neutral salts, but dissolve slowly in some basic solutions of certain concentrations, e.g. in NaOH, NaHCO3 and LiF. Some larger organic molecules, alkylamines and alkylammonium cations are found to intercalate and expand the lattice of GO membranes significantly, e.g. up to ∼35 Å in octadecylamine/methanol solution. Intercalation of solutes into the GO structure is one of the limiting factors for nano-filtration of certain molecules but it also allows modification of the inter-layer distance of GO membranes and tuning of their permeation properties. For example, GO membranes functionalized with alkylammonium cations are hydrophobized and they swell in non-polar solvents.

  9. Relationship between solution structure and phase behavior: a neutron scattering study of concentrated aqueous hexamethylenetetramine solutions.

    PubMed

    Burton, R C; Ferrari, E S; Davey, R J; Finney, J L; Bowron, D T

    2009-04-30

    The water-hexamethylenetetramine system displays features of significant interest in the context of phase equilibria in molecular materials. First, it is possible to crystallize two solid phases depending on temperature, both hexahydrate and anhydrous forms. Second, saturated aqueous solutions in equilibrium with these forms exhibit a negative dependence of solubility (retrograde) on temperature. In this contribution, neutron scattering experiments (with isotopic substitution) of concentrated aqueous hexamethylenetetramine solutions combined with empirical potential structure refinement (EPSR) were used to investigate the time-averaged atomistic details of this system. Through the derivation of radial distribution functions, quantitative details emerge of the solution coordination, its relationship to the nature of the solid phases, and of the underlying cause of the solubility behavior of this molecule.

  10. Travelling Wave Solutions in Multigroup Age-Structured Epidemic Models

    NASA Astrophysics Data System (ADS)

    Ducrot, Arnaut; Magal, Pierre; Ruan, Shigui

    2010-01-01

    Age-structured epidemic models have been used to describe either the age of individuals or the age of infection of certain diseases and to determine how these characteristics affect the outcomes and consequences of epidemiological processes. Most results on age-structured epidemic models focus on the existence, uniqueness, and convergence to disease equilibria of solutions. In this paper we investigate the existence of travelling wave solutions in a deterministic age-structured model describing the circulation of a disease within a population of multigroups. Individuals of each group are able to move with a random walk which is modelled by the classical Fickian diffusion and are classified into two subclasses, susceptible and infective. A susceptible individual in a given group can be crisscross infected by direct contact with infective individuals of possibly any group. This process of transmission can depend upon the age of the disease of infected individuals. The goal of this paper is to provide sufficient conditions that ensure the existence of travelling wave solutions for the age-structured epidemic model. The case of two population groups is numerically investigated which applies to the crisscross transmission of feline immunodeficiency virus (FIV) and some sexual transmission diseases.

  11. Poly(isophthalic acid)(ethylene oxide) as a Macromolecular Modulator for Metal-Organic Polyhedra.

    PubMed

    Chen, Teng-Hao; Wang, Le; Trueblood, Jonathan V; Grassian, Vicki H; Cohen, Seth M

    2016-08-01

    A new strategy was developed by using a polymer ligand, poly(isophthalic acid)(ethylene oxide), to modulate the growth of metal-organic polyhedra (MOP) crystals. This macromolecular modulator can effectively control the crystal habit of several different Cu24L24 (L = isophthalic acid derivatives) MOPs. The polymer also directed the formation of MOP structures under reaction conditions that only produce metal-organic frameworks in the absence of modulator. Moreover, the polymer also enabled the deposition of MOP crystals on glass surfaces. This macromolecular modulator strategy provides an innovative approach to control the morphology and assembly of MOP particles. PMID:27400759

  12. Steady state solutions to dynamically loaded periodic structures

    NASA Technical Reports Server (NTRS)

    Kalinowski, A. J.

    1980-01-01

    The general problem of solving for the steady state (time domain) dynamic response (i.e., NASTRAN rigid format-8) of a general elastic periodic structure subject to a phase difference loading of the type encountered in traveling wave propagation problems was studied. Two types of structural configurations were considered; in the first type, the structure has a repeating pattern over a span that is long enough to be considered, for all practical purposes, as infinite; in the second type, the structure has structural rotational symmetry in the circumferential direction. The theory and a corresponding set of DMAP instructions which permits the NASTRAN user to automatically alter the rigid format-8 sequence to solve the intended class of problems are presented. Final results are recovered as with any ordinary rigid format-8 solution, except that the results are only printed for the typical periodic segment of the structure. A simple demonstration problem having a known exact solution is used to illustrate the implementation of the procedure.

  13. Effect of Ternary Solutes on the Evolution of Structure and Gel Formation in Amphiphilic Copolymer Solutions

    NASA Astrophysics Data System (ADS)

    Meznarich, Norman Anthony Kang

    Aqueous solutions of polyoxyethylene-polyoxypropylene-polyoxyethylene (PEO-PPO-PEO) amphiphilic triblock copolymers (commercially known as Pluronic surfactants) undergo reversible and temperature-dependent micellization and arrangement into cubic ordered lattices known as "micelle gels". The macroscopic behavior of the ordering is a transition from a liquid to a gel. While the phase behavior and gel structure of pure Pluronic surfactant solutions have been well studied, less is known about the effects of added ternary solutes. In this dissertation, a comprehensive investigation into the effects of the added pharmaceutical methylparaben on solutions of F127 ranging from 10 to 30 wt% was conducted in order to better understand the behavior of F127 in multicomponent pharmaceutical formulations. The viscoelastic properties of F127 gel formation were studied using rheometry, where heating rates of 0.1, 1, and 10 degrees C/min were also used to probe the kinetics of the gel transition. In solutions containing methylparaben, F127 gelation occurred at up to 15 degrees C lower temperatures and was accelerated by a factor of three to four. Small angle x-ray scattering (SAXS) was used to characterize the structure of the ordered domains, and how they were affected by the presence of dissolved pharmaceuticals. It was found that ordered domain formation changed from heterogeneous nucleation and growth to possible homogeneous nucleation and growth. A roughly 2% reduction in the cubic lattice parameter was also observed for solutions containing methylparaben. Differential scanning calorimetry (DSC) experiments were performed on a series of different Pluronic surfactants in order to characterize the micellization behavior as a function of PPO center block length and PEO/PPO ratio. Added methylparaben suppressed the micellization endotherm, the degree of suppression depending linearly on the amount of added methylparaben, as well as the length of the PPO center block and PEO

  14. Light scattering measurements supporting helical structures for chromatin in solution.

    PubMed

    Campbell, A M; Cotter, R I; Pardon, J F

    1978-05-01

    Laser light scattering measurements have been made on a series of polynucleosomes containing from 50 to 150 nucleosomes. Radii of gyration have been determined as a function of polynucleosome length for different ionic strength solutions. The results suggest that at low ionic strength the chromatin adopts a loosely helical structure rather than a random coil. The helix becomes more regular on increasing the ionic strength, the dimension resembling those proposed by Finch and Klug for their solenoid model.

  15. Understanding the defect structure of solution grown zinc oxide

    SciTech Connect

    Liew, Laura-Lynn; Sankar, Gopinathan; Handoko, Albertus D.; Goh, Gregory K.L.; Kohara, Shinji

    2012-05-15

    Zinc oxide (ZnO) is a wide bandgap semiconducting oxide with many potential applications in various optoelectronic devices such as light emitting diodes (LEDs) and field effect transistors (FETs). Much effort has been made to understand the ZnO structure and its defects. However, one major issue in determining whether it is Zn or O deficiency that provides ZnO its unique properties remains. X-ray absorption spectroscopy (XAS) is an ideal, atom specific characterization technique that is able to probe defect structure in many materials, including ZnO. In this paper, comparative studies of bulk and aqueous solution grown ({<=}90 Degree-Sign C) ZnO powders using XAS and x-ray pair distribution function (XPDF) techniques are described. The XAS Zn-Zn correlation and XPDF results undoubtedly point out that the solution grown ZnO contains Zn deficiency, rather than the O deficiency that were commonly reported. This understanding of ZnO short range order and structure will be invaluable for further development of solid state lighting and other optoelectronic device applications. - Graphical abstract: Highlights: Black-Right-Pointing-Pointer ZnO powders have been synthesized through an aqueous solution method. Black-Right-Pointing-Pointer Defect structure studied using XAS and XPDF. Black-Right-Pointing-Pointer Zn-Zn correlations are less in the ZnO powders synthesized in solution than bulk. Black-Right-Pointing-Pointer Zn vacancies are present in the powders synthesized. Black-Right-Pointing-Pointer EXAFS and XPDF, when used complementary, are useful characterization techniques.

  16. Solution structure of the Legionella pneumophila Mip-rapamycin complex

    PubMed Central

    Ceymann, Andreas; Horstmann, Martin; Ehses, Philipp; Schweimer, Kristian; Paschke, Anne-Katrin; Steinert, Michael; Faber, Cornelius

    2008-01-01

    Background Legionella pneumphila is the causative agent of Legionnaires' disease. A major virulence factor of the pathogen is the homodimeric surface protein Mip. It shows peptidyl-prolyl cis/trans isomerase activty and is a receptor of FK506 and rapamycin, which both inhibit its enzymatic function. Insight into the binding process may be used for the design of novel Mip inhibitors as potential drugs against Legionnaires' disease. Results We have solved the solution structure of free Mip77–213 and the Mip77–213-rapamycin complex by NMR spectroscopy. Mip77–213 showed the typical FKBP-fold and only minor rearrangements upon binding of rapamycin. Apart from the configuration of a flexible hairpin loop, which is partly stabilized upon binding, the solution structure confirms the crystal structure. Comparisons to the structures of free FKBP12 and the FKBP12-rapamycin complex suggested an identical binding mode for both proteins. Conclusion The structural similarity of the Mip-rapamycin and FKBP12-rapamycin complexes suggests that FKBP12 ligands may be promising starting points for the design of novel Mip inhibitors. The search for a novel drug against Legionnaires' disease may therefore benefit from the large variety of known FKBP12 inhibitors. PMID:18366641

  17. Solution structure of the HIV gp120 C5 domain.

    PubMed

    Guilhaudis, Laure; Jacobs, Amy; Caffrey, Michael

    2002-10-01

    In HIV the viral envelope protein is processed by a host cell protease to form gp120 and gp41. The C1 and C5 domains of gp120 are thought to directly interact with gp41 but are largely missing from the available X-ray structure. Biophysical studies of the HIV gp120 C5 domain (residues 489-511 of HIV-1 strain HXB2), which corresponds to the carboxy terminal region of gp120, have been undertaken. CD studies of the C5 domain suggest that it is unstructured in aqueous solutions but partially helical in trifluoroethanol/aqueous and hexafluoroisopropanol/aqueous buffers. The solution structure of the C5 peptide in 40% trifluoroethanol/aqueous buffer was determined by NMR spectroscopy. The resulting structure is a turn helix structural motif, consistent with the CD results. Fluorescence titration experiments suggest that HIV C5 forms a 1 : 1 complex with the HIV gp41 ectodomain in the presence of cosolvent with an apparent Kd of approximately 1.0 micro m. The absence of complex formation in the absence of cosolvent indicates that formation of the turn-helix structural motif of C5 is necessary for complex formation. Examination of the C5 structure provides insight into the interaction between gp120 and gp41 and provides a possible target site for future drug therapies designed to disrupt the gp120/gp41 complex. In addition, the C5 structure lends insight into the site of HIV envelope protein maturation by the host enzymes furin and PC7, which provides other possible targets for drug therapies.

  18. Brownian Dynamics Simulation of Protein Solutions: Structural and Dynamical Properties

    SciTech Connect

    Mereghetti, Paolo; Gabdoulline, Razif; Wade, Rebecca C.

    2010-12-01

    The study of solutions of biomacromolecules provides an important basis for understanding the behavior of many fundamental cellular processes, such as protein folding, self-assembly, biochemical reactions, and signal transduction. Here, we describe a Brownian dynamics simulation procedure and its validation for the study of the dynamic and structural properties of protein solutions. In the model used, the proteins are treated as atomically detailed rigid bodies moving in a continuum solvent. The protein-protein interaction forces are described by the sum of electrostatic interaction, electrostatic desolvation, nonpolar desolvation, and soft-core repulsion terms. The linearized Poisson-Boltzmann equation is solved to compute electrostatic terms. Simulations of homogeneous solutions of three different proteins with varying concentrations, pH, and ionic strength were performed. The results were compared to experimental data and theoretical values in terms of long-time self-diffusion coefficients, second virial coefficients, and structure factors. The results agree with the experimental trends and, in many cases, experimental values are reproduced quantitatively. There are no parameters specific to certain protein types in the interaction model, and hence the model should be applicable to the simulation of the behavior of mixtures of macromolecules in cell-like crowded environments.

  19. Use of Capillaries for Macromolecular Crystallization in a Cryogenic Dewar

    NASA Technical Reports Server (NTRS)

    Ciszak, Ewa; Hammons, Aaron S.; Hong, Young Soo

    2002-01-01

    The enhanced gaseous nitrogen (EGN) dewar is a cryogenic dry shipper with a sealed cylinder inserted inside along with a temperature monitoring device, and is intended for macromolecular crystallization experiments on the International Space Station. Within the dewar, each crystallization experiment is contained as a solution within a plastic capillary tube. The standard procedure for loading samples in these tubes has involved rapid freezing of the precipitant and biomolecular solution, e.g., protein, directly in liquid nitrogen; this method, however, often resulted in uncontrolled formation of air voids, These air pockets, or bubbles, can lead to irreproducible crystallization results. A novel protocol has been developed to prevent formation of bubbles, and this has been tested in the laboratory as well as aboard the International Space Station during a 42-day long mission of July/August 2001. The gain or loss of mass from solutions within the plastic capillaries revealed that mass transport occurred among separated tubes, and that this mass transport was dependent upon the hygroscopic character of the solution contained in any given tube. The surface area of the plastic capillary tube also related to the observed mass transport. Furthermore, the decreased mass of solutions of-protein correlated to observed formation of protein crystals.

  20. Solution structure of murine macrophage inflammatory protein-2.

    PubMed

    Shao, W; Jerva, L F; West, J; Lolis, E; Schweitzer, B I

    1998-06-01

    The solution structure of murine macrophage inflammatory protein-2 (MIP-2), a heparin-binding chemokine that is secreted in response to inflammatory stimuli, has been determined using two-dimensional homonuclear and heteronuclear NMR spectroscopy. Structure calculations were carried out by means of torsion-angle molecular dynamics using the program X-PLOR. The structure is based on a total of 2390 experimental restraints, comprising 2246 NOE-derived distance restraints, 44 distance restraints for 22 hydrogen bonds, and 100 torsion angle restraints. The structure is well-defined, with the backbone (N, Calpha, C) and heavy atom atomic rms distribution about the mean coordinates for residues 9-69 of the dimer being 0.57 +/- 0.16 A and 0.96 +/- 0.12 A, respectively. The N- and C-terminal residues (1-8 and 70-73, respectively) are disordered. The overall structure of the MIP-2 dimer is similar to that reported previously for the NMR structures of MGSA and IL-8 and consists of a six-stranded antiparallel beta-sheet (residue 25-29, 39-44, and 48-52) packed against two C-terminal antiparallel alpha-helices. A best fit superposition of the NMR structure of MIP-2 on the structures of MGSA, NAP-2, and the NMR and X-ray structures of IL-8 are 1.11, 1.02, 1.27, and 1.19 A, respectively, for the monomers, and 1.28, 1.10, 1.55, and 1.36 A, respectively, for the dimers (IL-8 residues 7-14 and 16-67, NAP-2 residues 25-84). At the tertiary level, the main differences between the MIP-2 solution structure and the IL-8, MGSA, and NAP-2 structures involve the N-terminal loop between residues 9-23 and the loops formed by residues 30-38 and residues 53-58. At the quaternary level, the difference between MIP-2 and IL-8, MGSA, or NAP-2 results from differing interhelical angles and separations.

  1. Solution structure of murine macrophage inflammatory protein-2.

    PubMed

    Shao, W; Jerva, L F; West, J; Lolis, E; Schweitzer, B I

    1998-06-01

    The solution structure of murine macrophage inflammatory protein-2 (MIP-2), a heparin-binding chemokine that is secreted in response to inflammatory stimuli, has been determined using two-dimensional homonuclear and heteronuclear NMR spectroscopy. Structure calculations were carried out by means of torsion-angle molecular dynamics using the program X-PLOR. The structure is based on a total of 2390 experimental restraints, comprising 2246 NOE-derived distance restraints, 44 distance restraints for 22 hydrogen bonds, and 100 torsion angle restraints. The structure is well-defined, with the backbone (N, Calpha, C) and heavy atom atomic rms distribution about the mean coordinates for residues 9-69 of the dimer being 0.57 +/- 0.16 A and 0.96 +/- 0.12 A, respectively. The N- and C-terminal residues (1-8 and 70-73, respectively) are disordered. The overall structure of the MIP-2 dimer is similar to that reported previously for the NMR structures of MGSA and IL-8 and consists of a six-stranded antiparallel beta-sheet (residue 25-29, 39-44, and 48-52) packed against two C-terminal antiparallel alpha-helices. A best fit superposition of the NMR structure of MIP-2 on the structures of MGSA, NAP-2, and the NMR and X-ray structures of IL-8 are 1.11, 1.02, 1.27, and 1.19 A, respectively, for the monomers, and 1.28, 1.10, 1.55, and 1.36 A, respectively, for the dimers (IL-8 residues 7-14 and 16-67, NAP-2 residues 25-84). At the tertiary level, the main differences between the MIP-2 solution structure and the IL-8, MGSA, and NAP-2 structures involve the N-terminal loop between residues 9-23 and the loops formed by residues 30-38 and residues 53-58. At the quaternary level, the difference between MIP-2 and IL-8, MGSA, or NAP-2 results from differing interhelical angles and separations. PMID:9622482

  2. Conformational equilibria of alkanes in aqueous solution: relationship to water structure near hydrophobic solutes.

    PubMed Central

    Ashbaugh, H S; Garde, S; Hummer, G; Kaler, E W; Paulaitis, M E

    1999-01-01

    Conformational free energies of butane, pentane, and hexane in water are calculated from molecular simulations with explicit waters and from a simple molecular theory in which the local hydration structure is estimated based on a proximity approximation. This proximity approximation uses only the two nearest carbon atoms on the alkane to predict the local water density at a given point in space. Conformational free energies of hydration are subsequently calculated using a free energy perturbation method. Quantitative agreement is found between the free energies obtained from simulations and theory. Moreover, free energy calculations using this proximity approximation are approximately four orders of magnitude faster than those based on explicit water simulations. Our results demonstrate the accuracy and utility of the proximity approximation for predicting water structure as the basis for a quantitative description of n-alkane conformational equilibria in water. In addition, the proximity approximation provides a molecular foundation for extending predictions of water structure and hydration thermodynamic properties of simple hydrophobic solutes to larger clusters or assemblies of hydrophobic solutes. PMID:10423414

  3. The interplay of intrinsic disorder and macromolecular crowding on α-synuclein fibril formation

    NASA Astrophysics Data System (ADS)

    Shirai, Nobu C.; Kikuchi, Macoto

    2016-02-01

    α-synuclein (α-syn) is an intrinsically disordered protein which is considered to be one of the causes of Parkinson's disease. This protein forms amyloid fibrils when in a highly concentrated solution. The fibril formation of α-syn is induced not only by increases in α-syn concentration but also by macromolecular crowding. In order to investigate the coupled effect of the intrinsic disorder of α-syn and macromolecular crowding, we construct a lattice gas model of α-syn in contact with a crowding agent reservoir based on statistical mechanics. The main assumption is that α-syn can be expressed as coarse-grained particles with internal states coupled with effective volume; and disordered states are modeled by larger particles with larger internal entropy than other states. Thanks to the simplicity of the model, we can exactly calculate the number of conformations of crowding agents, and this enables us to prove that the original grand canonical ensemble with a crowding agent reservoir is mathematically equivalent to a canonical ensemble without crowding agents. In this expression, the effect of macromolecular crowding is absorbed in the internal entropy of disordered states; it is clearly shown that the crowding effect reduces the internal entropy. Based on Monte Carlo simulation, we provide scenarios of crowding-induced fibril formation. We also discuss the recent controversy over the existence of helically folded tetramers of α-syn, and suggest that macromolecular crowding is the key to resolving the controversy.

  4. Macromolecular Crowding Modulates Folding Mechanism of α/β Protein Apoflavodoxin

    NASA Astrophysics Data System (ADS)

    Homouz, D.; Stagg, L.; Wittungstafshede, P.; Cheung, M.

    2009-01-01

    Protein dynamics in cells may be different from that in dilute solutions in vitro since the environment in cells is highly concentrated with other macromolecules. This volume exclusion due to macromolecular crowding is predicted to affect both equilibrium and kinetic processes involving protein conformational changes. To quantify macromolecular crowding effects on protein folding mechanisms, here we have investigated the folding energy landscape of an alpha/beta protein, apoflavodoxin, in the presence of inert macromolecular crowding agents using in silico and in vitro approaches. By coarse-grained molecular simulations and topology-based potential interactions, we probed the effects of increased volume fraction of crowding agents (phi_c) as well as of crowding agent geometry (sphere or spherocylinder) at high phi_c. Parallel kinetic folding experiments with purified Desulfovibro desulfuricans apoflavodoxin in vitro were performed in the presence of Ficoll (sphere) and Dextran (spherocylinder) synthetic crowding agents. In conclusion, we have identified in silico crowding conditions that best enhance protein stability and discovered that upon manipulation of the crowding conditions, folding routes experiencing topological frustrations can be either enhanced or relieved. The test-tube experiments confirmed that apoflavodoxin's time-resolved folding path is modulated by crowding agent geometry. We propose that macromolecular crowding effects may be a tool for manipulation of protein folding and function in living cells.

  5. A primer in macromolecular linguistics.

    PubMed

    Searls, David B

    2013-03-01

    Polymeric macromolecules, when viewed abstractly as strings of symbols, can be treated in terms of formal language theory, providing a mathematical foundation for characterizing such strings both as collections and in terms of their individual structures. In addition this approach offers a framework for analysis of macromolecules by tools and conventions widely used in computational linguistics. This article introduces the ways that linguistics can be and has been applied to molecular biology, covering the relevant formal language theory at a relatively nontechnical level. Analogies between macromolecules and human natural language are used to provide intuitive insights into the relevance of grammars, parsing, and analysis of language complexity to biology.

  6. A primer in macromolecular linguistics.

    PubMed

    Searls, David B

    2013-03-01

    Polymeric macromolecules, when viewed abstractly as strings of symbols, can be treated in terms of formal language theory, providing a mathematical foundation for characterizing such strings both as collections and in terms of their individual structures. In addition this approach offers a framework for analysis of macromolecules by tools and conventions widely used in computational linguistics. This article introduces the ways that linguistics can be and has been applied to molecular biology, covering the relevant formal language theory at a relatively nontechnical level. Analogies between macromolecules and human natural language are used to provide intuitive insights into the relevance of grammars, parsing, and analysis of language complexity to biology. PMID:23034580

  7. Spatial structures in microtubular solutions requiring a sustained energy source

    NASA Astrophysics Data System (ADS)

    Tabony, J.; Job, D.

    1990-08-01

    MICROTUBULES are believed to be the principal organizers of the cell interior1. Cells respond to a variety of stimuli by modifying the spatial distribution of the microtubules. These effects are central to cell division and morphogenesis2, and embryo development3. During embryo development, macroscopic patterns are frequently observed3. Here we report that microtubular solutions spontaneously form alternating white and dark stripes about 1 mm wide and 1 cm long. Small-angle neutron scattering measurements show that in each segment the microtubules are aligned obliquely to the direction of the stripe, and that the white and dark stripes differ in having mutually orthogonal orientations. The formation of these structures requires an initial reservoir of organic phosphate. Phosphorus NMR measurements show that the process is accompanied by the energy-liberating conversion of organic to inorganic phosphate. These observations, together with similarities to the dissipative spatial structure formed by the Belousov-Zhabotinski reaction4-6, provide strong evidence that the observed structures are energy-dissipative in nature. Dissipative structures are thought to be critical to the appearence of complex living organisms7,8. Our results strongly suggest that microtubules are capable of forming such structures. Microtubular dissipative structures may occur during mitosis and embryo morphogenesis.

  8. The structure of aqueous sodium hydroxide solutions: a combined solution x-ray diffraction and simulation study.

    PubMed

    Megyes, Tünde; Bálint, Szabolcs; Grósz, Tamás; Radnai, Tamás; Bakó, Imre; Sipos, Pál

    2008-01-28

    To determine the structure of aqueous sodium hydroxide solutions, results obtained from x-ray diffraction and computer simulation (molecular dynamics and Car-Parrinello) have been compared. The capabilities and limitations of the methods in describing the solution structure are discussed. For the solutions studied, diffraction methods were found to perform very well in describing the hydration spheres of the sodium ion and yield structural information on the anion's hydration structure. Classical molecular dynamics simulations were not able to correctly describe the bulk structure of these solutions. However, Car-Parrinello simulation proved to be a suitable tool in the detailed interpretation of the hydration sphere of ions and bulk structure of solutions. The results of Car-Parrinello simulations were compared with the findings of diffraction experiments.

  9. On the atomic structure of cocaine in solution.

    PubMed

    Johnston, Andrew J; Busch, Sebastian; Pardo, Luis Carlos; Callear, Samantha K; Biggin, Philip C; McLain, Sylvia E

    2016-01-14

    Cocaine is an amphiphilic drug which has the ability to cross the blood-brain barrier (BBB). Here, a combination of neutron diffraction and computation has been used to investigate the atomic scale structure of cocaine in aqueous solutions. Both the observed conformation and hydration of cocaine appear to contribute to its ability to cross hydrophobic layers afforded by the BBB, as the average conformation yields a structure which might allow cocaine to shield its hydrophilic regions from a lipophilic environment. Specifically, the carbonyl oxygens and amine group on cocaine, on average, form ∼5 bonds with the water molecules in the surrounding solvent, and the top 30% of water molecules within 4 Å of cocaine are localized in the cavity formed by an internal hydrogen bond within the cocaine molecule. This water mediated internal hydrogen bonding suggests a mechanism of interaction between cocaine and the BBB that negates the need for deprotonation prior to interaction with the lipophilic portions of this barrier. This finding also has important implications for understanding how neurologically active molecules are able to interact with both the blood stream and BBB and emphasizes the use of structural measurements in solution in order to understand important biological function.

  10. Crystallization of macromolecular complexes: combinatorial complex crystallization

    NASA Astrophysics Data System (ADS)

    Stura, Enrico A.; Graille, Marc; Charbonnier, Jean-Baptiste

    2001-11-01

    The usefulness of antibody complexation, as a way of increasing the chances of crystallization needs to be re-evaluated after many antibody complexes have been crystallized and their structure determined. It is somewhat striking that among these, only a small number is a complex with a large protein antigen. The problem is that the effort of raising, cleaving and purifying an Fab is rewarded only by an extra chance of getting crystals; depending on the relative likelihood of crystallization of the complexed and uncomplexed protein. The example of the complex between HIV gp120, CD4 and an Fab fragment from a neutralizing antibody suggests that further complexation of an antigen-antibody complex with a third protein could, by increasing the number of possible combinations, improve the likelihood of crystallization. We propose the use of Ig-binding proteins as a way of extending the method from HIV gp120 to all proteins for which there are monoclonal antibodies. We discuss this technique, combinatorial complex crystallization (CCC), as part of a multi-component system for the enhancement of crystallization of macromolecular complexes. The method makes use of single Ig-binding domains from Staphylococcus aureus protein A (SpA), Peptostreptococcus magnus protein L (PpL) and the streptococcal protein G (SpG). The generality of the method depends on the ability of these domains to interact with a large repertoire of antibodies without affecting antigen binding. There is strong evidence to suggest that these Ig-binding domains bind outside the antigen-combining site of the antibody without perturbing antigen binding. It is clear from the crystal structure of the single SpG domain complexed with an Fab that the interaction involves mainly the immunoglobulin CH1 domain, a region not involved in antigen recognition. We have recently determined the structure of the complex between a human Fab and the domain D from SpA and found that steric hindrance is unlikely even for large

  11. Macromolecular Crystallization with Microfluidic Free-Interface Diffusion

    SciTech Connect

    Segelke, B

    2005-02-24

    Fluidigm released the Topaz 1.96 and 4.96 crystallization chips in the fall of 2004. Topaz 1.96 and 4.96 are the latest evolution of Fluidigm's microfluidics crystallization technologies that enable ultra low volume rapid screening for macromolecular crystallization. Topaz 1.96 and 4.96 are similar to each other but represent a major redesign of the Topaz system and have of substantially improved ease of automation and ease of use, improved efficiency and even further reduced amount of material needed. With the release of the new Topaz system, Fluidigm continues to set the standard in low volume crystallization screening which is having an increasing impact in the field of structural genomics, and structural biology more generally. In to the future we are likely to see further optimization and increased utility of the Topaz crystallization system, but we are also likely to see further innovation and the emergence of competing technologies.

  12. Single-particle cryo-electron microscopy of macromolecular complexes.

    PubMed

    Skiniotis, Georgios; Southworth, Daniel R

    2016-02-01

    Recent technological breakthroughs in image acquisition have enabled single-particle cryo-electron microscopy (cryo-EM) to achieve near-atomic resolution structural information for biological complexes. The improvements in image quality coupled with powerful computational methods for sorting distinct particle populations now also allow the determination of compositional and conformational ensembles, thereby providing key insights into macromolecular function. However, the inherent instability and dynamic nature of biological assemblies remain a tremendous challenge that often requires tailored approaches for successful implementation of the methodology. Here, we briefly describe the fundamentals of single-particle cryo-EM with an emphasis on covering the breadth of techniques and approaches, including low- and high-resolution methods, aiming to illustrate specific steps that are crucial for obtaining structural information by this method.

  13. Solution to certain problems in the failure of composite structures

    NASA Astrophysics Data System (ADS)

    Goodsell, Johnathan

    The present work contains the solution of two problems in composite structures. In the first, an approximate elasticity solution for prediction of the displacement, stress and strain fields within the m-layer, symmetric and balanced angle-ply composite laminate of finite-width subjected anticlastic bending deformation is developed. The solution is shown to recover classical laminated plate theory predictions at interior regions of the laminate and thereby illustrates the boundary layer character of this interlaminar phenomenon. The results exhibit the anticipated response in congruence with the solutions for uniform axial extension and uniform temperature change, where divergence of the interlaminar shearing stress is seen to occur at the intersection of the free-edge and planes between lamina of +theta and -theta orientation. The analytical results show excellent agreement with the finite-element predictions for the same boundary-value problem and thereby provide an efficient and compact solution available for parametric studies of the influence of geometry and material properties. The solution is combined with previously developed solutions for uniform axial extension and uniform temperature change of the identical laminate and the combined solution is exercised to compare the relative magnitudes of free-edge phenomenon arising from the different loading conditions, to study very thick laminates and laminates where the laminate width is less than the laminate thickness. Significantly, it was demonstrated that the solution is valid for arbitrary stacking sequence and the solution was exercised to examine antisymmetric and non-symmetric laminates. Finally, the solution was exercised to determine the dimensions of the boundary layer for very large numbers of layers. It was found that the dimension of the boundary layer width in bending is approximately twice that in uniform axial extension and uniform temperature change. In the second, the intrinsic flaw concept is

  14. Solution structure of the human signaling protein RACK1

    PubMed Central

    2010-01-01

    Background The adaptor protein RACK1 (receptor of activated kinase 1) was originally identified as an anchoring protein for protein kinase C. RACK1 is a 36 kDa protein, and is composed of seven WD repeats which mediate its protein-protein interactions. RACK1 is ubiquitously expressed and has been implicated in diverse cellular processes involving: protein translation regulation, neuropathological processes, cellular stress, and tissue development. Results In this study we performed a biophysical analysis of human RACK1 with the aim of obtaining low resolution structural information. Small angle X-ray scattering (SAXS) experiments demonstrated that human RACK1 is globular and monomeric in solution and its low resolution structure is strikingly similar to that of an homology model previously calculated by us and to the crystallographic structure of RACK1 isoform A from Arabidopsis thaliana. Both sedimentation velocity and sedimentation equilibrium analytical ultracentrifugation techniques showed that RACK1 is predominantly a monomer of around 37 kDa in solution, but also presents small amounts of oligomeric species. Moreover, hydrodynamic data suggested that RACK1 has a slightly asymmetric shape. The interaction of RACK1 and Ki-1/57 was tested by sedimentation equilibrium. The results suggested that the association between RACK1 and Ki-1/57(122-413) follows a stoichiometry of 1:1. The binding constant (KB) observed for RACK1-Ki-1/57(122-413) interaction was of around (1.5 ± 0.2) × 106 M-1 and resulted in a dissociation constant (KD) of (0.7 ± 0.1) × 10-6 M. Moreover, the fluorescence data also suggests that the interaction may occur in a cooperative fashion. Conclusion Our SAXS and analytical ultracentrifugation experiments indicated that RACK1 is predominantly a monomer in solution. RACK1 and Ki-1/57(122-413) interact strongly under the tested conditions. PMID:20529362

  15. New solution method for steady-state canopy structural loads

    SciTech Connect

    Sundberg, W.D.

    1986-08-01

    A new computer code has been written to perform structural analysis canopies. Although an existing code, CANO, has been available, the new code has better convergence reliability, is more understandably written, and is easier to use. The equations have been reformulated for the new solution method. The new code assumes a symmetric canopy, a steady-state condition, and no strength in the vertical direction. It computes the inflated shape, loads in the horizontal members, radial members, vent lines, and suspension lines, and total drag. Constructed geometry, material properties, dynamic pressure, and pressure distribution are required as input.

  16. The Promise of Macromolecular Crystallization in Micro-fluidic Chips

    NASA Technical Reports Server (NTRS)

    vanderWoerd, Mark; Ferree, Darren; Pusey, Marc

    2003-01-01

    Micro-fluidics, or lab on a chip technology, is proving to be a powerful, rapid, and efficient approach to a wide variety of bio-analytical and microscale bio-preparative needs. The low materials consumption, combined with the potential for packing a large number of experiments in a few cubic centimeters, makes it an attractive technique for both initial screening and subsequent optimization of macromolecular crystallization conditions. Screening operations, which require equilibrating macromolecule solution with a standard set of premixed solutions, are relatively straightforward and have been successfully demonstrated in a micro-fluidics platform. More complex optimization methods, where crystallization solutions are independently formulated from a range of stock solutions, are considerably more complex and have yet to be demonstrated. To be competitive with either approach, a micro-fluidics system must offer ease of operation, be able to maintain a sealed environment over several weeks to months, and give ready access for the observation of crystals as they are grown.

  17. Interfacial structures of acidic and basic aqueous solutions

    SciTech Connect

    Tian, C.; Ji, N.; Waychunas, G.; Shen, Y.R.

    2008-10-20

    Phase-sensitive sum-frequency vibrational spectroscopy was used to study water/vapor interfaces of HCl, HI, and NaOH solutions. The measured imaginary part of the surface spectral responses provided direct characterization of OH stretch vibrations and information about net polar orientations of water species contributing to different regions of the spectrum. We found clear evidence that hydronium ions prefer to emerge at interfaces. Their OH stretches contribute to the 'ice-like' band in the spectrum. Their charges create a positive surface field that tends to reorient water molecules more loosely bonded to the topmost water layer with oxygen toward the interface, and thus enhances significantly the 'liquid-like' band in the spectrum. Iodine ions in solution also like to appear at the interface and alter the positive surface field by forming a narrow double-charge layer with hydronium ions. In NaOH solution, the observed weak change of the 'liquid-like' band and disappearance of the 'ice-like' band in the spectrum indicates that OH{sup -} ions must also have excess at the interface. How they are incorporated in the interfacial water structure is however not clear.

  18. Structure and dynamics of of solution polymerized polyureas

    NASA Astrophysics Data System (ADS)

    Choi, Taeyi; Jeong, Youmi; Runt, James

    2011-03-01

    Polyureas consisting of alternating soft and hard (urea containing) segments exhibit physical properties that are closely related to their microphase separated structure, which consist of rigid (high Tg and sometimes crystalline) hard domains embedded in a matrix dominated by flexible polyether segments. Polyurea properties can be controlled over a rather broad range by varying the chemical structures, molecular weight of the components, and reaction stoichiometry. In the present study, we focus primarily on linear polyureas synthesized using methylene diphenyl diisocyanate and polytetramethylene oxide-di-p-aminobenzoate using a solution polymerization method. Soft segment (diamine) molecular weights were varied from 460 to 860 to 1200 g/mol and characterize their morphology, hydrogen bonding, mechanical behavior and dielectric properties upon varying molecular weight of diamines. This presentation will focus on our latest findings, particularly details of the microphase separated morphology and molecular dynamics as measured using dielectric relaxation spectroscopy This work is supported by Office of Naval Research.

  19. Advances in macromolecular data storage

    NASA Astrophysics Data System (ADS)

    Mansuripur, Masud

    2014-09-01

    user-data in a scheme that benefits from the massive parallelism of thousands of Read/Write stations within the same three-dimensionally micro-structured device.

  20. Solution structures of rat amylin peptide: simulation, theory, and experiment.

    PubMed

    Reddy, Allam S; Wang, Lu; Lin, Yu-Shan; Ling, Yun; Chopra, Manan; Zanni, Martin T; Skinner, James L; De Pablo, Juan J

    2010-02-01

    Amyloid deposits of amylin in the pancreas are an important characteristic feature found in patients with Type-2 diabetes. The aggregate has been considered important in the disease pathology and has been studied extensively. However, the secondary structures of the individual peptide have not been clearly identified. In this work, we present detailed solution structures of rat amylin using a combination of Monte Carlo and molecular dynamics simulations. A new Monte Carlo method is presented to determine the free energy of distinct biomolecular conformations. Both folded and random-coil conformations of rat amylin are observed in water and their relative stability is examined in detail. The former contains an alpha-helical segment comprised of residues 7-17. We find that at room temperature the folded structure is more stable, whereas at higher temperatures the random-coil structure predominates. From the configurations and weights we calculate the alpha-carbon NMR chemical shifts, with results that are in reasonable agreement with experiments of others. We also calculate the infrared spectrum in the amide I stretch regime, and the results are in fair agreement with the experimental line shape presented herein.

  1. Uniqueness of the macromolecular crystallographic phase problem.

    PubMed

    Millane, Rick P; Arnal, Romain D

    2015-11-01

    Uniqueness of the phase problem in macromolecular crystallography, and its relationship to the case of single particle imaging, is considered. The crystallographic problem is characterized by a constraint ratio that depends only on the size and symmetry of the molecule and the unit cell. The results are used to evaluate the effect of various real-space constraints. The case of an unknown molecular envelope is considered in detail. The results indicate the quite wide circumstances under which ab initio phasing should be possible. PMID:26522408

  2. Automated macromolecular crystal detection system and method

    DOEpatents

    Christian, Allen T.; Segelke, Brent; Rupp, Bernard; Toppani, Dominique

    2007-06-05

    An automated macromolecular method and system for detecting crystals in two-dimensional images, such as light microscopy images obtained from an array of crystallization screens. Edges are detected from the images by identifying local maxima of a phase congruency-based function associated with each image. The detected edges are segmented into discrete line segments, which are subsequently geometrically evaluated with respect to each other to identify any crystal-like qualities such as, for example, parallel lines, facing each other, similarity in length, and relative proximity. And from the evaluation a determination is made as to whether crystals are present in each image.

  3. Fock spaces for modeling macromolecular complexes

    NASA Astrophysics Data System (ADS)

    Kinney, Justin

    Large macromolecular complexes play a fundamental role in how cells function. Here I describe a Fock space formalism for mathematically modeling these complexes. Specifically, this formalism allows ensembles of complexes to be defined in terms of elementary molecular ``building blocks'' and ``assembly rules.'' Such definitions avoid the massive redundancy inherent in standard representations, in which all possible complexes are manually enumerated. Methods for systematically computing ensembles of complexes from a list of components and interaction rules are described. I also show how this formalism readily accommodates coarse-graining. Finally, I introduce diagrammatic techniques that greatly facilitate the application of this formalism to both equilibrium and non-equilibrium biochemical systems.

  4. Structure of polymer layers adsorbed from concentrated solutions

    NASA Astrophysics Data System (ADS)

    Auvray, Loïc; Auroy, Philippe; Cruz, Margarida

    1992-06-01

    We study by neutron scattering the interfacial strucuture of poly(dimethylsiloxane) layers irreversibly adsorbed from concentrated solutions or melts. We first measure the thickness h of the layers swollen by a good solvent as a function of the chain polymerisation index N and of the polymer volume fraction in the initial solution Φ. The relation h ≈ N^{0.8}Φ^{0.3}, recently predicted from an analogy between irreversibly adsorbed layers and grafted polymer brushes, describes well our results. We can therefore deduce that there is at least one large loop of about N monomers per adsorbed chain. We also study the shape of the polymer concentration profile in the layers by measuring on two samples the polymer-solid partial structure factor, that is proportional to the Fourier transform of the profile. The model of pseudobrushes predicts a concentration decay varying with the distance of the wall z as z^{-2/5}. This power law profile accounts quantitatively for the angular variation of the polymer-solid cross structure factor but it is difficult to distinguish it without anbiguity from less singular profiles. It implies that the adsorption of PDMS onto silica is sufficiently strong and fast to quench completely the loop distribution in the initial layer. Nous étudions par diffusion de neutrons la structure interfaciale de couches de poly(diméthylsiloxane) irréversiblement adsorbées sur de la silice à partir de solutions semidiluées et de fondus. Nous mesurons d'abord l'épaisseur h des couches gonflées par un bon solvant en fonction du degré de polymérisation des chaînes N et de la fraction volumique dans la solution initiale Φ. La relation h≈ N^{0.8}Φ^{0.3} récemment prédite à partir de l'analogie entre couches irréversiblement adsorbées et brosses de polymères greffés décrit bien nos résultats. Nous en déduisons qu'il existe au moins une grande boucle d'environ N monomères par chaîne adsorbée. Nous étudions aussi la forme du profil de

  5. Solution structure of an A-tract DNA bend.

    PubMed

    MacDonald, D; Herbert, K; Zhang, X; Pologruto, T; Lu, P; Polgruto, T

    2001-03-01

    The solution structure of a DNA dodecamer d(GGCAAAAAACGG)/d(CCGTTTTTTGCC) containing an A-tract has been determined by NMR spectroscopy with residual dipolar couplings. The structure shows an overall helix axis bend of 19 degrees in a geometry consistent with solution and gel electrophoresis experiments. Fourteen degrees of the bending occurs in the GC regions flanking the A-tract. The remaining 5 degrees is spread evenly over its six AT base-pairs. The A-tract is characterized by decreasing minor groove width from the 5' to the 3' direction along the A strand. This is a result of propeller twist in the AT pairs and the increasing negative inclination of the adenine bases at the 3' side of the run of adenine bases. The four central thymine bases all have negative inclination throughout the A-tract with an average value of -6.1 degrees. Although this negative inclination makes the geometry of the A-tract different from all X-ray structures, the proton on N6 of adenine and the O4 of thymine one step down the helix are within distance to form bifurcated hydrogen bonds. The 5' bend of 4 degrees occurs at the junction between the GC flank and the A-tract through a combination of tilt and roll. The larger 3' bend, 10 degrees, occurs in two base steps: the first composed of tilt, -4.1 degrees, and the second a combination of tilt, -4.2 degrees, and roll, 6.0 degrees. This second step is a direct consequence of the change in inclination between an adjacent cytosine base, which has an inclination of -12 degrees, and the next base, a guanine, which has 3 degrees inclination. This bend is a combination of tilt and roll. The large change in inclination allows the formation of a hydrogen bond between the protons of N4 of the 3' cytosine and the O6 of the next 3' base, a guanine, stabilizing the roll component in the bend. These structural features differ from existing models for A-tract bends.For comparison, we also determined the structure of the control sequence, d

  6. Quaternary structure of carbonmonoxyhemoglobins in solution: structural changes induced by the allosteric effector inositol hexaphosphate.

    PubMed

    Gong, Qingguo; Simplaceanu, Virgil; Lukin, Jonathan A; Giovannelli, Janel L; Ho, Nancy T; Ho, Chien

    2006-04-25

    We have applied the residual dipolar coupling (RDC) method to investigate the solution quaternary structures of (2)H- and (15)N-labeled human normal adult recombinant hemoglobin (rHb A) and a low-oxygen-affinity mutant recombinant hemoglobin, rHb(alpha96Val-->Trp), both in the carbonmonoxy form, in the absence and presence of an allosteric effector, inositol hexaphosphate (IHP), using a stretched polyacrylamide gel as the alignment medium. Our recent RDC results [Lukin, J. A., Kontaxis, G., Simplaceanu, V., Yuan, Y., Bax, A., and Ho, C. (2003) Proc. Natl. Acad. Sci. U.S.A. 100, 517-520] indicate that the quaternary structure of HbCO A in solution is a dynamic ensemble between two previously determined crystal structures, R (crystals grown under high-salt conditions) and R2 (crystals grown under low-salt conditions). On the basis of a comparison of the geometric coordinates of the T, R, and R2 structures, it has been suggested that the oxygenation of Hb A follows the transition pathway from T to R and then to R2, with R being the intermediate structure [Srinivasan, R., and Rose, G. D. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 11113-11117]. The results presented here suggest that IHP can shift the solution quaternary structure of HbCO A slightly toward the R structure. The solution quaternary structure of rHbCO(alpha96Val-->Trp) in the absence of IHP is similar to that of HbCO A in the presence of IHP, consistent with rHbCO(alpha96Val-->Trp) having an affinity for oxygen lower than that of Hb A. Moreover, IHP has a much stronger effect in shifting the solution quaternary structure of rHbCO(alpha96Val-->Trp) toward the R structure and toward the T structure, consistent with IHP causing a more pronounced decrease in its oxygen affinity. The results presented in this work, as well as other results recently reported in the literature, clearly indicate that there are multiple quaternary structures for the ligated form of hemoglobin. These results also provide new insights

  7. Type IV kerogens as analogues for organic macromolecular materials in aqueously altered carbonaceous chondrites.

    PubMed

    Matthewman, Richard; Martins, Zita; Sephton, Mark A

    2013-04-01

    Understanding the processes involved in the evolution of organic matter in the early Solar System requires extensive experimental work. The scientifically valuable carbonaceous chondrites are principal targets for organic analyses, but these meteorites are rare. Meteoritic analog materials available in larger quantities, on which experiments can be performed, would be highly beneficial. The bulk of the organic inventory of carbonaceous chondrites is made up of solvent-insoluble macromolecular material. This high-molecular-weight entity provides a record of thermal and aqueous parent-body alteration of precursor organic structures present at the birth of the Solar System. To identify an effective analogue for this macromolecular material, we analyzed a series of terrestrial kerogens by pyrolysis-gas chromatography-mass spectrometry. Type I and II kerogens are unsuitable analogues owing to their highly aliphatic nature. Type III kerogens show some similarities to meteoritic macromolecular materials but display a substantial biological heritage. Type IV kerogens, in this study derived from Mesozoic paleosols and produced by the reworking and oxidation of organic matter, represent an effective analogue. Some isomeric differences exist between meteoritic macromolecular materials and type IV kerogens, and stepped pyrolysis indicates variations in thermal stability. In addition to being a suitable material for novel experimentation, type IV kerogens also have the potential to aid in the optimization of instruments for deployment on Mars. PMID:23551239

  8. Extracting trends from two decades of microgravity macromolecular crystallization history

    NASA Technical Reports Server (NTRS)

    Judge, Russell A.; Snell, Edward H.; van der Woerd, Mark J.

    2005-01-01

    Since the 1980s hundreds of macromolecular crystal growth experiments have been performed in the reduced acceleration environment of an orbiting spacecraft. Significant enhancements in structural knowledge have resulted from X-ray diffraction of the crystals grown. Similarly, many samples have shown no improvement or degradation in comparison to those grown on the ground. A complex series of interrelated factors affect these experiments and by building a comprehensive archive of the results it was aimed to identify factors that result in success and those that result in failure. Specifically, it was found that dedicated microgravity missions increase the chance of success when compared with those where crystallization took place as a parasitic aspect of the mission. It was also found that the chance of success could not be predicted based on any discernible property of the macromolecule available to us.

  9. Large-volume protein crystal growth for neutron macromolecular crystallography

    DOE PAGES

    Ng, Joseph D.; Baird, James K.; Coates, Leighton; Garcia-Ruiz, Juan M.; Hodge, Teresa A.; Huang, Sijay

    2015-03-30

    Neutron macromolecular crystallography (NMC) is the prevailing method for the accurate determination of the positions of H atoms in macromolecules. As neutron sources are becoming more available to general users, finding means to optimize the growth of protein crystals to sizes suitable for NMC is extremely important. Historically, much has been learned about growing crystals for X-ray diffraction. However, owing to new-generation synchrotron X-ray facilities and sensitive detectors, protein crystal sizes as small as in the nano-range have become adequate for structure determination, lessening the necessity to grow large crystals. Here, some of the approaches, techniques and considerations for themore » growth of crystals to significant dimensions that are now relevant to NMC are revisited. We report that these include experimental strategies utilizing solubility diagrams, ripening effects, classical crystallization techniques, microgravity and theoretical considerations.« less

  10. Large-volume protein crystal growth for neutron macromolecular crystallography

    SciTech Connect

    Ng, Joseph D.; Baird, James K.; Coates, Leighton; Garcia-Ruiz, Juan M.; Hodge, Teresa A.; Huang, Sijay

    2015-03-30

    Neutron macromolecular crystallography (NMC) is the prevailing method for the accurate determination of the positions of H atoms in macromolecules. As neutron sources are becoming more available to general users, finding means to optimize the growth of protein crystals to sizes suitable for NMC is extremely important. Historically, much has been learned about growing crystals for X-ray diffraction. However, owing to new-generation synchrotron X-ray facilities and sensitive detectors, protein crystal sizes as small as in the nano-range have become adequate for structure determination, lessening the necessity to grow large crystals. Here, some of the approaches, techniques and considerations for the growth of crystals to significant dimensions that are now relevant to NMC are revisited. We report that these include experimental strategies utilizing solubility diagrams, ripening effects, classical crystallization techniques, microgravity and theoretical considerations.

  11. Large-volume protein crystal growth for neutron macromolecular crystallography.

    PubMed

    Ng, Joseph D; Baird, James K; Coates, Leighton; Garcia-Ruiz, Juan M; Hodge, Teresa A; Huang, Sijay

    2015-04-01

    Neutron macromolecular crystallography (NMC) is the prevailing method for the accurate determination of the positions of H atoms in macromolecules. As neutron sources are becoming more available to general users, finding means to optimize the growth of protein crystals to sizes suitable for NMC is extremely important. Historically, much has been learned about growing crystals for X-ray diffraction. However, owing to new-generation synchrotron X-ray facilities and sensitive detectors, protein crystal sizes as small as in the nano-range have become adequate for structure determination, lessening the necessity to grow large crystals. Here, some of the approaches, techniques and considerations for the growth of crystals to significant dimensions that are now relevant to NMC are revisited. These include experimental strategies utilizing solubility diagrams, ripening effects, classical crystallization techniques, microgravity and theoretical considerations.

  12. Hydration structure of salt solutions from ab initio molecular dynamics

    SciTech Connect

    Bankura, Arindam; Carnevale, Vincenzo; Klein, Michael L.

    2013-01-07

    The solvation structures of Na{sup +}, K{sup +}, and Cl{sup -} ions in aqueous solution have been investigated using density functional theory (DFT) based Car-Parrinello (CP) molecular dynamics (MD) simulations. CPMD trajectories were collected for systems containing three NaCl or KCl ion pairs solvated by 122 water molecules using three different but commonly employed density functionals (BLYP, HCTH, and PBE) with electron correlation treated at the level of the generalized gradient approximation (GGA). The effect of including dispersion forces was analyzed through the use of an empirical correction to the DFT-GGA scheme. Special attention was paid to the hydration characteristics, especially the structural properties of the first solvation shell of the ions, which was investigated through ion-water radial distribution functions, coordination numbers, and angular distribution functions. There are significant differences between the present results obtained from CPMD simulations and those provided by classical MD based on either the CHARMM force field or a polarizable model. Overall, the computed structural properties are in fair agreement with the available experimental results. In particular, the observed coordination numbers 5.0-5.5, 6.0-6.4, and 6.0-6.5 for Na{sup +}, K{sup +}, and Cl{sup -}, respectively, are consistent with X-ray and neutron scattering studies but differ somewhat from some of the many other recent computational studies of these important systems. Possible reasons for the differences are discussed.

  13. Hydration structure of salt solutions from ab initio molecular dynamics

    NASA Astrophysics Data System (ADS)

    Bankura, Arindam; Carnevale, Vincenzo; Klein, Michael L.

    2013-01-01

    The solvation structures of Na^+, K^+, and Cl^- ions in aqueous solution have been investigated using density functional theory (DFT) based Car-Parrinello (CP) molecular dynamics (MD) simulations. CPMD trajectories were collected for systems containing three NaCl or KCl ion pairs solvated by 122 water molecules using three different but commonly employed density functionals (BLYP, HCTH, and PBE) with electron correlation treated at the level of the generalized gradient approximation (GGA). The effect of including dispersion forces was analyzed through the use of an empirical correction to the DFT-GGA scheme. Special attention was paid to the hydration characteristics, especially the structural properties of the first solvation shell of the ions, which was investigated through ion-water radial distribution functions, coordination numbers, and angular distribution functions. There are significant differences between the present results obtained from CPMD simulations and those provided by classical MD based on either the CHARMM force field or a polarizable model. Overall, the computed structural properties are in fair agreement with the available experimental results. In particular, the observed coordination numbers 5.0-5.5, 6.0-6.4, and 6.0-6.5 for Na^+, K^+, and Cl^-, respectively, are consistent with X-ray and neutron scattering studies but differ somewhat from some of the many other recent computational studies of these important systems. Possible reasons for the differences are discussed.

  14. EIGER detector: application in macromolecular crystallography.

    PubMed

    Casanas, Arnau; Warshamanage, Rangana; Finke, Aaron D; Panepucci, Ezequiel; Olieric, Vincent; Nöll, Anne; Tampé, Robert; Brandstetter, Stefan; Förster, Andreas; Mueller, Marcus; Schulze-Briese, Clemens; Bunk, Oliver; Wang, Meitian

    2016-09-01

    The development of single-photon-counting detectors, such as the PILATUS, has been a major recent breakthrough in macromolecular crystallography, enabling noise-free detection and novel data-acquisition modes. The new EIGER detector features a pixel size of 75 × 75 µm, frame rates of up to 3000 Hz and a dead time as low as 3.8 µs. An EIGER 1M and EIGER 16M were tested on Swiss Light Source beamlines X10SA and X06SA for their application in macromolecular crystallography. The combination of fast frame rates and a very short dead time allows high-quality data acquisition in a shorter time. The ultrafine ϕ-slicing data-collection method is introduced and validated and its application in finding the optimal rotation angle, a suitable rotation speed and a sufficient X-ray dose are presented. An improvement of the data quality up to slicing at one tenth of the mosaicity has been observed, which is much finer than expected based on previous findings. The influence of key data-collection parameters on data quality is discussed. PMID:27599736

  15. EIGER detector: application in macromolecular crystallography

    PubMed Central

    Casanas, Arnau; Warshamanage, Rangana; Finke, Aaron D.; Panepucci, Ezequiel; Olieric, Vincent; Nöll, Anne; Tampé, Robert; Brandstetter, Stefan; Förster, Andreas; Mueller, Marcus; Schulze-Briese, Clemens; Bunk, Oliver; Wang, Meitian

    2016-01-01

    The development of single-photon-counting detectors, such as the PILATUS, has been a major recent breakthrough in macromolecular crystallography, enabling noise-free detection and novel data-acquisition modes. The new EIGER detector features a pixel size of 75 × 75 µm, frame rates of up to 3000 Hz and a dead time as low as 3.8 µs. An EIGER 1M and EIGER 16M were tested on Swiss Light Source beamlines X10SA and X06SA for their application in macromolecular crystallography. The combination of fast frame rates and a very short dead time allows high-quality data acquisition in a shorter time. The ultrafine φ-slicing data-collection method is introduced and validated and its application in finding the optimal rotation angle, a suitable rotation speed and a sufficient X-ray dose are presented. An improvement of the data quality up to slicing at one tenth of the mosaicity has been observed, which is much finer than expected based on previous findings. The influence of key data-collection parameters on data quality is discussed. PMID:27599736

  16. The solution structure of the copper clioquinol complex.

    PubMed

    Pushie, M Jake; Nienaber, Kurt H; Summers, Kelly L; Cotelesage, Julien J H; Ponomarenko, Olena; Nichol, Helen K; Pickering, Ingrid J; George, Graham N

    2014-04-01

    Clioquinol (5-chloro-7-iodo-8-hydroxyquinoline) recently has shown promising results in the treatment of Alzheimer's disease and in cancer therapy, both of which also are thought to be due to clioquinol's ability as a lipophilic copper chelator. Previously, clioquinol was used as an anti-fungal and anti-protozoal drug that was responsible for an epidemic of subacute myelo-optic neuropathy (SMON) in Japan during the 1960s, probably a myeloneuropathy arising from a clioquinol-induced copper deficiency. Previous X-ray absorption spectroscopy of solutions of copper chelates of clioquinol suggested unusual coordination chemistry. Here we use a combination of electron paramagnetic, UV-visible and X-ray absorption spectroscopies to provide clarification of the chelation chemistry between clioquinol and copper. We find that the solution structures for the copper complexes formed with stoichiometric and excess clioquinol are conventional 8-hydroxyquinolate chelates. Thus, the promise of clioquinol in new treatments for Alzheimer's disease and in cancer therapy is not likely to be due to any novel chelation chemistry, but rather due to other factors including the high lipophilicity of the free ligand and chelate complexes.

  17. Characterization of Chitin and Chitosan Molecular Structure in Aqueous Solution

    SciTech Connect

    Franca, Eduardo D.; Lins, Roberto D.; Freitas, Luiz C.; Straatsma, t. P.

    2008-11-08

    Molecular dynamics simulations have been used to characterize the structure of chitin and chitosan fibers in aqueous solutions. Chitin fibers, whether isolated or in the form of a β-chitin nanoparticle, adopt the so-called 2-fold helix with Φ and φ values similar to its crystalline state. In solution, the intramolecular hydrogen bond HO3(n)•••O5(n+1) responsible for the 2-fold helical motif is stabilized by hydrogen bonds with water molecules in a well-defined orientation. On the other hand, chitosan can adopt five distinct helical motifs and its conformational equilibrium is highly dependent on pH. The hydrogen bond pattern and solvation around the O3 atom of insoluble chitosan (basic pH) are nearly identical to these quantities in chitin. Our findings suggest that the solubility and conformation of these polysaccharides are related to the stability of the intrachain HO3(n)•••O5(n+1) hydrogen bond, which is affect by the water exchange around the O3-HO3 hydroxyl group.

  18. Structure and phase behavior of aqueous methylcellulose solutions

    NASA Astrophysics Data System (ADS)

    McAllister, John; Schmidt, Peter; Lodge, Timothy; Bates, Frank

    2015-03-01

    Cellulose ethers (CE) constitute a multi-billion dollar industry, and have found end uses in a broad array of applications from construction materials, food products, personal care products, and pharmaceuticals for more than 80 years. Methylcellulose (MC, with the trade name METHOCEL™) is a CE in which there is a partial substitution of -OH groups with -OCH3 groups. This results in a polymer that is water-soluble at low temperatures, and aqueous solutions of MC display gelation and phase separation at higher temperatures. The nature of MC gelation has been debated for many years, and this project has made significant advances in the understanding of the solution properties of CEs. We have characterized a fibrillar structure of MC gels by cryogenic transmission electron microscopy (cryo-TEM) and small angle neutron scattering (SANS). Using light scattering, turbidity measurements, and dynamic mechanical spectroscopy (DMS) we report that MC microphase separates by nucleation and growth of fibril aggregates, and is a different process from LCST phase separation.

  19. Solid-state NMR in macromolecular systems: insights on how molecular entities move.

    PubMed

    Hansen, Michael Ryan; Graf, Robert; Spiess, Hans Wolfgang

    2013-09-17

    sciences have emphasized structure. By contrast, following X-ray crystallographers, researchers studying proteins using solution NMR introduced the combination of NMR with computer simulation before that became common practice in solid-state NMR. Today's simulation methods can handle partially ordered or even disordered systems common in synthetic polymers. Thus, the multitechnique approaches employed in NMR of synthetic and biological macromolecules have converged. Therefore, this Account will be relevant to both researchers studying synthetic macromolecular and supramolecular systems and those studying biological complexes.

  20. Rapid automated superposition of shapes and macromolecular models using spherical harmonics

    PubMed Central

    Konarev, Petr V.; Petoukhov, Maxim V.; Svergun, Dmitri I.

    2016-01-01

    A rapid algorithm to superimpose macromolecular models in Fourier space is proposed and implemented (SUPALM). The method uses a normalized integrated cross-term of the scattering amplitudes as a proximity measure between two three-dimensional objects. The reciprocal-space algorithm allows for direct matching of heterogeneous objects including high- and low-resolution models represented by atomic coordinates, beads or dummy residue chains as well as electron microscopy density maps and inhomogeneous multi-phase models (e.g. of protein–nucleic acid complexes). Using spherical harmonics for the computation of the amplitudes, the method is up to an order of magnitude faster than the real-space algorithm implemented in SUPCOMB by Kozin & Svergun [J. Appl. Cryst. (2001 ▸), 34, 33–41]. The utility of the new method is demonstrated in a number of test cases and compared with the results of SUPCOMB. The spherical harmonics algorithm is best suited for low-resolution shape models, e.g. those provided by solution scattering experiments, but also facilitates a rapid cross-validation against structural models obtained by other methods. PMID:27275142

  1. Analytical solution of thermal magnetization on memory stabilizer structures

    NASA Astrophysics Data System (ADS)

    Tomita, Yu; Viteri, C. Ricardo; Brown, Kenneth R.

    2010-10-01

    We return to the question of how the choice of stabilizer generators affects the preservation of information on structures whose degenerate ground state encodes a classical redundancy code. Controlled-not gates are used to transform the stabilizer Hamiltonian into a Hamiltonian consisting of uncoupled single spins and/or pairs of spins. This transformation allows us to obtain an analytical partition function and derive closed-form equations for the relative magnetization and susceptibility. These equations are in agreement with the numerical results presented in Viteri [Phys. Rev. APLRAAN1050-294710.1103/PhysRevA.80.042313 80, 042313 (2009)] for finite size systems. Analytical solutions show that there is no finite critical temperature, Tc=0, for all of the memory structures in the thermodynamic limit. This is in contrast to the previously predicted finite critical temperatures based on extrapolation. The mismatch is a result of the infinite system being a poor approximation even for astronomically large finite-size systems, where spontaneous magnetization still arises below an apparent finite critical temperature. We extend our analysis to the canonical stabilizer Hamiltonian. Interestingly, Hamiltonians with two-body interactions have a higher apparent critical temperature than the many-body Hamiltonian.

  2. The NMR solution structure of recombinant RGD-hirudin

    SciTech Connect

    Song, Xia; Mo, Wei; Liu, Xingang; Zhu, Lina; Yan, Xiaomin; Song, Houyan . E-mail: hysong@shmu.edu.cn; Dai, Linsen . E-mail: lsdai@fudan.edu.cn

    2007-08-17

    The solution structure of a new recombinant RGD-hirudin, which has the activities of anti-thrombin and anti-platelet aggregation, was determined by {sup 1}H nuclear magnetic resonance spectroscopy and compared with the conformations of recombinant wild-type hirudin and hirudin (variant 2, Lys47) of the hirudin thrombin complex. On the basis of total 1284 distance and dihedral angle constraints derived from a series of NMR spectra, 20 conformers were computed with ARIA/CNS programs. The structure of residues 3-30 and 37-48 form a molecular core with two antiparallel {beta}-sheets as the other two hirudins. However, significant differences were found in the surface electrostatic charge distributions among the three hirudins, especially in the RGD segment of recombinant RGD-hirudin. This difference may be greatly beneficial to its additional function of anti-platelet aggregation. The difference in extended C-terminal makes its both ionic and hydrophobic interactions with the fibrinogen recognition exosite of thrombin more effective.

  3. Analytical solution of thermal magnetization on memory stabilizer structures

    SciTech Connect

    Tomita, Yu; Viteri, C. Ricardo; Brown, Kenneth R.

    2010-10-15

    We return to the question of how the choice of stabilizer generators affects the preservation of information on structures whose degenerate ground state encodes a classical redundancy code. Controlled-not gates are used to transform the stabilizer Hamiltonian into a Hamiltonian consisting of uncoupled single spins and/or pairs of spins. This transformation allows us to obtain an analytical partition function and derive closed-form equations for the relative magnetization and susceptibility. These equations are in agreement with the numerical results presented in Viteri et al. [Phys. Rev. A 80, 042313 (2009)] for finite size systems. Analytical solutions show that there is no finite critical temperature, T{sub c}=0, for all of the memory structures in the thermodynamic limit. This is in contrast to the previously predicted finite critical temperatures based on extrapolation. The mismatch is a result of the infinite system being a poor approximation even for astronomically large finite-size systems, where spontaneous magnetization still arises below an apparent finite critical temperature. We extend our analysis to the canonical stabilizer Hamiltonian. Interestingly, Hamiltonians with two-body interactions have a higher apparent critical temperature than the many-body Hamiltonian.

  4. Macromolecular crowding fails to fold a globular protein in cells.

    PubMed

    Schlesinger, Alexander P; Wang, Yaqiang; Tadeo, Xavier; Millet, Oscar; Pielak, Gary J

    2011-06-01

    Proteins perform their functions in cells where macromolecular solutes reach concentrations of >300 g/L and occupy >30% of the volume. The volume excluded by these macromolecules stabilizes globular proteins because the native state occupies less space than the denatured state. Theory predicts that crowding can increase the ratio of folded to unfolded protein by a factor of 100, amounting to 3 kcal/mol of stabilization at room temperature. We tested the idea that volume exclusion dominates the crowding effect in cells using a variant of protein L, a 7 kDa globular protein with seven lysine residues replaced by glutamic acids; 84% of the variant molecules populate the denatured state in dilute buffer at room temperature, compared with 0.1% for the wild-type protein. We then used in-cell NMR spectroscopy to show that the cytoplasm of Escherichia coli does not overcome even this modest (∼1 kcal/mol) free-energy deficit. The data are consistent with the idea that nonspecific interactions between cytoplasmic components can overcome the excluded-volume effect. Evidence for these interactions is provided by the observations that adding simple salts folds the variant in dilute solution but increasing the salt concentration inside E. coli does not fold the protein. Our data are consistent with the results of other studies of protein stability in cells and suggest that stabilizing excluded-volume effects, which must be present under crowded conditions, can be ameliorated by nonspecific interactions between cytoplasmic components.

  5. Nitric Oxide Release Part I. Macromolecular Scaffolds

    PubMed Central

    Riccio, Daniel A.; Schoenfisch, Mark H.

    2012-01-01

    Summary The roles of nitric oxide (NO) in physiology and pathophysiology merit the use of NO as a therapeutic for certain biomedical applications. Unfortunately, limited NO payloads, too rapid NO release, and the lack of targeted NO delivery have hindered the clinical utility of NO gas and low molecular weight NO donor compounds. A wide-variety of NO-releasing macromolecular scaffolds has thus been developed to improve NO’s pharmacological potential. In this tutorial review, we provide an overview of the most promising NO release scaffolds including protein, organic, inorganic, and hybrid organic-inorganic systems. The NO release vehicles selected for discussion were chosen based on their enhanced NO storage, tunable NO release characteristics, and potential as therapeutics. PMID:22362355

  6. Panorama of ancient metazoan macromolecular complexes

    PubMed Central

    Wan, Cuihong; Borgeson, Blake; Phanse, Sadhna; Tu, Fan; Drew, Kevin; Clark, Greg; Xiong, Xuejian; Kagan, Olga; Kwan, Julian; Berzginov, Alexandr; Chessman, Kyle; Pal, Swati; Cromar, Graham; Papoulas, Ophelia; Ni, Zuyao; Boutz, Daniel R.; Stoilova, Snejana; Havugimana, Pierre C.; Guo, Xinghua; Malty, Ramy H.; Sarov, Mihail; Greenblatt, Jack; Babu, Mohan; Derry, Brent; Tillier, Elisabeth; Wallingford, John B.; Parkinson, John; Marcotte, Edward M.; Emili, Andrew

    2016-01-01

    Macromolecular complexes are essential to conserved biological processes, but their prevalence across animals is unclear. By combining extensive biochemical fractionation with quantitative mass spectrometry, we directly examined the composition of soluble multiprotein complexes among diverse metazoan models. Using an integrative approach, we then generated a draft conservation map consisting of >1 million putative high-confidence co-complex interactions for species with fully sequenced genomes that encompasses functional modules present broadly across all extant animals. Clustering revealed a spectrum of conservation, ranging from ancient Eukaryal assemblies likely serving cellular housekeeping roles for at least 1 billion years, ancestral complexes that have accrued contemporary components, and rarer metazoan innovations linked to multicellularity. We validated these projections by independent co-fractionation experiments in evolutionarily distant species, by affinity-purification and by functional analyses. The comprehensiveness, centrality and modularity of these reconstructed interactomes reflect their fundamental mechanistic significance and adaptive value to animal cell systems. PMID:26344197

  7. Macromolecular crowding explains overflow metabolism in cells

    PubMed Central

    Vazquez, Alexei; Oltvai, Zoltán N.

    2016-01-01

    Overflow metabolism is a metabolic phenotype of cells characterized by mixed oxidative phosphorylation (OxPhos) and fermentative glycolysis in the presence of oxygen. Recently, it was proposed that a combination of a protein allocation constraint and a higher proteome fraction cost of energy generation by OxPhos relative to fermentation form the basis of overflow metabolism in the bacterium, Escherichia coli. However, we argue that the existence of a maximum or optimal macromolecular density is another essential requirement. Here we re-evaluate our previous theory of overflow metabolism based on molecular crowding following the proteomic fractions formulation. We show that molecular crowding is a key factor in explaining the switch from OxPhos to overflow metabolism. PMID:27484619

  8. The spliceosome: a flexible, reversible macromolecular machine.

    PubMed

    Hoskins, Aaron A; Moore, Melissa J

    2012-05-01

    With more than a hundred individual RNA and protein parts and a highly dynamic assembly and disassembly pathway, the spliceosome is arguably the most complicated macromolecular machine in the eukaryotic cell. This complexity has made kinetic and mechanistic analysis of splicing incredibly challenging. Yet, recent technological advances are now providing tools for understanding this process in much greater detail. Ranging from genome-wide analyses of splicing and creation of an orthogonal spliceosome in vivo, to purification of active spliceosomes and observation of single molecules in vitro, such new experimental approaches are yielding significant insight into the inner workings of this remarkable machine. These experiments are rewriting the textbooks, with a new picture emerging of a dynamic, malleable machine heavily influenced by the identity of its pre-mRNA substrate.

  9. From “Simple” DNA-Protein Interactions to the Macromolecular Machines of Gene Expression

    PubMed Central

    von Hippel, Peter H.

    2008-01-01

    The physicochemical concepts that underlie our present ideas on the structure and assembly of the “macromolecular machines of gene expression” are developed, starting with the structure and folding of the individual protein and DNA components, the thermodynamics and kinetics of their conformational rearrangements during complex assembly, and the molecular basis of the sequence specificity and recognition interactions of the final assemblies that include the DNA genome. The role of diffusion in reduced dimensions in the kinetics of the assembly of macromolecular machines from their components is also considered, and diffusion-driven reactions are compared with those fueled by ATP binding and hydrolysis, as well as by the specific covalent chemical modifications involved in rearranging chromatin and modifying signal transduction networks in higher organisms. PMID:17477836

  10. Advances in electron microscopy: A qualitative view of instrumentation development for macromolecular imaging and tomography.

    PubMed

    Schröder, Rasmus R

    2015-09-01

    Macromolecular imaging and tomography of ice embedded samples has developed into a mature imaging technology, in structural biology today widely referred to simply as cryo electron microscopy.(1) While the pioneers of the technique struggled with ill-suited instruments, state-of-the-art cryo microscopes are now readily available and an increasing number of groups are producing excellent high-resolution structural data of macromolecular complexes, of cellular organelles, or the morphology of whole cells. Instrumentation developers, however, are offering yet more novel electron optical devices, such as energy filters and monochromators, aberration correctors or physical phase plates. Here we discuss how current instrumentation has already changed cryo EM, and how newly available instrumentation - often developed in other fields of electron microscopy - may further develop the use and applicability of cryo EM to the imaging of single isolated macromolecules of smaller size or molecules embedded in a crowded cellular environment.

  11. Reconstruction of SAXS Profiles from Protein Structures

    PubMed Central

    Putnam, Daniel K.; Lowe, Edward W.

    2013-01-01

    Small angle X-ray scattering (SAXS) is used for low resolution structural characterization of proteins often in combination with other experimental techniques. After briefly reviewing the theory of SAXS we discuss computational methods based on 1) the Debye equation and 2) Spherical Harmonics to compute intensity profiles from a particular macromolecular structure. Further, we review how these formulas are parameterized for solvent density and hydration shell adjustment. Finally we introduce our solution to compute SAXS profiles utilizing GPU acceleration. PMID:24688746

  12. The influence of interchain coupling on intramolecular oscillation mobility in coupled macromolecular chains: The case of coplanar parallel chains

    NASA Astrophysics Data System (ADS)

    Čevizović, D.; Petković, S.; Galović, S.; Chizhov, A.; Reshetnyak, A.

    2015-10-01

    We enlarge our results from the study of the hopping mechanism of the oscillation excitation transport in 1D model of one biologica-likel macromolecular chain to the case of a system composed from two 1D parallel macromolecular chains with consideration of the properties of intramolecular oscillation excitations. We suppose, that due to the exciton interaction with thermal oscillation (generated by mechanical phonon subsystem) of structural elements (consisting of the peptide group) of the chains, the exciton becomes by self trapped and forms the polaron state. We suggest a model which generalizes the modified Holstein polaron model to the case of two macromolecular chains and find that because of the interchain coupling, the exciton energy band is splitted into two subbands. The hopping process of exciton migration along the macromolecular chains is studied in dependence of system parameters and temperature. We pay an special attention to the temperature range (near T = 300 K) in which living cells operate. It is found that for the certain values of the system parameters there exists the abrupt change of the exciton migration nature from practically free (light) exciton motion to an immobile (heavy, dressed by phonon cloud) quasiparticle We discuss an application of the obtained results to the exciton transport both within deoxyribonucleic acid molecule and in the 2D polymer films organized from such macromolecular chains.

  13. Phylogenetic Diversity in the Macromolecular Composition of Microalgae.

    PubMed

    Finkel, Zoe V; Follows, Mick J; Liefer, Justin D; Brown, Chris M; Benner, Ina; Irwin, Andrew J

    2016-01-01

    The elemental stoichiometry of microalgae reflects their underlying macromolecular composition and influences competitive interactions among species and their role in the food web and biogeochemistry. Here we provide a new estimate of the macromolecular composition of microalgae using a hierarchical Bayesian analysis of data compiled from the literature. The median macromolecular composition of nutrient-sufficient exponentially growing microalgae is 32.2% protein, 17.3% lipid, 15.0% carbohydrate, 17.3% ash, 5.7% RNA, 1.1% chlorophyll-a and 1.0% DNA as percent dry weight. Our analysis identifies significant phylogenetic differences in macromolecular composition undetected by previous studies due to small sample sizes and the large inherent variability in macromolecular pools. The phylogenetic differences in macromolecular composition lead to variations in carbon-to-nitrogen ratios that are consistent with independent observations. These phylogenetic differences in macromolecular and elemental composition reflect adaptations in cellular architecture and biochemistry; specifically in the cell wall, the light harvesting apparatus, and storage pools. PMID:27228080

  14. Phylogenetic Diversity in the Macromolecular Composition of Microalgae

    PubMed Central

    Finkel, Zoe V.; Follows, Mick J.; Liefer, Justin D.; Brown, Chris M.; Benner, Ina; Irwin, Andrew J.

    2016-01-01

    The elemental stoichiometry of microalgae reflects their underlying macromolecular composition and influences competitive interactions among species and their role in the food web and biogeochemistry. Here we provide a new estimate of the macromolecular composition of microalgae using a hierarchical Bayesian analysis of data compiled from the literature. The median macromolecular composition of nutrient-sufficient exponentially growing microalgae is 32.2% protein, 17.3% lipid, 15.0% carbohydrate, 17.3% ash, 5.7% RNA, 1.1% chlorophyll-a and 1.0% DNA as percent dry weight. Our analysis identifies significant phylogenetic differences in macromolecular composition undetected by previous studies due to small sample sizes and the large inherent variability in macromolecular pools. The phylogenetic differences in macromolecular composition lead to variations in carbon-to-nitrogen ratios that are consistent with independent observations. These phylogenetic differences in macromolecular and elemental composition reflect adaptations in cellular architecture and biochemistry; specifically in the cell wall, the light harvesting apparatus, and storage pools. PMID:27228080

  15. Structure of Solutions of Multidimensional Conservation Laws with Discontinuous Flux and Applications to Uniqueness

    NASA Astrophysics Data System (ADS)

    Crasta, Graziano; De Cicco, Virginia; De Philippis, Guido; Ghiraldin, Francesco

    2016-08-01

    We investigate the structure of solutions of conservation laws with discontinuous flux under quite general assumption on the flux. We show that any entropy solution admits traces on the discontinuity set of the coefficients and we use this to prove the validity of a generalized Kato inequality for any pair of solutions. Applications to uniqueness of solutions are then given.

  16. Visualization of the atomic structure of solid solutions with the NaCl structure

    NASA Astrophysics Data System (ADS)

    Babanov, Yu. A.; Ponomarev, D. A.; Ustinov, V. V.

    2015-04-01

    It has been shown how an atomic cluster for a solid solution with a rock salt structure can be constructed using the Pauling model. Simulation has been performed for 343000 ions of Ni x Zn1 - x O3 ( x = 0, 0.3, 0.5, 0.7, 1.0) oxide substitutional solid solutions. Coordinates of all cluster ions are obtained and distribution functions of ion pairs (Ni-O, Ni-Ni, Ni-Zn, Zn-Zn, Zn-O, O-O) are constructed as functions of distance. The shape of the normal distribution indicates the existence of bounded chaos in the system of oxide solid solutions. The width of the Gaussian distribution function is determined by the difference of metal ionic radii. The results are in agreement with both X-ray diffraction and EXAFS spectroscopy data.

  17. a Procedural Solution to Model Roman Masonry Structures

    NASA Astrophysics Data System (ADS)

    Cappellini, V.; Saleri, R.; Stefani, C.; Nony, N.; De Luca, L.

    2013-07-01

    The paper will describe a new approach based on the development of a procedural modelling methodology for archaeological data representation. This is a custom-designed solution based on the recognition of the rules belonging to the construction methods used in roman times. We have conceived a tool for 3D reconstruction of masonry structures starting from photogrammetric surveying. Our protocol considers different steps. Firstly we have focused on the classification of opus based on the basic interconnections that can lead to a descriptive system used for their unequivocal identification and design. Secondly, we have chosen an automatic, accurate, flexible and open-source photogrammetric pipeline named Pastis Apero Micmac - PAM, developed by IGN (Paris). We have employed it to generate ortho-images from non-oriented images, using a user-friendly interface implemented by CNRS Marseille (France). Thirdly, the masonry elements are created in parametric and interactive way, and finally they are adapted to the photogrammetric data. The presented application, currently under construction, is developed with an open source programming language called Processing, useful for visual, animated or static, 2D or 3D, interactive creations. Using this computer language, a Java environment has been developed. Therefore, even if the procedural modelling reveals an accuracy level inferior to the one obtained by manual modelling (brick by brick), this method can be useful when taking into account the static evaluation on buildings (requiring quantitative aspects) and metric measures for restoration purposes.

  18. Computational tools for analysing structural changes in proteins in solution.

    PubMed

    Noé, Frank; Schwarzl, Sonja M; Fischer, Stefan; Smith, Jeremy C

    2003-01-01

    Many important structural changes in proteins involve long-time dynamics, which are outside the timescale presently accessible by a straightforward integration of Newton's equations of motion. This problem is addressed with minimisation-based algorithms, which are applied on possible reaction pathways using atomic-detail models. For reasons of efficiency, an implicit treatment of solvent is imperative. We present the charge reparameterisation protocol, which is a method that approximates the interaction energies obtained by a numerical solution of the Poisson-Boltzmann equation. Furthermore, we present a number of methods that can be used to compute possible reaction pathways associated with a particular conformational change. Two of them, the self-penalty walk and the nudged elastic band method, define an objective function, which is minimised to find optimal paths. A third method, conjugate peak refinement, is a heuristic method, which finds minimum energy paths without the use of an explicit objective function. Finally, we discuss problems and limitations with these methods and give a perspective on future research.

  19. Solution structures of stromelysin complexed to thiadiazole inhibitors.

    PubMed Central

    Stockman, B. J.; Waldon, D. J.; Gates, J. A.; Scahill, T. A.; Kloosterman, D. A.; Mizsak, S. A.; Jacobsen, E. J.; Belonga, K. L.; Mitchell, M. A.; Mao, B.; Petke, J. D.; Goodman, L.; Powers, E. A.; Ledbetter, S. R.; Kaytes, P. S.; Vogeli, G.; Marshall, V. P.; Petzold, G. L.; Poorman, R. A.

    1998-01-01

    Unregulated or overexpressed matrix metalloproteinases (MMPs), including stromelysin, collagenase, and gelatinase. have been implicated in several pathological conditions including arthritis and cancer. Small-molecule MMP inhibitors may have therapeutic value in the treatment of these diseases. In this regard, the solution structures of two stromelysin/ inhibitor complexes have been investigated using 1H, 13C, and 15N NMR spectroscopy. Both-inhibitors are members of a novel class of matrix metalloproteinase inhibitor that contain a thiadiazole group and that interact with stromelysin in a manner distinct from other classes of inhibitors. The inhibitors coordinate the catalytic zinc atom through their exocyclic sulfur atom, with the remainder of the ligand extending into the S1-S3 side of the active site. The binding of inhibitor containing a protonated or fluorinated aromatic ring was investigated using 1H and 19F NMR spectroscopy. The fluorinated ring was found to have a reduced ring-flip rate compared to the protonated version. A strong, coplanar interaction between the fluorinated ring of the inhibitor and the aromatic ring of Tyr155 is proposed to account for the reduced ring-flip rate and for the increase in binding affinity observed for the fluorinated inhibitor compared to the protonated inhibitor. Binding interactions observed for the thiadiazole class of ligands have implications for the design of matrix metalloproteinase inhibitors. PMID:9827994

  20. Automated macromolecular model building for X-ray crystallography using ARP/wARP version 7

    PubMed Central

    Langer, Gerrit G; Cohen, Serge X; Lamzin, Victor S; Perrakis, Anastassis

    2008-01-01

    ARP/wARP is a software suite to build macromolecular models in X-ray crystallography electron density maps. Structural genomics initiatives and the study of complex macromolecular assemblies and membrane proteins all rely on advanced methods for 3D structure determination. ARP/wARP meets these needs by providing the tools to obtain a macromolecular model automatically, with a reproducible computational procedure. ARP/wARP 7.0 tackles several tasks: iterative protein model building including a high-level decision-making control module; fast construction of the secondary structure of a protein; building flexible loops in alternate conformations; fully automated placement of ligands, including a choice of the best fitting ligand from a “cocktail”; and finding ordered water molecules. All protocols are easy to handle by a non-expert user through a graphical user interface or a command line. The time required is typically a few minutes although iterative model building may take a few hours. PMID:18600222

  1. Valence Electronic Structure of Aqueous Solutions: Insights from Photoelectron Spectroscopy

    NASA Astrophysics Data System (ADS)

    Seidel, Robert; Winter, Bernd; Bradforth, Stephen E.

    2016-05-01

    The valence orbital electron binding energies of water and of embedded solutes are crucial quantities for understanding chemical reactions taking place in aqueous solution, including oxidation/reduction, transition-metal coordination, and radiation chemistry. Their experimental determination based on liquid-photoelectron spectroscopy using soft X-rays is described, and we provide an overview of valence photoelectron spectroscopy studies reported to date. We discuss principal experimental aspects and several theoretical approaches to compute the measured binding energies of the least tightly bound molecular orbitals. Solutes studied are presented chronologically, from simple electrolytes, via transition-metal ion solutions and several organic and inorganic molecules, to biologically relevant molecules, including aqueous nucleotides and their components. In addition to the lowest vertical ionization energies, the measured valence photoelectron spectra also provide information on adiabatic ionization energies and reorganization energies for the oxidation (ionization) half-reaction. For solutes with low solubility, resonantly enhanced ionization provides a promising alternative pathway.

  2. Valence Electronic Structure of Aqueous Solutions: Insights from Photoelectron Spectroscopy.

    PubMed

    Seidel, Robert; Winter, Bernd; Bradforth, Stephen E

    2016-05-27

    The valence orbital electron binding energies of water and of embedded solutes are crucial quantities for understanding chemical reactions taking place in aqueous solution, including oxidation/reduction, transition-metal coordination, and radiation chemistry. Their experimental determination based on liquid-photoelectron spectroscopy using soft X-rays is described, and we provide an overview of valence photoelectron spectroscopy studies reported to date. We discuss principal experimental aspects and several theoretical approaches to compute the measured binding energies of the least tightly bound molecular orbitals. Solutes studied are presented chronologically, from simple electrolytes, via transition-metal ion solutions and several organic and inorganic molecules, to biologically relevant molecules, including aqueous nucleotides and their components. In addition to the lowest vertical ionization energies, the measured valence photoelectron spectra also provide information on adiabatic ionization energies and reorganization energies for the oxidation (ionization) half-reaction. For solutes with low solubility, resonantly enhanced ionization provides a promising alternative pathway.

  3. Comparison of the crystal and solution structures of two RNA oligonucleotides.

    PubMed Central

    Rife, J P; Stallings, S C; Correll, C C; Dallas, A; Steitz, T A; Moore, P B

    1999-01-01

    Until recently, there were no examples of RNAs whose structures had been determined by both NMR and x-ray crystallography, and thus there was no experimental basis for assessing the accuracy of RNA solution structures. A comparison of the solution and the crystal structures of two RNAs is presented, which demonstrates that NMR can produce solution structures that resemble crystal structures and thus validates the application to RNA of a methodology developed initially for the determination of protein conformations. Models for RNA solution structures are appreciably affected by the parameters used for their refinement that describe intramolecular interactions. For the RNAs of interest here, the more realistic those parameters, the greater the similarity between solution structures and crystal structures. PMID:9876123

  4. Radiation damage in macromolecular crystallography: what is it and why should we care?

    PubMed Central

    Garman, Elspeth F.

    2010-01-01

    Radiation damage inflicted during diffraction data collection in macromolecular crystallography has re-emerged in the last decade as a major experimental and computational challenge, as even for crystals held at 100 K it can result in severe data-quality degradation and the appearance in solved structures of artefacts which affect biological interpretations. Here, the observable symptoms and basic physical processes involved in radiation damage are described and the concept of absorbed dose as the basic metric against which to monitor the experimentally observed changes is outlined. Investigations into radiation damage in macromolecular crystallography are ongoing and the number of studies is rapidly increasing. The current literature on the subject is compiled as a resource for the interested researcher. PMID:20382986

  5. Romp: The Method of Choice for Precise Macromolecular Engineering and Synthesis of Smart Materials

    NASA Astrophysics Data System (ADS)

    Khosravi, Ezat; Castle, Thomas C.; Kujawa, Margaret; Leejarkpai, Jan; Hutchings, Lian R.; Hine, Peter J.

    The recent advances in olefin metathesis highlight the impact of Ring Opening Metathesis Polymerisation (ROMP) as a powerful technique for macromolecular engineering and synthesis of smart materials with well-defined structures. ROMP has attracted a considerable research attention recently particularly by industry largely due to the development of well-defined metal complexes as initiators and also because of the award of the Noble Prize for Chemistry in 2005 to three scientists (Chauvin, Grubbs, Schrock) for their contributions in this area. This chapter discusses several interesting examples in order to demonstrate that ROMP is a power tool in macromolecular engineering and that it allows the design and synthesis of polymers with novel topologies.

  6. Sequential recovery of macromolecular components of the nucleolus.

    PubMed

    Bai, Baoyan; Laiho, Marikki

    2015-01-01

    The nucleolus is involved in a number of cellular processes of importance to cell physiology and pathology, including cell stress responses and malignancies. Studies of macromolecular composition of the nucleolus depend critically on the efficient extraction and accurate quantification of all macromolecular components (e.g., DNA, RNA, and protein). We have developed a TRIzol-based method that efficiently and simultaneously isolates these three macromolecular constituents from the same sample of purified nucleoli. The recovered and solubilized protein can be accurately quantified by the bicinchoninic acid assay and assessed by polyacrylamide gel electrophoresis or by mass spectrometry. We have successfully applied this approach to extract and quantify the responses of all three macromolecular components in nucleoli after drug treatments of HeLa cells, and conducted RNA-Seq analysis of the nucleolar RNA.

  7. Methods for methotrexate determination in macromolecular conjugates drug carrier.

    PubMed

    Ciekot, Jarosław; Goszczyński, Tomasz; Boratyńskit, Janusz

    2012-01-01

    In this paper, two simple, cost-effective and fast methods for quantification of methotrexate (MTX) in the macromolecular conjugates are presented. The method for analysis of total MTX in preparations was based on absorption spectrophotometry. Validation was performed by measuring absorbance at 372 nm of the sodium bicarbonate solution. Curve describing drug concentration against absorption had a linear character in the range of 1.204-40.13 microM. The reproducibility and precision of method was 0.1558 to 3.086%. The recovery of the method was between 99.56 and 104.7%. The limit of quantitation method was 1.050 microM. The method for free MTX determination was based on size exclusion chromatography and UV-VIS detection at the wavelength of 302 nm. Superdex Peptide column (150 x 4.6 mm) and a mobile phase 0.1 M sodium bicarbonate with a flow rate of 0.4 mL/min was used. In the free drug determination method, the curve had a linear character in the range of 2.006-200.6 microM. The reproducibility and precision of method was 0.3761 to 2.452%. The recovery of the method was between 93.18 and 104.5%. The limit of quantitation method was 0.9203 microM. PMID:23285700

  8. JBluIce-EPICS control system for macromolecular crystallography.

    SciTech Connect

    Stepanov, S.; Makarov, O.; Hilgart, M.; Pothineni, S.; Urakhchin, A.; Devarapalli, S.; Yoder, D.; Becker, M.; Ogata, C.; Sanishvili, R.; Nagarajan, V.; Smith, J. L.; Fischetti, R. F.

    2011-01-01

    The trio of macromolecular crystallography beamlines constructed by the General Medicine and Cancer Institutes Collaborative Access Team (GM/CA-CAT) in Sector 23 of the Advanced Photon Source (APS) have been in growing demand owing to their outstanding beam quality and capacity to measure data from crystals of only a few micrometres in size. To take full advantage of the state-of-the-art mechanical and optical design of these beamlines, a significant effort has been devoted to designing fast, convenient, intuitive and robust beamline controls that could easily accommodate new beamline developments. The GM/CA-CAT beamline controls are based on the power of EPICS for distributed hardware control, the rich Java graphical user interface of Eclipse RCP and the task-oriented philosophy as well as the look and feel of the successful SSRL BluIce graphical user interface for crystallography. These beamline controls feature a minimum number of software layers, the wide use of plug-ins that can be written in any language and unified motion controls that allow on-the-fly scanning and optimization of any beamline component. This paper describes the ways in which BluIce was combined with EPICS and converted into the Java-based JBluIce, discusses the solutions aimed at streamlining and speeding up operations and gives an overview of the tools that are provided by this new open-source control system for facilitating crystallographic experiments, especially in the field of microcrystallography.

  9. The Lunar Internal Structure Model: Problems and Solutions

    NASA Astrophysics Data System (ADS)

    Nefedyev, Yuri; Gusev, Alexander; Petrova, Natalia; Varaksina, Natalia

    decomposition of gravitational field of the Moon of members up to 165th order with a high degree of accuracy. Judging from the given data, the distinctive feature of the Moon’s gravitational field is that harmonics of the third and even the fourth order are comparable with harmonics of the second order, except for member J2. General conclusion: according to recent data, the true figure of the Moon is much more complex than a three-axis ellipsoid. Gravitational field and dynamic figure of the multilayered Moon: One of the main goals of selenodesy is the study of a dynamic figure of the Moon which determines distribution of the mass within the Moon’s body. A dynamic figure is shaped by the inertia ellipsoid set by values of resultant moments of inertia of the Moon A, B, C and their orientation in space. Selenoid satellites (SS) open new and most perspective opportunities in the study of gravitational field and the Moon’s figure. SSs “Moon 10”, “Apollo”, “Clementine”, “Lunar Prospector” trajectory tracking data processing has allowed for identification of coefficients in decomposition of gravitational field of the Moon of members up to 165th order with a high degree of accuracy. Judging from the given data, the distinctive feature of the Moon’s gravitational field is that harmonics of the third and even the fourth order are comparable with harmonics of the second order. Difference from zero of c-coefficients proves asymmetry of gravitational fields on the visible and invisible sides of the Moon. As a first attempt at solving the problem, the report presents the survey of internal structure of the Moon, tabulated values of geophysical parameters and geophysical profile of the Moon, including liquid lunar core, analytical solution of Clairaut’s equation for the two-layer model of the Moon; mathematical and bifurcational analysis of solution based on physically justified task options; original debugged software in VBA programming language for computer

  10. Elucidating transient macromolecular interactions using paramagnetic relaxation enhancement

    PubMed Central

    Clore, G. Marius; Tang, Chun; Iwahara, Junji

    2007-01-01

    Recent advances in the use of paramagnetic relaxation enhancement (PRE) in structure refinement and in the analysis of transient dynamic processes involved in macromolecular complex formation are presented. In the slow exchange regime, we show, using the SRY/DNA complex as an example, that the PRE provides a powerful tool that can lead to significant increases in the reliability and accuracy of NMR structure determinations. Refinement necessitates the use of an ensemble representation of the paramagnetic center and a model free extension of the Solomon-Bloembergen equations. In the fast exchange regime, the PRE provides insight into dynamic processes and the existence of transient, low population intermediate species. The PRE allows one to characterize dynamic non-specific binding of a protein to DNA; to directly demonstrate that the search process whereby a transcription factor locates its specific DNA target site involves both intramolecular (sliding) and intermolecular (hopping and intersegment transfer) translocation; and to detect and visualize the distribution of an ensemble of transient encounter complexes in protein-protein association. PMID:17913493

  11. Acute iron poisoning. Rescue with macromolecular chelators.

    PubMed Central

    Mahoney, J R; Hallaway, P E; Hedlund, B E; Eaton, J W

    1989-01-01

    Acute iron intoxication is a frequent, sometimes life-threatening, form of poisoning. Present therapy, in severe cases, includes oral and intravenous administration of the potent iron chelator, deferoxamine. Unfortunately, high dose intravenous deferoxamine causes acute hypotension additive with that engendered by the iron poisoning itself. To obviate this problem, we have covalently attached deferoxamine to high molecular weight carbohydrates such as dextran and hydroxyethyl starch. These macromolecular forms of deferoxamine do not cause detectable decreases in blood pressure of experimental animals, even when administered intravenously in very large doses, and persist in circulation much longer than the free drug. These novel iron-chelating substances, but not deferoxamine itself, will prevent mortality from otherwise lethal doses of iron administered to mice either orally or intraperitoneally. Further reflecting this enhanced therapeutic efficacy, the high molecular weight iron chelators also abrogate iron-mediated hepatotoxicity, suppressing the release of alanine aminotransferase. We conclude that high molecular weight derivatives of deferoxamine hold promise for the effective therapy of acute iron intoxication and may also be useful in other clinical circumstances in which control of free, reactive iron is therapeutically desirable. PMID:2794068

  12. Solution structure of monomeric BsaL, the type III secretion needle protein of Burkholderia pseudomallei.

    PubMed

    Zhang, Lingling; Wang, Yu; Picking, Wendy L; Picking, William D; De Guzman, Roberto N

    2006-06-01

    Many gram-negative bacteria that are important human pathogens possess type III secretion systems as part of their required virulence factor repertoire. During the establishment of infection, these pathogens coordinately assemble greater than 20 different proteins into a macromolecular structure that spans the bacterial inner and outer membranes and, in many respects, resembles and functions like a syringe. This type III secretion apparatus (TTSA) is used to inject proteins into a host cell's membrane and cytoplasm to subvert normal cellular processes. The external portion of the TTSA is a needle that is composed of a single type of protein that is polymerized in a helical fashion to form an elongated tube with a central channel of 2-3 nm in diameter. TTSA needle proteins from a variety of bacterial pathogens share sequence conservation; however, no atomic structure for any TTSA needle protein is yet available. Here, we report the structure of a TTSA needle protein called BsaL from Burkholderia pseudomallei determined by nuclear magnetic resonance (NMR) spectroscopy. The central part of the protein assumes a helix-turn-helix core domain with two well-defined alpha-helices that are joined by an ordered, four-residue linker. This forms a two-helix bundle that is stabilized by interhelix hydrophobic contacts. Residues that flank this presumably exposed core region are not completely disordered, but adopt a partial helical conformation. The atomic structure of BsaL and its sequence homology with other TTSA needle proteins suggest potentially unique structural dynamics that could be linked with a universal mechanism for control of type III secretion in diverse gram-negative bacterial pathogens.

  13. Effects of macromolecular crowding on a small lipid binding protein probed at the single-amino acid level.

    PubMed

    Pérez Santero, Silvia; Favretto, Filippo; Zanzoni, Serena; Chignola, Roberto; Assfalg, Michael; D'Onofrio, Mariapina

    2016-09-15

    Macromolecular crowding is a distinctive feature of the cellular interior, influencing the behaviour of biomacromolecules. Despite significant advancements in the description of the effects of crowding on global protein properties, the influence of cellular components on local protein attributes has received limited attention. Here, we describe a residue-level systematic interrogation of the structural, dynamic, and binding properties of the liver fatty acid binding protein (LFABP) in crowded solutions. Two-dimensional NMR spectral fingerprints and relaxation data were collected on LFABP in the presence of polymeric and biomolecular crowders. Non-interacting crowders produced minimal site-specific spectral perturbations on ligand-free and lipid-bound LFABP. Conformational adaptations upon ligand binding reproduced those observed in dilute solution, but a perturbation of the free oleate state resulted in less favorable uptake. When LFABP engaged in direct interactions with background molecules, changes in local chemical environments were detected for residues of the internal binding pocket and of the external surface. Enhanced complexity was introduced by investigating LFABP in cell lysates, and in membrane-bounded compartments. LFABP was able to capture ligands from prokaryotic and eukaryotic cell lysates, and from artificial cells (water-in-oil emulsion droplets). The data suggest that promiscuous interactions are a major factor influencing protein function in the cell. PMID:27457417

  14. The structure of ionic aqueous solutions at interfaces: an intrinsic structure analysis.

    PubMed

    Bresme, Fernando; Chacón, Enrique; Tarazona, Pedro; Wynveen, Aaron

    2012-09-21

    We investigate the interfacial structure of ionic solutions consisting of alkali halide ions in water at concentrations in the range 0.2-1.0 molal and at 300 K. Combining molecular dynamics simulations of point charge ion models and a recently introduced computational approach that removes the averaging effect of interfacial capillary waves, we compute the intrinsic structure of the aqueous interface. The interfacial structure is more complex than previously inferred from the analysis of mean profiles. We find a strong alternating double layer structure near the interface, which depends on the cation and anion size. Relatively small changes in the ion diameter disrupt the double layer structure, promoting the adsorption of anions or inducing the density enhancement of small cations with diameters used in simulation studies of lithium solutions. The density enhancement of the small cations is mediated by their strong water solvation shell, with one or more water molecules "anchoring" the ion to the outermost water layer. We find that the intrinsic interfacial electrostatic potential features very strong oscillations with a minimum at the liquid surface that is ∼4 times stronger than the electrostatic potential in the bulk. For the water model employed in this work, SPC/E, the electrostatic potential at the water surface is ∼-2 V, equivalent to ∼80 k(B)T (for T = 300 K), much stronger than previously considered. Furthermore, we show that the utilization of the intrinsic surface technique provides a route to extract ionic potentials of mean force that are not affected by the thermal fluctuations, which limits the accuracy of most past approaches including the popular umbrella sampling technique. PMID:22998280

  15. Structural qualia: a solution to the hard problem of consciousness

    PubMed Central

    Loorits, Kristjan

    2014-01-01

    The hard problem of consciousness has been often claimed to be unsolvable by the methods of traditional empirical sciences. It has been argued that all the objects of empirical sciences can be fully analyzed in structural terms but that consciousness is (or has) something over and above its structure. However, modern neuroscience has introduced a theoretical framework in which also the apparently non-structural aspects of consciousness, namely the so called qualia or qualitative properties, can be analyzed in structural terms. That framework allows us to see qualia as something compositional with internal structures that fully determine their qualitative nature. Moreover, those internal structures can be identified which certain neural patterns. Thus consciousness as a whole can be seen as a complex neural pattern that misperceives some of its own highly complex structural properties as monadic and qualitative. Such neural pattern is analyzable in fully structural terms and thereby the hard problem is solved. PMID:24672510

  16. The use of a mini-κ goniometer head in macromolecular crystallography diffraction experiments

    PubMed Central

    Brockhauser, Sandor; Ravelli, Raimond B. G.; McCarthy, Andrew A.

    2013-01-01

    Most macromolecular crystallography (MX) diffraction experiments at synchrotrons use a single-axis goniometer. This markedly contrasts with small-molecule crystallography, in which the majority of the diffraction data are collected using multi-axis goniometers. A novel miniaturized κ-gonio­meter head, the MK3, has been developed to allow macromolecular crystals to be aligned. It is available on the majority of the structural biology beamlines at the ESRF, as well as elsewhere. In addition, the Strategy for the Alignment of Crystals (STAC) software package has been developed to facilitate the use of the MK3 and other similar devices. Use of the MK3 and STAC is streamlined by their incorporation into online analysis tools such as EDNA. The current use of STAC and MK3 on the MX beamlines at the ESRF is discussed. It is shown that the alignment of macromolecular crystals can result in improved diffraction data quality compared with data obtained from randomly aligned crystals. PMID:23793150

  17. Conformational Statistics of Semi-Flexible Macromolecular Chains with Internal Joints

    PubMed Central

    Zhou, Yu

    2010-01-01

    Fluctuations in the bending angles at internal irregularities of DNA and RNA (such as symmetric loops, bulges, and nicks/gaps) have been observed from various experiments. However, little effort has been made to computationally predict and explain the statistical behavior of semi-flexible chains with internal defects. In this paper, we describe the general structure of these macromolecular chains as inextensible elastic chains with one or more internal joints which have limited ranges of rotation, and propose a method to compute the probability density functions of the end-to-end pose of these macromolecular chains. Our method takes advantage of the operational properties of the non-commutative Fourier transform for the group of rigid-body motions in three-dimensional space, SE(3). Two representative types of joints, the hinge for planar rotation and the ball joint for spatial rotation, are discussed in detail. The proposed method applies to various stiffness models of semi-flexible chain-like macromolecules. Examples are calculated using the Kratky-Porod model with specified stiffness, angular fluctuation, and joint locations. Entropic effects associated with internal angular fluctuations of semi-flexible macromolecular chains with internal joints can be computed using this formulation. Our method also provides a potential tool to detect the existence of internal irregularities. PMID:21243113

  18. Macromolecular Imaging Agents Containing Lanthanides: Can Conceptual Promise Lead to Clinical Potential?

    PubMed Central

    Bryson, Joshua; Reineke, Jeffrey W.; Reineke, Theresa M.

    2012-01-01

    Macromolecular magnetic resonance imaging (MRI) contrast agents are increasingly being used to improve the resolution of this noninvasive diagnostic technique. All clinically-approved T1 contrast agents are small molecule chelates of gadolinium [Gd(III)] that affect bound water proton relaxivity. Both the small size and monomeric nature of these agents ultimately limits the image resolution enhancement that can be achieved for both contrast enhancement and pharmacokinetic/biodistribution reasons. The multimeric nature of macromolecules, such as polymers, dendrimers, and noncovalent complexes of small molecule agents with proteins, have been shown to significantly increase the image contrast and resolution due to their large size and ability to incorporate multiple Gd(III) chlelation sites. Also, macromolecular agents are advantageous as they have the ability to be designed to be nontoxic, hydrophilic, easily purified, aggregation-resistant, and have controllable three-dimensional macromolecular structure housing the multiple lanthanide chelation sites. For these reasons, large molecule diagnostics have the ability to significantly increase the relaxivity of water protons within the targeted tissues and thus the image resolution for many diagnostic applications. The FDA approval of a contrast agent that consists of a reversible, non-covalent coupling of a small Gd(III) chelate with serum albumin for blood pool imaging (marketed under the trade names of Vasovist and Ablivar) proved to be one of the first diagnostic agent to capitalize on these benefits from macromolecular association in humans. However, much research and development is necessary to optimize the safety of these unique agents for in vivo use and potential clinical development. To this end, recent work in the field of polymer, dendrimer, and noncovalent complex-based imaging agents are reviewed herein and the future outlook of this field is discussed. PMID:23467737

  19. Application of finite-element-based solution technologies for viscoplastic structural analyses

    NASA Technical Reports Server (NTRS)

    Arya, V. K.

    1990-01-01

    Finite-element solution technology developed for use in conjunction with advanced viscoplastic models is described. The development of such solution technology is necessary for performing stress/life analyses of engineering structural problems where the complex geometries and loadings make the conventional analytical solutions difficult. The versatility of the solution technology is demonstrated by applying it to viscoplastic models possessing different mathematical structures and encompassing isotropic and anisotropic material. The computational results qualitatively replicate deformation behavior observed in experiments on prototypical structural components.

  20. Synchrotron radiation macromolecular crystallography: science and spin-offs

    PubMed Central

    Helliwell, John R.; Mitchell, Edward P.

    2015-01-01

    A current overview of synchrotron radiation (SR) in macromolecular crystallography (MX) instrumentation, methods and applications is presented. Automation has been and remains a central development in the last decade, as have the rise of remote access and of industrial service provision. Results include a high number of Protein Data Bank depositions, with an increasing emphasis on the successful use of microcrystals. One future emphasis involves pushing the frontiers of using higher and lower photon energies. With the advent of X-ray free-electron lasers, closely linked to SR developments, the use of ever smaller samples such as nanocrystals, nanoclusters and single molecules is anticipated, as well as the opening up of femtosecond time-resolved diffraction structural studies. At SR sources, a very high-throughput assessment for the best crystal samples and the ability to tackle just a few micron and sub-micron crystals will become widespread. With higher speeds and larger detectors, diffraction data volumes are becoming long-term storage and archiving issues; the implications for today and the future are discussed. Together with the rise of the storage ring to its current pre-eminence in MX data provision, the growing tendency of central facility sites to offer other centralized facilities complementary to crystallography, such as cryo-electron microscopy and NMR, is a welcome development. PMID:25866664

  1. Synchrotron radiation macromolecular crystallography: science and spin-offs.

    PubMed

    Helliwell, John R; Mitchell, Edward P

    2015-03-01

    A current overview of synchrotron radiation (SR) in macromolecular crystallography (MX) instrumentation, methods and applications is presented. Automation has been and remains a central development in the last decade, as have the rise of remote access and of industrial service provision. Results include a high number of Protein Data Bank depositions, with an increasing emphasis on the successful use of microcrystals. One future emphasis involves pushing the frontiers of using higher and lower photon energies. With the advent of X-ray free-electron lasers, closely linked to SR developments, the use of ever smaller samples such as nanocrystals, nanoclusters and single molecules is anticipated, as well as the opening up of femtosecond time-resolved diffraction structural studies. At SR sources, a very high-throughput assessment for the best crystal samples and the ability to tackle just a few micron and sub-micron crystals will become widespread. With higher speeds and larger detectors, diffraction data volumes are becoming long-term storage and archiving issues; the implications for today and the future are discussed. Together with the rise of the storage ring to its current pre-eminence in MX data provision, the growing tendency of central facility sites to offer other centralized facilities complementary to crystallography, such as cryo-electron microscopy and NMR, is a welcome development.

  2. A Beam line for Macromolecular Crystallography in ALBA

    SciTech Connect

    Juanhuix, Jordi; Ferrer, Salvador

    2007-01-19

    ALBA is a third generation 3 GeV storage ring being built near Barcelona and foreseen to be operational in 2010. Out of the seven beamlines already funded in ALBA, one will be dedicated to macromolecular crystallography (MX). The beamline, dubbed XALOC, shall cope with a broad range of crystal structures and sizes. To this aim, a flexible optical design involving variable focusing optics has been incorporated into the beamline optics. The photon source will be a 2 m long, in-vacuum undulator with a period of 21.3 mm. The optics will consist in a Si(111), double-crystal monochromator cryogenically cooled, and a pair of mirrors placed in a Kirkpatrick-Baez configuration. The beamline will deliver a high flux beam in the 5-15 keV energy range, with an energy resolution of {delta}E/E {approx}2 x 10-4. In addition to the main beamline, it is being considered the possibility to use a diamond laue monochromator to provide photons at a fixed wavelength to an ancillary branch. This report shows the present status of the beamline design.

  3. JBlulce Data Acquisition Software for Macromolecular Crystallography

    SciTech Connect

    2010-06-01

    JBlulce (Java Beam Line Universal Integrated Configuration Environment is a data acquisition software for macromolecular crystallography conforming user interface of the SSRL Blulce that has become a de-factor standard in the field. Besides this interface conformity, JBlulce is a unique system in terms of architecture, speec, capability and osftware implementation. It features only two software layers, the JBlulce clients and the EPICS servers, as compared to three layers present in Blulc and most of similar systems. This layers reduction provides a faster communication with hardware and an easier access to advanced hardware capabilities like on-the-fly scanning. Then JBlulc clients are designed to operate in parallel with the other beamline controls which streamlines the tasks performed by staff such as beamline preparation, maitenance, audting and user assistance. Another distinction is the deployment of multiple plugins that can be written in any programming languag thus involving more staff into the development. further on, JBlulce makes use of unified motion controls allowing for easy scanning and optimizing of any beamline component. Finally, the graphic interface is implemented in Java making full use of rich Java libraries and Jave IDE for debugging. to compare, Blulce user interface is implemented with aging Tcl/tk language providing very restricted capabilities. JBlulce makes full use of the industrial power and wide drivers selection of EPICS in controlling hardware; all hardware commuication is routed via multiple EPICS servers residing on local area network. JBlulce also includes several EPICS State Notation servers aimed at making hardware communication more robust. Besides using EPICS for controlling hardware, JBlulce extensively uses EPICS databases for efficien communications between multiple instances of JBlulce clients and JBlulce pplugins that can run in parallel on different computers. All of the above makes JBlulce one of the biggest and most

  4. JBlulce Data Acquisition Software for Macromolecular Crystallography

    2010-06-01

    JBlulce (Java Beam Line Universal Integrated Configuration Environment is a data acquisition software for macromolecular crystallography conforming user interface of the SSRL Blulce that has become a de-factor standard in the field. Besides this interface conformity, JBlulce is a unique system in terms of architecture, speec, capability and osftware implementation. It features only two software layers, the JBlulce clients and the EPICS servers, as compared to three layers present in Blulc and most of similarmore » systems. This layers reduction provides a faster communication with hardware and an easier access to advanced hardware capabilities like on-the-fly scanning. Then JBlulc clients are designed to operate in parallel with the other beamline controls which streamlines the tasks performed by staff such as beamline preparation, maitenance, audting and user assistance. Another distinction is the deployment of multiple plugins that can be written in any programming languag thus involving more staff into the development. further on, JBlulce makes use of unified motion controls allowing for easy scanning and optimizing of any beamline component. Finally, the graphic interface is implemented in Java making full use of rich Java libraries and Jave IDE for debugging. to compare, Blulce user interface is implemented with aging Tcl/tk language providing very restricted capabilities. JBlulce makes full use of the industrial power and wide drivers selection of EPICS in controlling hardware; all hardware commuication is routed via multiple EPICS servers residing on local area network. JBlulce also includes several EPICS State Notation servers aimed at making hardware communication more robust. Besides using EPICS for controlling hardware, JBlulce extensively uses EPICS databases for efficien communications between multiple instances of JBlulce clients and JBlulce pplugins that can run in parallel on different computers. All of the above makes JBlulce one of the biggest

  5. Analytical solitary wave solutions of the nonlinear Kronig-Penney model in photonic structures.

    PubMed

    Kominis, Y

    2006-06-01

    A phase space method is employed for the construction of analytical solitary wave solutions of the nonlinear Kronig-Penney model in a photonic structure. This class of solutions is obtained under quite generic conditions, while the method is applicable to a large variety of systems. The location of the solutions on the spectral band gap structure as well as on the low dimensional space of system's conserved quantities is studied, and robust solitary wave propagation is shown.

  6. Macromolecular networks and intelligence in microorganisms

    PubMed Central

    Westerhoff, Hans V.; Brooks, Aaron N.; Simeonidis, Evangelos; García-Contreras, Rodolfo; He, Fei; Boogerd, Fred C.; Jackson, Victoria J.; Goncharuk, Valeri; Kolodkin, Alexey

    2014-01-01

    Living organisms persist by virtue of complex interactions among many components organized into dynamic, environment-responsive networks that span multiple scales and dimensions. Biological networks constitute a type of information and communication technology (ICT): they receive information from the outside and inside of cells, integrate and interpret this information, and then activate a response. Biological networks enable molecules within cells, and even cells themselves, to communicate with each other and their environment. We have become accustomed to associating brain activity – particularly activity of the human brain – with a phenomenon we call “intelligence.” Yet, four billion years of evolution could have selected networks with topologies and dynamics that confer traits analogous to this intelligence, even though they were outside the intercellular networks of the brain. Here, we explore how macromolecular networks in microbes confer intelligent characteristics, such as memory, anticipation, adaptation and reflection and we review current understanding of how network organization reflects the type of intelligence required for the environments in which they were selected. We propose that, if we were to leave terms such as “human” and “brain” out of the defining features of “intelligence,” all forms of life – from microbes to humans – exhibit some or all characteristics consistent with “intelligence.” We then review advances in genome-wide data production and analysis, especially in microbes, that provide a lens into microbial intelligence and propose how the insights derived from quantitatively characterizing biomolecular networks may enable synthetic biologists to create intelligent molecular networks for biotechnology, possibly generating new forms of intelligence, first in silico and then in vivo. PMID:25101076

  7. Macromolecular Topography Leaps into the Digital Age

    NASA Technical Reports Server (NTRS)

    Lovelace, J.; Bellamy, H.; Snell, E. H.; Borgstahl, G.

    2003-01-01

    A low-cost, real-time digital topography system is under development which will replace x-ray film and nuclear emulsion plates. The imaging system is based on an inexpensive surveillance camera that offers a 1000x1000 array of 8 im square pixels, anti-blooming circuitry, and very quick read out. Currently, the system directly converts x-rays to an image with no phosphor. The system is small and light and can be easily adapted to work with other crystallographic equipment. Preliminary images have been acquired of cubic insulin at the NSLS x26c beam line. NSLS x26c was configured for unfocused monochromatic radiation. Six reflections were collected with stills spaced from 0.002 to 0.001 degrees apart across the entire oscillation range that the reflections were in diffracting condition. All of the reflections were rotated to the vertical to reduce Lorentz and beam related effects. This particular CCD is designed for short exposure applications (much less than 1 sec) and so has a relatively high dark current leading to noisy raw images. The images are processed to remove background and other system noise with a multi-step approach including the use of wavelets, histogram, and mean window filtering. After processing, animations were constructed with the corresponding reflection profile to show the diffraction of the crystal volume vs. the oscillation angle as well as composite images showing the parts of the crystal with the strongest diffraction for each reflection. The final goal is to correlate features seen in reflection profiles captured with fine phi slicing to those seen in the topography images. With this development macromolecular topography finally comes into the digital age.

  8. Macromolecular networks and intelligence in microorganisms.

    PubMed

    Westerhoff, Hans V; Brooks, Aaron N; Simeonidis, Evangelos; García-Contreras, Rodolfo; He, Fei; Boogerd, Fred C; Jackson, Victoria J; Goncharuk, Valeri; Kolodkin, Alexey

    2014-01-01

    Living organisms persist by virtue of complex interactions among many components organized into dynamic, environment-responsive networks that span multiple scales and dimensions. Biological networks constitute a type of information and communication technology (ICT): they receive information from the outside and inside of cells, integrate and interpret this information, and then activate a response. Biological networks enable molecules within cells, and even cells themselves, to communicate with each other and their environment. We have become accustomed to associating brain activity - particularly activity of the human brain - with a phenomenon we call "intelligence." Yet, four billion years of evolution could have selected networks with topologies and dynamics that confer traits analogous to this intelligence, even though they were outside the intercellular networks of the brain. Here, we explore how macromolecular networks in microbes confer intelligent characteristics, such as memory, anticipation, adaptation and reflection and we review current understanding of how network organization reflects the type of intelligence required for the environments in which they were selected. We propose that, if we were to leave terms such as "human" and "brain" out of the defining features of "intelligence," all forms of life - from microbes to humans - exhibit some or all characteristics consistent with "intelligence." We then review advances in genome-wide data production and analysis, especially in microbes, that provide a lens into microbial intelligence and propose how the insights derived from quantitatively characterizing biomolecular networks may enable synthetic biologists to create intelligent molecular networks for biotechnology, possibly generating new forms of intelligence, first in silico and then in vivo.

  9. Special quasirandom structure modeling of fluorite-structured oxide solid solutions with aliovalent cation substitutions

    NASA Astrophysics Data System (ADS)

    Wolff-Goodrich, Silas; Hanken, Benjamin E.; Solomon, Jonathan M.; Asta, Mark

    2015-07-01

    The accuracy of the special quasirandom structure (SQS) approach for modeling the structure and energetics of fluorite-structured oxide solid solutions with aliovalent cation substitutions is assessed in an ionic-pair potential study of urania and ceria based systems mixed with trivalent rare-earth ions. Mixing enthalpies for SQS supercells containing 96 and 324 lattice sites were calculated using ionic pair potentials for U0.5La0.5O1.75, U0.5Y0.5O1.75, Ce0.5La0.5O1.75, Ce0.5Y0.5O1.75, and Ce0.5Gd0.5O1.75, which all have stoichiometries of pyrochlores. The SQS results were compared to benchmark values for random substitutional disorder obtained using large supercell models. The calculations show significant improvement of the mixing enthalpy for the larger 324 site SQS, which is attributed to a better description of the structural distortions, as characterized by the radial distribution functions in relaxed systems.

  10. Integrated Force Method Solution to Indeterminate Structural Mechanics Problems

    NASA Technical Reports Server (NTRS)

    Patnaik, Surya N.; Hopkins, Dale A.; Halford, Gary R.

    2004-01-01

    Strength of materials problems have been classified into determinate and indeterminate problems. Determinate analysis primarily based on the equilibrium concept is well understood. Solutions of indeterminate problems required additional compatibility conditions, and its comprehension was not exclusive. A solution to indeterminate problem is generated by manipulating the equilibrium concept, either by rewriting in the displacement variables or through the cutting and closing gap technique of the redundant force method. Compatibility improvisation has made analysis cumbersome. The authors have researched and understood the compatibility theory. Solutions can be generated with equal emphasis on the equilibrium and compatibility concepts. This technique is called the Integrated Force Method (IFM). Forces are the primary unknowns of IFM. Displacements are back-calculated from forces. IFM equations are manipulated to obtain the Dual Integrated Force Method (IFMD). Displacement is the primary variable of IFMD and force is back-calculated. The subject is introduced through response variables: force, deformation, displacement; and underlying concepts: equilibrium equation, force deformation relation, deformation displacement relation, and compatibility condition. Mechanical load, temperature variation, and support settling are equally emphasized. The basic theory is discussed. A set of examples illustrate the new concepts. IFM and IFMD based finite element methods are introduced for simple problems.

  11. Reverse engineering chemical structures from molecular descriptors : how many solutions?

    SciTech Connect

    Brown, William Michael; Martin, Shawn Bryan; Faulon, Jean-Loup Michel

    2005-06-01

    Physical, chemical and biological properties are the ultimate information of interest for chemical compounds. Molecular descriptors that map structural information to activities and properties are obvious candidates for information sharing. In this paper, we consider the feasibility of using molecular descriptors to safely exchange chemical information in such a way that the original chemical structures cannot be reverse engineered. To investigate the safety of sharing such descriptors, we compute the degeneracy (the number of structure matching a descriptor value) of several 2D descriptors, and use various methods to search for and reverse engineer structures. We examine degeneracy in the entire chemical space taking descriptors values from the alkane isomer series and the PubChem database. We further use a stochastic search to retrieve structures matching specific topological index values. Finally, we investigate the safety of exchanging of fragmental descriptors using deterministic enumeration.

  12. NMR solution structure of the neurotrypsin Kringle domain.

    PubMed

    Ozhogina, Olga A; Grishaev, Alexander; Bominaar, Emile L; Patthy, László; Trexler, Maria; Llinás, Miguel

    2008-11-25

    Neurotrypsin is a multidomain protein that serves as a brain-specific serine protease. Here we report the NMR structure of its kringle domain, NT/K. The data analysis was performed with the BACUS (Bayesian analysis of coupled unassigned spins) algorithm. This study presents the first application of BACUS to the structure determination of a 13C unenriched protein for which no prior experimental 3D structure was available. NT/K adopts the kringle fold, consisting of an antiparallel beta-sheet bridged by an overlapping pair of disulfides. The structure reveals the presence of a surface-exposed left-handed polyproline II helix that is closely packed to the core beta-structure. This feature distinguishes NT/K from other members of the kringle fold and points toward a novel functional role for a kringle domain. Functional divergence among kringle domains is discussed on the basis of their surface and electrostatic characteristics. PMID:18956887

  13. Growing market acceptance for fiber optic solutions in civil structures

    NASA Astrophysics Data System (ADS)

    Graver, Thomas; Inaudi, Daniele; Doornink, Justin

    2004-12-01

    Owners must manage and ensure the safety of their civil structures even as use of many structures extends well beyond their design lifetime. Traditionally, most structures rely on strict maintenance procedures, visual inspections, and very few sensors. But maintenance is very expensive, visual inspections can miss critical problems, and conventional sensors can fail in harsh environments. Can fiber-optic sensing (FOS) address these issues? This is not a new question, but there are some new answers. This paper highlights several structures where FOS is used, and describes the associated successes and challenges for each application. Many successes are coupled to improved FOS tools: better sensor packages, simpler and less expensive instrumentation, improved installation techniques, and more efficient data analysis tools. Examples of each are provided. Particular attention is given to the economics of instrumenting civil structures - when and how it pays. Conclusions include recommendations for future developments that will further accelerate FOS acceptance and use.

  14. Remote Access to the PXRR Macromolecular Crystallography Facilities at the NSLS

    SciTech Connect

    Soares, A.S.; Schneider, D. K.; Skinner, J. M.; Cowan, M.; Buono, R.; Robinson, H. H.; Heroux, A.; Carlucci-Dayton, M.; Saxena, A.; Sweet, R. M.

    2008-09-01

    The most recent surge of innovations that have simplified and streamlined the process of determining macromolecular structures by crystallography owes much to the efforts of the structural genomics community. However, this was only the last step in a long evolution that saw the metamorphosis of crystallography from an heroic effort that involved years of dedication and skill into a straightforward measurement that is occasionally almost trivial. Many of the steps in this remarkable odyssey involved reducing the physical labor that is demanded of experimenters in the field. Other steps reduced the technical expertise required for conducting those experiments.

  15. Remote Access to the PXRR Macromolecular Crystallography Facilities at the NSLS

    SciTech Connect

    A Soares; D Schneider; J Skinner; M Cowan; R Buono; H Robinson; A Heroux; M Carlucci-Dayton; A Saxena; R Sweet

    2011-12-31

    The most recent surge of innovations that have simplified and streamlined the process of determining macromolecular structures by crystallography owes much to the efforts of the structural genomics community. However, this was only the last step in a long evolution that saw the metamorphosis of crystallography from an heroic effort that involved years of dedication and skill into a straightforward measurement that is occasionally almost trivial. Many of the steps in this remarkable odyssey involved reducing the physical labor that is demanded of experimenters in the field. Other steps reduced the technical expertise required for conducting those experiments.

  16. Structure and symmetry of crystalline solid solutions: general revision

    SciTech Connect

    Vaida, M.; Shimon, L.J.W.; Weisinger-Lewin, Y.; Frolow, F.; Lahav, M.; Leiserowitz, L.; McMullan, R.K.

    1988-09-16

    Mixed single crystals composed of host and guest organic molecules of similar structure and shapes are shown to comprise sectors with different host-guest distributions and to have symmetries lower than that of the host crystal. These properties are determined by the structure of the guest and the surface structures of the crystal faces through which the guest molecules are occluded. This general concept is illustrated by studies of three mixed crystal systems, (E)-cinnamamide-(E)-2-thienylacrylamide, (E)-cinnamamide-(E)-3-thienylacrylamide, and (S)-asparagine-(S)-aspartic acid, with x-ray and neutron diffraction and solid-state photochemistry. 19 references, 4 figures, 2 tables.

  17. MUFOLD: A new solution for protein 3D structure prediction

    PubMed Central

    Zhang, Jingfen; Wang, Qingguo; Barz, Bogdan; He, Zhiquan; Kosztin, Ioan; Shang, Yi; Xu, Dong

    2010-01-01

    There have been steady improvements in protein structure prediction during the past 2 decades. However, current methods are still far from consistently predicting structural models accurately with computing power accessible to common users. Toward achieving more accurate and efficient structure prediction, we developed a number of novel methods and integrated them into a software package, MUFOLD. First, a systematic protocol was developed to identify useful templates and fragments from Protein Data Bank for a given target protein. Then, an efficient process was applied for iterative coarse-grain model generation and evaluation at the Cα or backbone level. In this process, we construct models using interresidue spatial restraints derived from alignments by multidimensional scaling, evaluate and select models through clustering and static scoring functions, and iteratively improve the selected models by integrating spatial restraints and previous models. Finally, the full-atom models were evaluated using molecular dynamics simulations based on structural changes under simulated heating. We have continuously improved the performance of MUFOLD by using a benchmark of 200 proteins from the Astral database, where no template with >25% sequence identity to any target protein is included. The average root-mean-square deviation of the best models from the native structures is 4.28 Å, which shows significant and systematic improvement over our previous methods. The computing time of MUFOLD is much shorter than many other tools, such as Rosetta. MUFOLD demonstrated some success in the 2008 community-wide experiment for protein structure prediction CASP8. PMID:19927325

  18. JBluIce–EPICS control system for macromolecular crystallography

    PubMed Central

    Stepanov, Sergey; Makarov, Oleg; Hilgart, Mark; Pothineni, Sudhir Babu; Urakhchin, Alex; Devarapalli, Satish; Yoder, Derek; Becker, Michael; Ogata, Craig; Sanishvili, Ruslan; Venugopalan, Nagarajan; Smith, Janet L.; Fischetti, Robert F.

    2011-01-01

    The trio of macromolecular crystallography beamlines constructed by the General Medicine and Cancer Institutes Collaborative Access Team (GM/CA-CAT) in Sector 23 of the Advanced Photon Source (APS) have been in growing demand owing to their outstanding beam quality and capacity to measure data from crystals of only a few micrometres in size. To take full advantage of the state-of-the-art mechanical and optical design of these beamlines, a significant effort has been devoted to designing fast, convenient, intuitive and robust beamline controls that could easily accommodate new beamline developments. The GM/CA-CAT beamline controls are based on the power of EPICS for distributed hardware control, the rich Java graphical user interface of Eclipse RCP and the task-oriented philosophy as well as the look and feel of the successful SSRL BluIce graphical user interface for crystallography. These beamline controls feature a minimum number of software layers, the wide use of plug-ins that can be written in any language and unified motion controls that allow on-the-fly scanning and optimization of any beamline com­ponent. This paper describes the ways in which BluIce was combined with EPICS and converted into the Java-based JBluIce, discusses the solutions aimed at streamlining and speeding up operations and gives an overview of the tools that are provided by this new open-source control system for facilitating crystallo­graphic experiments, especially in the field of microcrystallo­graphy. PMID:21358048

  19. Solution structures of europium(III) complexes of ethylenediaminetetraacetic acid

    SciTech Connect

    Latva, M.; Kankara, J.; Haapakka, K.

    1996-04-01

    Coordination of ethylenediaminetetraacetic acid (EDTA) with europium(III) has been studied at different concentrations in solution using {sup 7}F{sub 0}{yields}{sup 5}D{sub 0} excitation spectroscopy and excited-state lifetime measurements. EDTA forms with Eu(III) ion three different species in equimolar solutions at room temperature. At low pH values EuEDTAH is formed and at higher pH values than 1.5 two EuEDTA{sup -} complexes, which differ from each other with one water molecule in the first coordination sphere of the Eu(III) ion, total coordination number and coordination geometry, are also formed. When the concentration of EDTA is higher than the concentration of Eu(III), an EuEDTA(EDTAH){sup 4-} species where the second EDTA is weakly coordinated to EuEDTA{sup -}, is formed. If the concentration of Eu(III) ion is higher than EDTA, the extra Eu(III) ions associate with EuEDTA{sup -} and link to one of the carboxylate groups of EDTA thus causing a shortening of the excited-state lifetime of the EuEDTA{sup -} complex.

  20. Polysaccharide-based micro/nanohydrogels for delivering macromolecular therapeutics.

    PubMed

    Ganguly, Kuntal; Chaturvedi, Kiran; More, Uttam A; Nadagouda, Mallikarjuna N; Aminabhavi, Tejraj M

    2014-11-10

    Increased interest in developing novel micro/nanohydrogel based formulations for delivering macromolecular therapeutics has led to multiple choices of biodegradable and biocompatible natural polymers. This interest is largely due to the availability of large number of highly pure recombinant proteins and peptides with tunable properties as well as RNA interference technology that are used in treating some of the deadly diseases that were difficult to be treated by the conventional approaches. The majority of marketed drugs that are now available are in the form of injectables that pose limited patient compliance and convenience. On the other hand, micro/nanotechnology based macromolecular delivery formulations offer many alternative routes of administration and advantages with improved patient compliance and efficient or targeted delivery of intracellular therapeutics to the site of action. This review outlines and critically evaluates the research findings on micro and nano-carrier polymeric hydrogels for the delivery of macromolecular therapeutics.

  1. Finite element solution of transient fluid-structure interaction problems

    NASA Technical Reports Server (NTRS)

    Everstine, Gordon C.; Cheng, Raymond S.; Hambric, Stephen A.

    1991-01-01

    A finite element approach using NASTRAN is developed for solving time-dependent fluid-structure interaction problems, with emphasis on the transient scattering of acoustic waves from submerged elastic structures. Finite elements are used for modeling both structure and fluid domains to facilitate the graphical display of the wave motion through both media. For the liquid, the use of velocity potential as the fundamental unknown results in a symmetric matrix equation. The approach is illustrated for the problem of transient scattering from a submerged elastic spherical shell subjected to an incident tone burst. The use of an analogy between the equations of elasticity and the wave equation of acoustics, a necessary ingredient to the procedure, is summarized.

  2. Solutions for Missing Data in Structural Equation Modeling

    ERIC Educational Resources Information Center

    Carter, Rufus Lynn

    2006-01-01

    Many times in both educational and social science research it is impossible to collect data that is complete. When administering a survey, for example, people may answer some questions and not others. This missing data causes a problem for researchers using structural equation modeling (SEM) techniques for data analyses. Because SEM and…

  3. Element-by-element Solution Procedures for Nonlinear Structural Analysis

    NASA Technical Reports Server (NTRS)

    Hughes, T. J. R.; Winget, J. M.; Levit, I.

    1984-01-01

    Element-by-element approximate factorization procedures are proposed for solving the large finite element equation systems which arise in nonlinear structural mechanics. Architectural and data base advantages of the present algorithms over traditional direct elimination schemes are noted. Results of calculations suggest considerable potential for the methods described.

  4. Macromolecular crystallography beamline X25 at the NSLS

    PubMed Central

    Héroux, Annie; Allaire, Marc; Buono, Richard; Cowan, Matthew L.; Dvorak, Joseph; Flaks, Leon; LaMarra, Steven; Myers, Stuart F.; Orville, Allen M.; Robinson, Howard H.; Roessler, Christian G.; Schneider, Dieter K.; Shea-McCarthy, Grace; Skinner, John M.; Skinner, Michael; Soares, Alexei S.; Sweet, Robert M.; Berman, Lonny E.

    2014-01-01

    Beamline X25 at the NSLS is one of the five beamlines dedicated to macromolecular crystallography operated by the Brookhaven National Laboratory Macromolecular Crystallography Research Resource group. This mini-gap insertion-device beamline has seen constant upgrades for the last seven years in order to achieve mini-beam capability down to 20 µm × 20 µm. All major components beginning with the radiation source, and continuing along the beamline and its experimental hutch, have changed to produce a state-of-the-art facility for the scientific community. PMID:24763654

  5. Macromolecular crystallography beamline X25 at the NSLS.

    PubMed

    Héroux, Annie; Allaire, Marc; Buono, Richard; Cowan, Matthew L; Dvorak, Joseph; Flaks, Leon; Lamarra, Steven; Myers, Stuart F; Orville, Allen M; Robinson, Howard H; Roessler, Christian G; Schneider, Dieter K; Shea-McCarthy, Grace; Skinner, John M; Skinner, Michael; Soares, Alexei S; Sweet, Robert M; Berman, Lonny E

    2014-05-01

    Beamline X25 at the NSLS is one of the five beamlines dedicated to macromolecular crystallography operated by the Brookhaven National Laboratory Macromolecular Crystallography Research Resource group. This mini-gap insertion-device beamline has seen constant upgrades for the last seven years in order to achieve mini-beam capability down to 20 µm × 20 µm. All major components beginning with the radiation source, and continuing along the beamline and its experimental hutch, have changed to produce a state-of-the-art facility for the scientific community.

  6. A macromolecular prodrug strategy for combinatorial drug delivery.

    PubMed

    Li, Nan-Nan; Lin, Jiantao; Gao, Di; Zhang, Li-Ming

    2014-03-01

    A novel macromolecular prodrug strategy was developed for the combinatorial delivery of two poorly water-soluble drugs, dexamethasone and doxorubicin. In this work, dexamethasone was firstly conjugated onto a water-soluble modified polysaccharide by an acid-labile hydrazone linkage. The resultant macromolecular prodrug had an amphiphilic character and could self-assemble into spherical polymeric micelles in aqueous system. With these micelles, doxorubicin was then encapsulated into their hydrophobic cores. For the conjugated dexamethasone and encapsulated doxorubicin, they could exhibit independent and acid-sensitive release characteristics. For the doxorubicin-loaded prodrug micelles, they were easily be internalized by living cells and showed obvious antitumor activity. PMID:24407691

  7. Effect of water structure on adsorption of thermosensitive polymer hydrogel in salt solutions

    SciTech Connect

    Nakano, Yoshio; Seida, Yoshimi

    1996-12-31

    Effects of temperature and additive salt on an adsorption property of thermosensitive polymer hydrogel were studied in terms of (1) phase transition temperature of the gel, (2) hydration structure of network of the gel in the various salt solutions, and (3) structure of water in the solutions. The adsorption properties of the gel were correlated fairly well with the phase transition temperature of the gel and the structure of water both on the network of the gel and in the bulk solution. 2 refs., 5 figs.

  8. Davisson-Germer Prize in Atomic or Surface Physics Lecture: Line 'Em All Up: Macromolecular Assembly at Liquid Interfaces

    NASA Astrophysics Data System (ADS)

    Richmond, Geraldine

    2013-03-01

    Advances in our molecular level understanding of the ubiquitous fluid interface comprised of a hydrophobic fluid medium, and an aqueous solution of soluble ions and solutes has been slow until recently. This more recent upsurge in interest and progress comes from advances in both experimental and computational techniques as well as the increasingly important role that this interface is playing in such areas as green chemistry, nanoparticle synthesis, improved oil and mineral recovery and water purification. The presentation will focus on our most recent efforts in understanding (1) the molecular structure of the interface between two immiscible liquids, (2) the penetration of aqueous phase ions into the interfacial region and their effect on its properties, and (3) the structure and dynamics of the adsorption of surfactants, polymers and nanoparticles at this interface. To gain insights into these processes we use a combination of vibrational sum frequency spectroscopy, surface tension measurements using the pendant drop method, and molecular dynamics simulations. The results demonstrate that weak interactions between interfacial oil and water molecules create an interface that exhibits a high degree of molecular structuring and ordering, and with properties quite different than what is observed at the air-water interface. As a consequence of these interfacial oil-water interactions, the interface provides a unique environment for the adsorption and assembly of ions, polymers and nanoparticles that are drawn to its inner-most regions. Examples of our studies that provide new insights into the unique nature of adsorption, adsorption dynamics and macromolecular assembly at this interface will be provided.

  9. Macromolecular Crowding Studies of Amino Acids Using NMR Diffusion Measurements and Molecular Dynamics Simulations

    NASA Astrophysics Data System (ADS)

    Virk, Amninder; Stait-Gardner, Timothy; Willis, Scott; Torres, Allan; Price, William

    2015-02-01

    Molecular crowding occurs when the total concentration of macromolecular species in a solution is so high that a considerable proportion of the volume is physically occupied and therefore not accessible to other molecules. This results in significant changes in the solution properties of the molecules in such systems. Macromolecular crowding is ubiquitous in biological systems due to the generally high intracellular protein concentrations. The major hindrance to understanding crowding is the lack of direct comparison of experimental data with theoretical or simulated data. Self-diffusion is sensitive to changes in the molecular weight and shape of the diffusing species, and the available diffusion space (i.e., diffusive obstruction). Consequently, diffusion measurements are a direct means for probing crowded systems including the self-association of molecules. In this work, nuclear magnetic resonance measurements of the self-diffusion of four amino acids (glycine, alanine, valine and phenylalanine) up to their solubility limit in water were compared directly with molecular dynamics simulations. The experimental data were then analyzed using various models of aggregation and obstruction. Both experimental and simulated data revealed that the diffusion of both water and the amino acids were sensitive to the amino acid concentration. The direct comparison of the simulated and experimental data afforded greater insights into the aggregation and obstruction properties of each amino acid.

  10. An Efficient Low Storage and Memory Treatment of Gridded Interaction Fields for Simulations of Macromolecular Association.

    PubMed

    Ozboyaci, Musa; Martinez, Michael; Wade, Rebecca C

    2016-09-13

    Computer simulations of molecular systems often make use of regular rectangular grids with equidistant spacing to store information on their molecular interaction fields, e.g., electrostatic potential. These grids provide an easy way to store the data as they do not require any particular specification of the structure of the data. However, such grids may easily become large, and the storage and memory demands may become so high that calculations become infeasible. To overcome this problem, we show here how the data structure DT-Grid can be adapted and applied to efficiently represent macromolecular interaction grids by exploiting the nonuniformity of information on the grid; at the same time, this data structure ensures fast random data access. We demonstrate use of the DT-Grid data structure for potential of mean force and Brownian dynamics simulations of protein-surface binding and macromolecular association with the SDA software. We further demonstrate that the DT-Grid structure enables systems of large size, such as a viral capsid, and high resolution grids to be handled that are beyond current computational feasibility. PMID:27463233

  11. EDM-DEDM and protein crystal structure solution.

    PubMed

    Caliandro, Rocco; Carrozzini, Benedetta; Cascarano, Giovanni Luca; Giacovazzo, Carmelo; Mazzone, Anna Maria; Siliqi, Dritan

    2009-05-01

    Electron-density modification (EDM) procedures are the classical tool for driving model phases closer to those of the target structure. They are often combined with automated model-building programs to provide a correct protein model. The task is not always performed, mostly because of the large initial phase error. A recently proposed procedure combined EDM with DEDM (difference electron-density modification); the method was applied to the refinement of phases obtained by molecular replacement, ab initio or SAD phasing [Caliandro, Carrozzini, Cascarano, Giacovazzo, Mazzone & Siliqi (2009), Acta Cryst. D65, 249-256] and was more effective in improving phases than EDM alone. In this paper, a novel fully automated protocol for protein structure refinement based on the iterative application of automated model-building programs combined with the additional power derived from the EDM-DEDM algorithm is presented. The cyclic procedure was successfully tested on challenging cases for which all other approaches had failed.

  12. Assembly and solution structure of the core retromer protein complex.

    PubMed

    Norwood, Suzanne J; Shaw, Daniel J; Cowieson, Nathan P; Owen, David J; Teasdale, Rohan D; Collins, Brett M

    2011-01-01

    Retromer is a peripheral membrane protein complex that has pleiotropic roles in endosomal membrane trafficking. The core of retromer possesses three subunits, VPS35, VPS29 and VPS26, that play different roles in binding to cargo, regulatory proteins and complex stabilization. We have performed an investigation of the thermodynamics of core retromer assembly using isothermal titration calorimetry (ITC) demonstrating that VPS35 acts as the central subunit to which VPS29 and VPS26 bind independently. Furthermore, we confirm that the conserved PRLYL motif of the large VPS35 subunit is critical for direct VPS26 interaction. Heat capacity measurements of VPS29 and VPS26 binding to VPS35 indicate extensive binding interfaces and suggest conformational alterations in VPS29 or VPS35 upon complex formation. Solution studies of the retromer core using small-angle X-ray scattering allow us to propose a model whereby VPS35 forms an extended platform with VPS29 and VPS26 bound at distal ends, with the potential for forming dimeric assemblies. PMID:20875039

  13. ANOVA-HDMR structure of the higher order nodal diffusion solution

    SciTech Connect

    Bokov, P. M.; Prinsloo, R. H.; Tomasevic, D. I.

    2013-07-01

    Nodal diffusion methods still represent a standard in global reactor calculations, but employ some ad-hoc approximations (such as the quadratic leakage approximation) which limit their accuracy in cases where reference quality solutions are sought. In this work we solve the nodal diffusion equations utilizing the so-called higher-order nodal methods to generate reference quality solutions and to decompose the obtained solutions via a technique known as High Dimensional Model Representation (HDMR). This representation and associated decomposition of the solution provides a new formulation of the transverse leakage term. The HDMR structure is investigated via the technique of Analysis of Variance (ANOVA), which indicates why the existing class of transversely-integrated nodal methods prove to be so successful. Furthermore, the analysis leads to a potential solution method for generating reference quality solutions at a much reduced calculational cost, by applying the ANOVA technique to the full higher order solution. (authors)

  14. The solution structure of an anti-CRISPR protein

    PubMed Central

    Maxwell, Karen L.; Garcia, Bianca; Bondy-Denomy, Joseph; Bona, Diane; Hidalgo-Reyes, Yurima; Davidson, Alan R.

    2016-01-01

    Bacterial CRISPR–Cas adaptive immune systems use small guide RNAs to protect against phage infection and invasion by foreign genetic elements. We previously demonstrated that a group of Pseudomonas aeruginosa phages encode anti-CRISPR proteins that inactivate the type I-F and I-E CRISPR–Cas systems using distinct mechanisms. Here, we present the three-dimensional structure of an anti-CRISPR protein and map a functional surface that is critical for its potent inhibitory activity. The interaction of the anti-CRISPR protein with the CRISPR–Cas complex through this functional surface is proposed to prevent the binding of target DNA. PMID:27725669

  15. Simulation of macromolecular liquids with the adaptive resolution molecular dynamics technique

    NASA Astrophysics Data System (ADS)

    Peters, J. H.; Klein, R.; Delle Site, L.

    2016-08-01

    We extend the application of the adaptive resolution technique (AdResS) to liquid systems composed of alkane chains of different lengths. The aim of the study is to develop and test the modifications of AdResS required in order to handle the change of representation of large molecules. The robustness of the approach is shown by calculating several relevant structural properties and comparing them with the results of full atomistic simulations. The extended scheme represents a robust prototype for the simulation of macromolecular systems of interest in several fields, from material science to biophysics.

  16. Simulation of macromolecular liquids with the adaptive resolution molecular dynamics technique.

    PubMed

    Peters, J H; Klein, R; Delle Site, L

    2016-08-01

    We extend the application of the adaptive resolution technique (AdResS) to liquid systems composed of alkane chains of different lengths. The aim of the study is to develop and test the modifications of AdResS required in order to handle the change of representation of large molecules. The robustness of the approach is shown by calculating several relevant structural properties and comparing them with the results of full atomistic simulations. The extended scheme represents a robust prototype for the simulation of macromolecular systems of interest in several fields, from material science to biophysics. PMID:27627414

  17. Phase Behavior of a Single Structured Ionomer Chain in Solution

    DOE PAGES

    Aryal, Dipak; Etampawala, Thusitha; Perahia, Dvora; Grest, Gary S.

    2014-08-14

    Structured polymers offer a means to tailor transport pathways within mechanically stable manifolds. Here we examine the building block of such a membrane, namely a single large pentablock co-polymer that consist of a center block of a randomly sulfonated polystyrene, designed for transport, tethered to poly-ethylene-r-propylene and end-capped by poly-t-butyl styrene, for mechanical stability,using molecular dynamics simulations. The polymer structure in a cyclohexane-heptane mixture, a technologically viable solvent, and in water, a poor solvent for all segments and a ubiquitous substance is extracted. In all solvents the pentablock collapsed into nearly spherical aggregates where the ionic block is segregated. Inmore » hydrophobic solvents, the ionic block resides in the center, surrounded by swollen intermix of flexible and end blocks. In water all blocks are collapsed with the sulfonated block residing on the surface. Our results demonstrate that solvents drive different local nano-segregation, providing a gateway to assemble membranes with controlled topology.« less

  18. Phase Behavior of a Single Structured Ionomer Chain in Solution

    SciTech Connect

    Aryal, Dipak; Etampawala, Thusitha; Perahia, Dvora; Grest, Gary S.

    2014-08-14

    Structured polymers offer a means to tailor transport pathways within mechanically stable manifolds. Here we examine the building block of such a membrane, namely a single large pentablock co-polymer that consist of a center block of a randomly sulfonated polystyrene, designed for transport, tethered to poly-ethylene-r-propylene and end-capped by poly-t-butyl styrene, for mechanical stability,using molecular dynamics simulations. The polymer structure in a cyclohexane-heptane mixture, a technologically viable solvent, and in water, a poor solvent for all segments and a ubiquitous substance is extracted. In all solvents the pentablock collapsed into nearly spherical aggregates where the ionic block is segregated. In hydrophobic solvents, the ionic block resides in the center, surrounded by swollen intermix of flexible and end blocks. In water all blocks are collapsed with the sulfonated block residing on the surface. Our results demonstrate that solvents drive different local nano-segregation, providing a gateway to assemble membranes with controlled topology.

  19. Structural Order of Water Molecules around Hydrophobic Solutes: Length-Scale Dependence and Solute-Solvent Coupling.

    PubMed

    Hande, Vrushali R; Chakrabarty, Suman

    2015-08-27

    It has been suggested that the structure and thermodynamics of the water molecules in the hydration layer of simple hydrophobic solutes undergo an order-disorder transition around a nanometer length-scale of the solute size. Using extensive atomistic molecular dynamics (MD) and replica exchange molecular dynamics (REMD) simulation studies, we have probed this order-disorder transition around model hydrophobic solutes of varying size and shape (spherical, planar, and linear), as well as flexible hydrophobic homopolymer chains (n-alkanes), where the conformational fluctuations are likely to create both spatial and temporal heterogeneity on the solvent accessible surface. We have explored the structural response of the water molecules in the hydration shell due to the local variations of the length-scale (or curvature) upon hydrophobic collapse and/or local conformational changes of these polymers. We have shown that the tetrahedral order of the water molecules in the hydration shell is practically independent of the polymer size in the extended state of the polymer due to the availability of a subnanometer cross-sectional length-scale, allowing the water molecules to form hydrogen bonds around the polymer chain. Beyond a certain length of the polymer chains, the collapsed states (associated with larger solute length-scale) start to induce disorder in the surface water molecules. We demonstrate that the local structure (both local number density and tetrahedral order) of the hydration layer is dynamically coupled to the local topology of the polymer. Thus, we envisage that in a flexible (bio)polymer, the hydration shell properties will be sensitive to the local conformational state of the molecule (both spatially and temporally), and the overall observed water structure and dynamics will be dependent on the topological/chemical heterogeneity, and the time-scale of fluctuations in the local curvature (length-scale) of the solvent accessible surface. Moreover, we have

  20. Molecular structures and protonation state of 2-Mercaptopyridine in aqueous solution

    NASA Astrophysics Data System (ADS)

    Eckert, S.; Miedema, P. S.; Quevedo, W.; O'Cinneide, B.; Fondell, M.; Beye, M.; Pietzsch, A.; Ross, M.; Khalil, M.; Föhlisch, A.

    2016-03-01

    The speciation of 2-Mercaptopyridine in aqueous solution has been investigated with nitrogen 1s Near Edge X-ray Absorption Fine Structure spectroscopy and time dependent Density Functional Theory. The prevalence of distinct species as a function of the solvent basicity is established. No indications of dimerization towards high concentrations are found. The determination of different molecular structures of 2-Mercaptopyridine in aqueous solution is put into the context of proton-transfer in keto-enol and thione-thiol tautomerisms.

  1. A Compact X-Ray System for Macromolecular Crystallography

    NASA Technical Reports Server (NTRS)

    Gubarev, Mikhail; Ciszak, Ewa; Ponomarev, Igor; Gibson, Walter; Joy, Marshall

    2000-01-01

    We describe the design and performance of a high flux x-ray system for a macromolecular crystallography that combines a microfocus x-ray generator (40 micrometer full width at half maximum spot size at a power level of 46.5 W) and a collimating polycapillary optic. The Cu Ka lpha x-ray flux produced by this optimized system through a 500,um diam orifice is 7.0 times greater than the x-ray flux previously reported by Gubarev et al. [M. Gubarev et al., J. Appl. Crystallogr. 33, 882 (2000)]. The x-ray flux from the microfocus system is also 2.6 times higher than that produced by a rotating anode generator equipped with a graded multilayer monochromator (green optic, Osmic Inc. CMF24-48-Cu6) and 40% less than that produced by a rotating anode generator with the newest design of graded multilayer monochromator (blue optic, Osmic, Inc. CMF12-38-Cu6). Both rotating anode generators operate at a power level of 5000 W, dissipating more than 100 times the power of our microfocus x-ray system. Diffraction data collected from small test crystals are of high quality. For example, 42 540 reflections collected at ambient temperature from a lysozyme crystal yielded R(sub sym)=5.0% for data extending to 1.70 A, and 4.8% for the complete set of data to 1.85 A. The amplitudes of the observed reflections were used to calculate difference electron density maps that revealed positions of structurally important ions and water molecules in the crystal of lysozyme using the phases calculated from the protein model.

  2. A Compact X-Ray System for Macromolecular Crystallography. 5

    NASA Technical Reports Server (NTRS)

    Gubarev, Mikhail; Ciszak, Ewa; Ponomarev, Igor; Joy, Marshall

    2000-01-01

    We describe the design and performance of a high flux x-ray system for macromolecular crystallography that combines a microfocus x-ray generator (40 gm FWHM spot size at a power level of 46.5Watts) and a 5.5 mm focal distance polycapillary optic. The Cu K(sub alpha) X-ray flux produced by this optimized system is 7.0 times above the X-ray flux previously reported. The X-ray flux from the microfocus system is also 3.2 times higher than that produced by the rotating anode generator equipped with a long focal distance graded multilayer monochromator (Green optic; CMF24-48-Cu6) and 30% less than that produced by the rotating anode generator with the newest design of graded multilayer monochromator (Blue optic; CMF12-38-Cu6). Both rotating anode generators operate at a power level of 5000 Watts, dissipating more than 100 times the power of our microfocus x-ray system. Diffraction data collected from small test crystals are of high quality. For example, 42,540 reflections collected at ambient temperature from a lysozyme crystal yielded R(sub sym) 5.0% for the data extending to 1.7A, and 4.8% for the complete set of data to 1.85A. The amplitudes of the reflections were used to calculate difference electron density maps that revealed positions of structurally important ions and water molecules in the crystal of lysozyme using the phases calculated from the protein model.

  3. Minimizing distortion in truss structures - A Hopfield network solution

    NASA Technical Reports Server (NTRS)

    Fu, B.; Hajela, P.

    1992-01-01

    Distortions in truss structures can result from random errors in element lengths that are typical of a manufacturing process. These distortions may be minimized by an optimal selection of elements from those available for placement between the prescribed nodes - a combinatorial optimization problem requiring significant investment of computational resource for all but the smallest problems. The present paper describes a formulation in which near-optimal element assignments are obtained as minimum-energy stable states, of an analogous Hopfield neural network. This requires mapping of the optimization problem into an energy function of the appropriate Liapunov form. The computational architecture is ideally suited to a parallel processor implementation and offers significant savings in computational effort. A numerical implementation of the approach is discussed with reference to planar truss problems.

  4. Minimizing distortion in truss structures -- a Hopfield network solution

    NASA Technical Reports Server (NTRS)

    Fu, B.; Hajela, P.

    1993-01-01

    Distortions in truss structures can result from random errors in elemental lengths that are typical of a manufacturing process. These distortions may be minimized by an optimal selection of elements from those available for placement between the prescribed nodes -- a combinatorial optimization problem requiring significant investment of computational resource for all but the smallest problems. The present paper describes a formulation in which near-optimal element assignments are obtained as minimum energy, stable states, of an analogous Hopfield neural network. This requires mapping of the optimization problem into an energy function of the appropriate Lyapunov form. The computational architecture is ideally suited to a parallel processor implementation and offers significant savings in computational effort. A numerical implementation of the approach is discussed with reference to planar truss problems.

  5. Effects of macromolecular crowding and osmolyte on human Tau fibrillation.

    PubMed

    Wu, Yingying; Teng, Ningning; Li, Sen

    2016-09-01

    Tau fibrillation is reported to be involved in neurodegenerative disorders, such as Alzheimer's disease, in which the natural environment is very crowded in the cells. Understanding the role of crowding environments in regulating Tau fibrillation is of great importance for elucidating the etiology of these diseases. In this experiment, the effects of macromolecular crowding and osmolyte reagents in the crowding environment on Tau fibrillation were studied by thioflavin T binding, SDS-PAGE and TEM assays. Ficoll 70 and Dextran 70 of different concentrations were used as macromolecular crowding reagents inside the cells and showed a strong enhancing effect on the fibrillation of normal and hyperphosphorylated Tau. The enhancing effect of Dextran is stronger than that of Ficoll 70 at the same concentration. In addition, the cellular osmolyte sucrose was found to protect Tau against fibrillation, and inhibit the enhancing effect of macromolecular crowding on Tau fibrillation. A possible model for the fibrillation process of Tau and the effect of macromolecular crowding and osmolyte on this process was proposed based on these experimental results. The information obtained from our study can enhance the understanding of how proteins aggregate and avoid aggregation in crowded physiological environments and might lead to a better understanding of the molecular mechanisms of Alzheimer's disease in vivo.

  6. Macromolecular Pt(IV) Prodrugs from Poly(organo)phosphazenes.

    PubMed

    Henke, Helena; Kryeziu, Kushtrim; Banfić, Jelena; Theiner, Sarah; Körner, Wilfried; Brüggemann, Oliver; Berger, Walter; Keppler, Bernhard K; Heffeter, Petra; Teasdale, Ian

    2016-08-01

    The preparation of novel macromolecular prodrugs via the conjugation of two platinum(IV) complexes to suitably functionalized poly(organo)phosphazenes is presented. The inorganic/organic polymers provide carriers with controlled dimensions due to the use of living cationic polymerization and allow the preparation of conjugates with excellent aqueous solubility but long-term hydrolytic degradability. The macromolecular Pt(IV) prodrugs are designed to undergo intracellular reduction and simultaneous release from the macromolecular carrier to present the active Pt(II) drug derivatives. In vitro investigations show a significantly enhanced intracellular uptake of Pt for the macromolecular prodrugs when compared to small molecule Pt complexes, which is also reflected in an increase in cytotoxicity. Interestingly, drug-resistant sublines also show a significantly smaller resistance against the conjugates compared to clinically established platinum drugs, indicating that an alternative uptake route of the Pt(IV) conjugates might also be able to overcome acquired resistance against Pt(II) drugs. In vivo studies of a selected conjugate show improved tumor shrinkage compared to the respective Pt(IV) complex. PMID:27169668

  7. Macromolecular Pt(IV) Prodrugs from Poly(organo)phosphazenes.

    PubMed

    Henke, Helena; Kryeziu, Kushtrim; Banfić, Jelena; Theiner, Sarah; Körner, Wilfried; Brüggemann, Oliver; Berger, Walter; Keppler, Bernhard K; Heffeter, Petra; Teasdale, Ian

    2016-08-01

    The preparation of novel macromolecular prodrugs via the conjugation of two platinum(IV) complexes to suitably functionalized poly(organo)phosphazenes is presented. The inorganic/organic polymers provide carriers with controlled dimensions due to the use of living cationic polymerization and allow the preparation of conjugates with excellent aqueous solubility but long-term hydrolytic degradability. The macromolecular Pt(IV) prodrugs are designed to undergo intracellular reduction and simultaneous release from the macromolecular carrier to present the active Pt(II) drug derivatives. In vitro investigations show a significantly enhanced intracellular uptake of Pt for the macromolecular prodrugs when compared to small molecule Pt complexes, which is also reflected in an increase in cytotoxicity. Interestingly, drug-resistant sublines also show a significantly smaller resistance against the conjugates compared to clinically established platinum drugs, indicating that an alternative uptake route of the Pt(IV) conjugates might also be able to overcome acquired resistance against Pt(II) drugs. In vivo studies of a selected conjugate show improved tumor shrinkage compared to the respective Pt(IV) complex.

  8. Modeling of Carbon Nanotubes (Nanofibers) as Macromolecular Coils

    NASA Astrophysics Data System (ADS)

    Mikitaev, A. K.; Kozlov, G. V.

    2015-12-01

    Modeling of carbon nanotubes (nanofibers) in polymer nanocomposites as macromolecular coils is performed. This approach offers an estimation of the real degree of anisotropy of these nanofillers and allows predicting the properties of the resulting nanocomposites. An important role of the nanofiller - polymer matrix interface is demonstrated.

  9. Macromolecular Pt(IV) Prodrugs from Poly(organo)phosphazenes

    PubMed Central

    Banfić, Jelena; Theiner, Sarah; Körner, Wilfried; Brüggemann, Oliver; Berger, Walter; Keppler, Bernhard K.; Heffeter, Petra; Teasdale, Ian

    2016-01-01

    The preparation of novel macromolecular prodrugs via the conjugation of two platinum(IV) complexes to suitably functionalized poly(organo)phosphazenes is presented. The inorganic/organic polymers provide carriers with controlled dimensions due to the use of living cationic polymerization and allow the preparation of conjugates with excellent aqueous solubility but long-term hydrolytic degradability. The macromolecular Pt(IV) prodrugs are designed to undergo intracellular reduction and simultaneous release from the macromolecular carrier to present the active Pt(II) drug derivatives. In vitro investigations show a significantly enhanced intracellular uptake of Pt for the macromolecular prodrugs when compared to small molecule Pt complexes, which is also reflected in an increase in cytotoxicity. Interestingly, drug-resistant sublines also show a significantly smaller resistance against the conjugates compared to clinically established platinum drugs, indicating that an alternative uptake route of the Pt(IV) conjugates might also be able to overcome acquired resistance against Pt(II) drugs. In vivo studies of a selected conjugate show improved tumor shrinkage compared to the respective Pt(IV) complex. PMID:27169668

  10. Modifications in structure and interaction of nanoparticle-protein-surfactant complexes in electrolyte solution

    NASA Astrophysics Data System (ADS)

    Mehan, Sumit; Kumar, S.; Aswal, V. K.; Schweins, R.

    2016-05-01

    SANS experiments of three-component system of anionic silica nanoparticles, anionic BSA protein and anionic SDS surfactants have been carried out without and with electrolyte in aqueous solution. In both the cases, the interaction of surfactant with protein results in formation of bead-necklace structure of protein-surfactant complexes in solution. These protein-surfactant complexes interact very differently with nanoparticles in absence and presence of electrolyte. In absence of electrolyte, nanoparticles remain in dispersed phase in solution, whereas with the addition of electrolyte the nanoparticles fractal aggregates are formed. SANS describes the phase behavior to be governed by competition of electrostatic and depletion interactions among the components solution.

  11. Exploiting special problem structure when reliably investigating the solution of systems of IVPs and DDEs

    NASA Astrophysics Data System (ADS)

    Enright, W. H.

    2016-06-01

    In recent years we have developed a class of reliable order p methods for the approximate solution of general systems of initial value problems (IVPs) and delay differential equations (DDEs). In the theoretical analysis of these methods we have identified several trade-offs that do arise and have to be addressed when applying these methods to problems that exhibit special structure. Similar trade-offs also arise when one is concerned with investigating other important properties of the solutions. We will give examples of such trade-offs that arise when investigating the sensitivities of the solutions, and when very accurate approximate solutions are required.

  12. Global structure of black hole and brane solutions in a multidimensional model with anisotropic fluid

    NASA Astrophysics Data System (ADS)

    Bolokhov, S. V.; Ivashchuk, V. D.

    We analyse the global causal structure of a family of multidimensional spherically-symmetric solutions with a horizon which appear in the model with 1-component anisotropic fluid. This family can be considered as a generalized analogs of the well-known black hole solutions (including the Reissner--Nordström one) and some black brane solutions. The structure of regular horizons and singular boundaries is studied, and the corresponding Carter--Penrose diagrams are constructed for various values of the parameters of the model.

  13. Solution structure of the cAMP-dependent protein kinase

    SciTech Connect

    Trewhella, J.; Olah, G.A.; Walsh, D.A.; Mitchell, R.D.

    1998-12-31

    This is the final report of a three-year, Laboratory Directed Research and Development (LDRD) project as Los Alamos National Laboratory (LANL). Protein phosphorylation is well established as one of the most important mechanisms of signal transduction and cellular regulation. Two of the key enzymes that catalyze these phosphorylation reactions are the cAMP- (PKA) and cGMP- (PKG) dependent protein kinases. PKA has served as the prototypic model of this class of enzymes that now comprises in excess of 300 phylogenetically related proteins. A large number of these protein kinases are critical for the regulation of cell function and a full analysis of their similarities and differences is essential to understand their diverse physiological roles. The cAMP-dependent protein kinase has the subunit structure R2C2, in which C and R refer to the catalytic and regulatory subunits, respectively. The cGMP-dependent protein kinase (PKG) is highly homologous to PKA but is distinguished from it by having the regulatory and catalytic domains on a contiguous polypeptide. The studies described here use small-angle scattering and Fourier Transform InfraRed (FTIR) spectroscopy to study domain movements and conformational changes in these enzymes in different functional states in order to elucidate the molecular bases for the regulation of their activities.

  14. A structured multi-block solution-adaptive mesh algorithm with mesh quality assessment

    NASA Technical Reports Server (NTRS)

    Ingram, Clint L.; Laflin, Kelly R.; Mcrae, D. Scott

    1995-01-01

    The dynamic solution adaptive grid algorithm, DSAGA3D, is extended to automatically adapt 2-D structured multi-block grids, including adaption of the block boundaries. The extension is general, requiring only input data concerning block structure, connectivity, and boundary conditions. Imbedded grid singular points are permitted, but must be prevented from moving in space. Solutions for workshop cases 1 and 2 are obtained on multi-block grids and illustrate both increased resolution of and alignment with the solution. A mesh quality assessment criteria is proposed to determine how well a given mesh resolves and aligns with the solution obtained upon it. The criteria is used to evaluate the grid quality for solutions of workshop case 6 obtained on both static and dynamically adapted grids. The results indicate that this criteria shows promise as a means of evaluating resolution.

  15. The density, viscosity and structural properties of aqueous ethambutol hydrochloride solutions

    NASA Astrophysics Data System (ADS)

    Deosarkar, S. D.; Puyad, A. L.; Kalyankar, T. M.

    2012-05-01

    Ethambutol (EMB) is a bacteriostatic antimycobacterial drug prescribed to treat tuberculosis. It is bacteriostatic against actively growing TB bacilli. The density and viscosity of aqueous ethambutol hydrochloride solutions have been studied at 298.15, 301.15 and 304.15 K and at different concentrations (0.255, 0.168, 0.128, 0.087, 0.041, and 0.023 mol dm-3). The apparent molar volume of these solutions for different temperatures and concentrations was calculated from the density data. The relative viscosities of drug solutions have been analysed by Jones-Dole equation. The limiting apparent molar volumes have been evaluated for different temperatures. The different properties have been used to study structural properties, structure formation and breaking properties of drug and solute-solvent interactions in solutions.

  16. Investigations on the structure of DMSO and acetone in aqueous solution

    SciTech Connect

    McLain, Sylvia E; Soper, Alan K

    2007-01-01

    Aqueous solutions of dimethyl sulfoxide (DMSO) and acetone have been investigated using neutron diffraction augmented with isotopic substitution and empirical potential structure refinement computer simulations. Each solute has been measured at two concentrations-1:20 and 1:2 solute:water mole ratios. At both concentrations for each solute, the tetrahedral hydrogen bonding network of water is largely unperturbed, though the total water molecule coordination number is reduced in the higher 1:2 concentrations. With higher concentrations of acetone, water tends to segregate into clusters, while in higher concentrations of DMSO the present study reconfirms that the structure of the liquid is dominated by DMSO-water interactions. This result may have implications for the highly nonideal behavior observed in the thermodynamic functions for 1:2 DMSO-water solutions.

  17. Large-scale analysis of macromolecular crowding effects on protein aggregation using a reconstituted cell-free translation system

    PubMed Central

    Niwa, Tatsuya; Sugimoto, Ryota; Watanabe, Lisa; Nakamura, Shugo; Ueda, Takuya; Taguchi, Hideki

    2015-01-01

    Proteins must fold into their native structures in the crowded cellular environment, to perform their functions. Although such macromolecular crowding has been considered to affect the folding properties of proteins, large-scale experimental data have so far been lacking. Here, we individually translated 142 Escherichia coli cytoplasmic proteins using a reconstituted cell-free translation system in the presence of macromolecular crowding reagents (MCRs), Ficoll 70 or dextran 70, and evaluated the aggregation propensities of 142 proteins. The results showed that the MCR effects varied depending on the proteins, although the degree of these effects was modest. Statistical analyses suggested that structural parameters were involved in the effects of the MCRs. Our dataset provides a valuable resource to understand protein folding and aggregation inside cells. PMID:26500644

  18. Robust, high-throughput solution structural analyses by small angle X-ray scattering (SAXS)

    SciTech Connect

    Hura, Greg L.; Menon, Angeli L.; Hammel, Michal; Rambo, Robert P.; Poole II, Farris L.; Tsutakawa, Susan E.; Jenney Jr, Francis E.; Classen, Scott; Frankel, Kenneth A.; Hopkins, Robert C.; Yang, Sungjae; Scott, Joseph W.; Dillard, Bret D.; Adams, Michael W. W.; Tainer, John A.

    2009-07-20

    We present an efficient pipeline enabling high-throughput analysis of protein structure in solution with small angle X-ray scattering (SAXS). Our SAXS pipeline combines automated sample handling of microliter volumes, temperature and anaerobic control, rapid data collection and data analysis, and couples structural analysis with automated archiving. We subjected 50 representative proteins, mostly from Pyrococcus furiosus, to this pipeline and found that 30 were multimeric structures in solution. SAXS analysis allowed us to distinguish aggregated and unfolded proteins, define global structural parameters and oligomeric states for most samples, identify shapes and similar structures for 25 unknown structures, and determine envelopes for 41 proteins. We believe that high-throughput SAXS is an enabling technology that may change the way that structural genomics research is done.

  19. Ground Based Program for the Physical Analysis of Macromolecular Crystal Growth

    NASA Technical Reports Server (NTRS)

    Malkin, Alexander J.

    1998-01-01

    During the past year we have focused on application of in situ Atomic Force Microscopy (AFM) for studies of the growth mechanisms and kinetics of crystallization for different macromolecular systems. Mechanisms of macrostep formation and their decay, which are important in understanding of defect formation, were studied on the surfaces of thaumatin, catalase, canavalin and lysozyme crystals. Experiments revealed that step bunching on crystalline surfaces occurred either due to two- or three-dimensional nucleation on the terraces of vicinal slopes or as a result of uneven step generation by complex dislocation sources. No step bunching arising from interaction of individual steps in the course of the experiment was observed. The molecular structure of the growth steps for thaumatin and lipase crystals were deduced. It was further shown that growth step advance occurs by incorporation of single protein molecules. In singular directions growth steps move by one-dimensional nucleation on step edges followed by lateral growth. One-dimensional nuclei have different sizes, less then a single unit cell, varying for different directions of step movement. There is no roughness due to thermal fluctuations, and each protein molecule which incorporated into the step remained. Growth kinetics for catalase crystals was investigated over wide supersaturation ranges. Strong directional kinetic anisotropy in the tangential step growth rates in different directions was seen. The influence of impurities on growth kinetics and cessation of macromolecular crystals was studied. Thus, for catalase, in addition to pronounced impurity effects on the kinetics of crystallization, we were also able to directly observe adsorption of some impurities. At low supersaturation we repeatedly observed filaments which formed from impurity molecules sedimenting on the surfaces. Similar filaments were observed on the surfaces of thaumatin, canavalin and STMV crystals as well, but the frequency was low

  20. Method for minimization of solution costs for transient dynamic analysis of nonlinear periodic structures

    SciTech Connect

    Van Kirk, J.S.; Bogard, W.T.; Wood, L.R.

    1982-01-01

    Techniques are described for the transient dynamic analysis of nonlinear periodic structures subjected to time history excitations. These techniques consider the special characteristics of periodic structures in conjunction with the psuedo-force approach in numerical integration to reduce computerized solution costs. A special purpose computer code, based on these techniques, is applied to a specific nonlinear periodic structure: the response of a pressurized water reactor core to a postulated main coolant pipe rupture. Comparisons with a general purpose finite element code, using consistent integration methods, demonstrate the savings in solution cost.

  1. High-accuracy deterministic solution of the Boltzmann equation for the shock wave structure

    NASA Astrophysics Data System (ADS)

    Malkov, E. A.; Bondar, Ye. A.; Kokhanchik, A. A.; Poleshkin, S. O.; Ivanov, M. S.

    2015-07-01

    A new deterministic method of solving the Boltzmann equation has been proposed. The method has been employed in numerical studies of the plane shock wave structure in a hard sphere gas. Results for Mach numbers and have been compared with predictions of the direct simulation Monte Carlo (DSMC) method, which has been used to obtain the reference solution. Particular attention in estimating the solution accuracy has been paid to a fine structural effect: the presence of a total temperature peak exceeding the temperature value further downstream. The results of solving the Boltzmann equation for the shock wave structure are in excellent agreement with the DSMC predictions.

  2. A Web Resource for Standardized Benchmark Datasets, Metrics, and Rosetta Protocols for Macromolecular Modeling and Design

    PubMed Central

    Pache, Roland A.; Ollikainen, Noah; Kundert, Kale; O'Meara, Matthew J.; Smith, Colin A.; Kortemme, Tanja

    2015-01-01

    The development and validation of computational macromolecular modeling and design methods depend on suitable benchmark datasets and informative metrics for comparing protocols. In addition, if a method is intended to be adopted broadly in diverse biological applications, there needs to be information on appropriate parameters for each protocol, as well as metrics describing the expected accuracy compared to experimental data. In certain disciplines, there exist established benchmarks and public resources where experts in a particular methodology are encouraged to supply their most efficient implementation of each particular benchmark. We aim to provide such a resource for protocols in macromolecular modeling and design. We present a freely accessible web resource (https://kortemmelab.ucsf.edu/benchmarks) to guide the development of protocols for protein modeling and design. The site provides benchmark datasets and metrics to compare the performance of a variety of modeling protocols using different computational sampling methods and energy functions, providing a “best practice” set of parameters for each method. Each benchmark has an associated downloadable benchmark capture archive containing the input files, analysis scripts, and tutorials for running the benchmark. The captures may be run with any suitable modeling method; we supply command lines for running the benchmarks using the Rosetta software suite. We have compiled initial benchmarks for the resource spanning three key areas: prediction of energetic effects of mutations, protein design, and protein structure prediction, each with associated state-of-the-art modeling protocols. With the help of the wider macromolecular modeling community, we hope to expand the variety of benchmarks included on the website and continue to evaluate new iterations of current methods as they become available. PMID:26335248

  3. Localization of protein aggregation in Escherichia coli is governed by diffusion and nucleoid macromolecular crowding effect.

    PubMed

    Coquel, Anne-Sophie; Jacob, Jean-Pascal; Primet, Mael; Demarez, Alice; Dimiccoli, Mariella; Julou, Thomas; Moisan, Lionel; Lindner, Ariel B; Berry, Hugues

    2013-04-01

    Aggregates of misfolded proteins are a hallmark of many age-related diseases. Recently, they have been linked to aging of Escherichia coli (E. coli) where protein aggregates accumulate at the old pole region of the aging bacterium. Because of the potential of E. coli as a model organism, elucidating aging and protein aggregation in this bacterium may pave the way to significant advances in our global understanding of aging. A first obstacle along this path is to decipher the mechanisms by which protein aggregates are targeted to specific intercellular locations. Here, using an integrated approach based on individual-based modeling, time-lapse fluorescence microscopy and automated image analysis, we show that the movement of aging-related protein aggregates in E. coli is purely diffusive (Brownian). Using single-particle tracking of protein aggregates in live E. coli cells, we estimated the average size and diffusion constant of the aggregates. Our results provide evidence that the aggregates passively diffuse within the cell, with diffusion constants that depend on their size in agreement with the Stokes-Einstein law. However, the aggregate displacements along the cell long axis are confined to a region that roughly corresponds to the nucleoid-free space in the cell pole, thus confirming the importance of increased macromolecular crowding in the nucleoids. We thus used 3D individual-based modeling to show that these three ingredients (diffusion, aggregation and diffusion hindrance in the nucleoids) are sufficient and necessary to reproduce the available experimental data on aggregate localization in the cells. Taken together, our results strongly support the hypothesis that the localization of aging-related protein aggregates in the poles of E. coli results from the coupling of passive diffusion-aggregation with spatially non-homogeneous macromolecular crowding. They further support the importance of "soft" intracellular structuring (based on macromolecular

  4. A Web Resource for Standardized Benchmark Datasets, Metrics, and Rosetta Protocols for Macromolecular Modeling and Design.

    PubMed

    Ó Conchúir, Shane; Barlow, Kyle A; Pache, Roland A; Ollikainen, Noah; Kundert, Kale; O'Meara, Matthew J; Smith, Colin A; Kortemme, Tanja

    2015-01-01

    The development and validation of computational macromolecular modeling and design methods depend on suitable benchmark datasets and informative metrics for comparing protocols. In addition, if a method is intended to be adopted broadly in diverse biological applications, there needs to be information on appropriate parameters for each protocol, as well as metrics describing the expected accuracy compared to experimental data. In certain disciplines, there exist established benchmarks and public resources where experts in a particular methodology are encouraged to supply their most efficient implementation of each particular benchmark. We aim to provide such a resource for protocols in macromolecular modeling and design. We present a freely accessible web resource (https://kortemmelab.ucsf.edu/benchmarks) to guide the development of protocols for protein modeling and design. The site provides benchmark datasets and metrics to compare the performance of a variety of modeling protocols using different computational sampling methods and energy functions, providing a "best practice" set of parameters for each method. Each benchmark has an associated downloadable benchmark capture archive containing the input files, analysis scripts, and tutorials for running the benchmark. The captures may be run with any suitable modeling method; we supply command lines for running the benchmarks using the Rosetta software suite. We have compiled initial benchmarks for the resource spanning three key areas: prediction of energetic effects of mutations, protein design, and protein structure prediction, each with associated state-of-the-art modeling protocols. With the help of the wider macromolecular modeling community, we hope to expand the variety of benchmarks included on the website and continue to evaluate new iterations of current methods as they become available.

  5. An Analytical Solution for Transient Thermal Response of an Insulated Structure

    NASA Technical Reports Server (NTRS)

    Blosser, Max L.

    2012-01-01

    An analytical solution was derived for the transient response of an insulated aerospace vehicle structure subjected to a simplified heat pulse. This simplified problem approximates the thermal response of a thermal protection system of an atmospheric entry vehicle. The exact analytical solution is solely a function of two non-dimensional parameters. A simpler function of these two parameters was developed to approximate the maximum structural temperature over a wide range of parameter values. Techniques were developed to choose constant, effective properties to represent the relevant temperature and pressure-dependent properties for the insulator and structure. A technique was also developed to map a time-varying surface temperature history to an equivalent square heat pulse. Using these techniques, the maximum structural temperature rise was calculated using the analytical solutions and shown to typically agree with finite element simulations within 10 to 20 percent over the relevant range of parameters studied.

  6. Nanoparticle imaging. 3D structure of individual nanocrystals in solution by electron microscopy.

    PubMed

    Park, Jungwon; Elmlund, Hans; Ercius, Peter; Yuk, Jong Min; Limmer, David T; Chen, Qian; Kim, Kwanpyo; Han, Sang Hoon; Weitz, David A; Zettl, A; Alivisatos, A Paul

    2015-07-17

    Knowledge about the synthesis, growth mechanisms, and physical properties of colloidal nanoparticles has been limited by technical impediments. We introduce a method for determining three-dimensional (3D) structures of individual nanoparticles in solution. We combine a graphene liquid cell, high-resolution transmission electron microscopy, a direct electron detector, and an algorithm for single-particle 3D reconstruction originally developed for analysis of biological molecules. This method yielded two 3D structures of individual platinum nanocrystals at near-atomic resolution. Because our method derives the 3D structure from images of individual nanoparticles rotating freely in solution, it enables the analysis of heterogeneous populations of potentially unordered nanoparticles that are synthesized in solution, thereby providing a means to understand the structure and stability of defects at the nanoscale. PMID:26185247

  7. 3D structure of individual nanocrystals in solution by electron microscopy

    NASA Astrophysics Data System (ADS)

    Park, Jungwon; Elmlund, Hans; Ercius, Peter; Yuk, Jong Min; Limmer, David T.; Chen, Qian; Kim, Kwanpyo; Han, Sang Hoon; Weitz, David A.; Zettl, A.; Alivisatos, A. Paul

    2015-07-01

    Knowledge about the synthesis, growth mechanisms, and physical properties of colloidal nanoparticles has been limited by technical impediments. We introduce a method for determining three-dimensional (3D) structures of individual nanoparticles in solution. We combine a graphene liquid cell, high-resolution transmission electron microscopy, a direct electron detector, and an algorithm for single-particle 3D reconstruction originally developed for analysis of biological molecules. This method yielded two 3D structures of individual platinum nanocrystals at near-atomic resolution. Because our method derives the 3D structure from images of individual nanoparticles rotating freely in solution, it enables the analysis of heterogeneous populations of potentially unordered nanoparticles that are synthesized in solution, thereby providing a means to understand the structure and stability of defects at the nanoscale.

  8. SASSIE: A program to study intrinsically disordered biological molecules and macromolecular ensembles using experimental scattering restraints

    NASA Astrophysics Data System (ADS)

    Curtis, Joseph E.; Raghunandan, Sindhu; Nanda, Hirsh; Krueger, Susan

    2012-02-01

    A program to construct ensembles of biomolecular structures that are consistent with experimental scattering data are described. Specifically, we generate an ensemble of biomolecular structures by varying sets of backbone dihedral angles that are then filtered using experimentally determined restraints to rapidly determine structures that have scattering profiles that are consistent with scattering data. We discuss an application of these tools to predict a set of structures for the HIV-1 Gag protein, an intrinsically disordered protein, that are consistent with small-angle neutron scattering experimental data. We have assembled these algorithms into a program called SASSIE for structure generation, visualization, and analysis of intrinsically disordered proteins and other macromolecular ensembles using neutron and X-ray scattering restraints. Program summaryProgram title: SASSIE Catalogue identifier: AEKL_v1_0 Program summary URL:http://cpc.cs.qub.ac.uk/summaries/AEKL_v1_0.html Program obtainable from: CPC Program Library, Queen's University, Belfast, N. Ireland Licensing provisions: GNU General Public License v3 No. of lines in distributed program, including test data, etc.: 3 991 624 No. of bytes in distributed program, including test data, etc.: 826 Distribution format: tar.gz Programming language: Python, C/C++, Fortran Computer: PC/Mac Operating system: 32- and 64-bit Linux (Ubuntu 10.04, Centos 5.6) and Mac OS X (10.6.6) RAM: 1 GB Classification: 3 External routines: Python 2.6.5, numpy 1.4.0, swig 1.3.40, scipy 0.8.0, Gnuplot-py-1.8, Tcl 8.5, Tk 8.5, Mac installation requires aquaterm 1.0 (or X window system) and Xcode 3 development tools. Nature of problem: Open source software to generate structures of disordered biological molecules that subsequently allow for the comparison of computational and experimental results is limiting the use of scattering resources. Solution method: Starting with an all atom model of a protein, for example, users can input

  9. Exact solution to the problem of N bodies forming a multi-layer rotating structure.

    PubMed

    Smulsky, Joseph J

    2015-01-01

    Exact solutions to the problem of the Newtonian gravitational interaction of N material points moving around N 2 concentric circular orbits are considered. Each circular orbit contains N 3 axisymmetrically located bodies having identical masses. The structure as a whole rotates around its symmetry axis. Such structures are identical to the homographic-dynamics configurations, or planar central configurations, known from literature. Conceptually, those structures can be considered as structures formed by mutually embedded polygons with point bodies placed at polygon vortices. For structures involving less than 20 bodies, solutions were obtained using Hamiltonian-mechanics methods. In the study, the forces acting on each body in the rotating structure from the side of all other bodies were found. The differential motion equations of the bodies were reduced to a system of linear algebraic equations for the body masses. Solutions in various forms were obtained. For specifying the initial parameters and for calculating all other characteristics of the structures, a computer program RtCrcSt2.for has been developed. Structures comprising up to one million bodies have been calculated. Graphical images of obtained structures are presented, and their properties are described. Stability problems for examined structures are considered, and possible application of obtained results to celestial- and space-mechanics problems is discussed. PMID:26203407

  10. Exact solution to the problem of N bodies forming a multi-layer rotating structure.

    PubMed

    Smulsky, Joseph J

    2015-01-01

    Exact solutions to the problem of the Newtonian gravitational interaction of N material points moving around N 2 concentric circular orbits are considered. Each circular orbit contains N 3 axisymmetrically located bodies having identical masses. The structure as a whole rotates around its symmetry axis. Such structures are identical to the homographic-dynamics configurations, or planar central configurations, known from literature. Conceptually, those structures can be considered as structures formed by mutually embedded polygons with point bodies placed at polygon vortices. For structures involving less than 20 bodies, solutions were obtained using Hamiltonian-mechanics methods. In the study, the forces acting on each body in the rotating structure from the side of all other bodies were found. The differential motion equations of the bodies were reduced to a system of linear algebraic equations for the body masses. Solutions in various forms were obtained. For specifying the initial parameters and for calculating all other characteristics of the structures, a computer program RtCrcSt2.for has been developed. Structures comprising up to one million bodies have been calculated. Graphical images of obtained structures are presented, and their properties are described. Stability problems for examined structures are considered, and possible application of obtained results to celestial- and space-mechanics problems is discussed.

  11. The NMR structure of cyclosporin A bound to cyclophilin in aqueous solution

    SciTech Connect

    Weber, C.; Wilder, G.; von Freyberg, B.; Braun, W.; Wuethrich, K. ); Traber, R.; Widmer, H. )

    1991-07-02

    Cyclosporin A bound to the presumed receptor protein cyclophilin was studied in aqueous solution at pH 6.0 by nuclear magnetic resonance spectroscopy using uniform {sup 15}N- or {sup 13}C-labeling of cyclosporin A and heteronuclear spectral editing techniques. With an input of 108 intramolecular NOEs and four vicinal {sup 3}J{sub HN{alpha}} coupling constants, the three-dimensional structure of cyclosporin A bound to cyclophilin was calculated with the distance geometry program DISMAN, and the structures resulting from 181 converged calculations were energy refined with the program FANTOM. A group of 120 conformers was selected on the basis of the residual constraint violations and energy criteria to represent the solution structure. The average of the pairwise root-mean-square distances calculated for the backbone atoms of the 120 structures was 0.58 {angstrom}. The structure represents a novel conformation of cyclosporin A, for which the backbone conformation is significantly different from the previously reported structures in single crystals and in chloroform solution. The structure has all peptide bonds in the trans form, contains no elements of regular secondary structure and no intramolecular hydrogen bonds, and exposes nearly all polar groups to its environment. The root-mean-square distance between the backbone atoms of the crystal structure of cyclosporin A and the mean of the 120 conformers representing the NMR structure of cyclosporin A bound to cyclophilin is 2.5 {angstrom}.

  12. The structure and dynamics in solution of Cu(I) pseudoazurin from Paracoccus pantotrophus.

    PubMed Central

    Thompson, G. S.; Leung, Y. C.; Ferguson, S. J.; Radford, S. E.; Redfield, C.

    2000-01-01

    The solution structure and backbone dynamics of Cu(I) pseudoazurin, a 123 amino acid electron transfer protein from Paracoccus pantotrophus, have been determined using NMR methods. The structure was calculated to high precision, with a backbone RMS deviation for secondary structure elements of 0.35+/-0.06 A, using 1,498 distance and 55 torsion angle constraints. The protein has a double-wound Greek-key fold with two alpha-helices toward its C-terminus, similar to that of its oxidized counterpart determined by X-ray crystallography. Comparison of the Cu(I) solution structure with the X-ray structure of the Cu(II) protein shows only small differences in the positions of some of the secondary structure elements. Order parameters S2, measured for amide nitrogens, indicate that the backbone of the protein is rigid on the picosecond to nanosecond timescale. PMID:10850794

  13. A brief history of macromolecular crystallography, illustrated by a family tree and its Nobel fruits.

    PubMed

    Jaskolski, Mariusz; Dauter, Zbigniew; Wlodawer, Alexander

    2014-09-01

    As a contribution to the celebration of the year 2014, declared by the United Nations to be 'The International Year of Crystallography', the FEBS Journal is dedicating this issue to papers showcasing the intimate union between macromolecular crystallography and structural biology, both in historical perspective and in current research. Instead of a formal editorial piece, by way of introduction, this review discusses the most important, often iconic, achievements of crystallographers that led to major advances in our understanding of the structure and function of biological macromolecules. We identified at least 42 scientists who received Nobel Prizes in Physics, Chemistry or Medicine for their contributions that included the use of X-rays or neutrons and crystallography, including 24 who made seminal discoveries in macromolecular sciences. Our spotlight is mostly, but not only, on the recipients of this most prestigious scientific honor, presented in approximately chronological order. As a summary of the review, we attempt to construct a genealogy tree of the principal lineages of protein crystallography, leading from the founding members to the present generation.

  14. A refined solution structure of hen lysozyme determined using residual dipolar coupling data.

    PubMed

    Schwalbe, H; Grimshaw, S B; Spencer, A; Buck, M; Boyd, J; Dobson, C M; Redfield, C; Smith, L J

    2001-04-01

    A high resolution NMR structure of hen lysozyme has been determined using 209 residual 1H-15N dipolar coupling restraints from measurements made in two different dilute liquid crystalline phases (bicelles) in conjunction with a data set of 1632 NOE distance restraints, 110 torsion angle restraints, and 60 hydrogen bond restraints. The ensemble of 50 low-energy calculated structures has an average backbone RMSD of 0.50+/-0.13A to the mean structure and of 1.49+/-0.10A to the crystal structure of hen lysozyme. To assess the importance of the dipolar coupling data in the structure determination, the final structures are compared with an ensemble calculated using an identical protocol but excluding the dipolar coupling restraints. The comparison shows that structures calculated with the dipolar coupling data are more similar to the crystal structure than those calculated without, and have better stereochemical quality. The structures also show improved quality factors when compared with additional dipolar coupling data that were not included in the structure calculations, with orientation-dependent 15N chemical shift changes measured in the bicelle solutions, and with T1/T2 values obtained from 15N relaxation measurements. Analysis of the ensemble of NMR structures and comparisons with crystal structures, 15N relaxation data, and molecular dynamics simulations of hen lysozyme provides a detailed description of the solution structure of this protein and insights into its dynamical behavior.

  15. Discussions on equivalent solutions and localized structures via the mapping method based on Riccati equation

    NASA Astrophysics Data System (ADS)

    Xu, Ling; Cheng, Xuan; Dai, Chao-Qing

    2015-12-01

    Although the mapping method based on Riccati equation was proposed to obtain variable separation solutions many years ago, two important problems have not been studied: i) the equivalence of variable separation solutions by means of the mapping method based on Riccati equation with the radical sign combined ansatz; and ii) lack of physical meanings for some localized structures constructed by variable separation solutions. In this paper, we re-study the (2+1)-dimensional Boiti-Leon-Pempinelli equation via the mapping method based on Riccati equation and prove that nine types of variable separation solutions are actually equivalent to each other. Moreover, we also re-study localized structures constructed by variable separation solutions. Results indicate that some localized structures reported in the literature are lacking real values due to the appearance of the divergent and un-physical phenomenon for the initial field. Therefore, we must be careful with the initial field to avoid the appearance of some un-physical or even divergent structures in it when we construct localized structures for the potential field.

  16. Local structure of the halite-sylvine solid solution according to the computer simulation data

    SciTech Connect

    Urusov, V. S. Leonenko, E. V.

    2008-09-15

    The structural, elastic, and thermodynamic properties of halite NaCl and sylvine KCl and the miscibility properties of the NaCl-KCl solid solution found by computer simulation are in good agreement with the experimental data. Analysis of the relaxation of the solid solution structure suggests that both anion and cation sublattices are distorted; however, the anion sublattice is distorted much more strongly. Calculations of the local bond valence at all types of ions in the solid solution show opposite deviations from the balance at cations, whereas the general balance is retained. The values of the electrostatic potential in the ion positions reflect weakening of bonding in the solid solution with respect to its pure components. In addition, with an increase in the average interatomic distance in the first coordination sphere around cations, the modulus of the electrostatic potential at cations decreases.

  17. Structure solution of DNA-binding proteins and complexes with ARCIMBOLDO libraries

    SciTech Connect

    Pröpper, Kevin; Meindl, Kathrin; Sammito, Massimo; Dittrich, Birger; Sheldrick, George M.; Pohl, Ehmke; Usón, Isabel

    2014-06-01

    The structure solution of DNA-binding protein structures and complexes based on the combination of location of DNA-binding protein motif fragments with density modification in a multi-solution frame is described. Protein–DNA interactions play a major role in all aspects of genetic activity within an organism, such as transcription, packaging, rearrangement, replication and repair. The molecular detail of protein–DNA interactions can be best visualized through crystallography, and structures emphasizing insight into the principles of binding and base-sequence recognition are essential to understanding the subtleties of the underlying mechanisms. An increasing number of high-quality DNA-binding protein structure determinations have been witnessed despite the fact that the crystallographic particularities of nucleic acids tend to pose specific challenges to methods primarily developed for proteins. Crystallographic structure solution of protein–DNA complexes therefore remains a challenging area that is in need of optimized experimental and computational methods. The potential of the structure-solution program ARCIMBOLDO for the solution of protein–DNA complexes has therefore been assessed. The method is based on the combination of locating small, very accurate fragments using the program Phaser and density modification with the program SHELXE. Whereas for typical proteins main-chain α-helices provide the ideal, almost ubiquitous, small fragments to start searches, in the case of DNA complexes the binding motifs and DNA double helix constitute suitable search fragments. The aim of this work is to provide an effective library of search fragments as well as to determine the optimal ARCIMBOLDO strategy for the solution of this class of structures.

  18. Evaporation induced self-assembly of ordered structures from a capillary-held solution

    NASA Astrophysics Data System (ADS)

    Hong, Suck Won

    The use of spontaneous self-assembly as a lithography and external fields-free means to construct well-ordered, often intriguing structures has received much attention as a result of the ease of producing complex structures with small feature sizes. Self-assembly via irreversible solvent evaporation of a droplet containing nonvolatile solutes (polymers, nanoparticles, and colloids) represents one such case. However, the flow instabilities within the evaporating droplet often result in irregular dissipative structures (e.g., convection patterns and fingering instabilities). Therefore, fully utilizing evaporation as a simple tool for creating well-ordered structures that have numerous technological applications requires delicate control over several factors, including the evaporative flux, solution concentration, interfacial interaction between the solute and the substrate, etc. In this study, we developed a simple route to produce highly regular polymeric structures in an easily controllable, cost-effective, and reproducible manner simply by allowing a drop to evaporate in a confined geometry consisting of a sphere on a Si surface (i.e., a sphere-on-Si geometry). The confined geometry provides unique environment for controlling the flow within the evaporating droplet, which, in turn, regulates the structure formation. A variety of polymers, including poly[2-methoxy-5-(2-ethylhexyloxy)-1,4-phenylenevinylene] (MEH-PPV), poly(ferrocenyldimethylsilane) (PFS), polystyrene (PS), poly(methyl methacrylate) (PMMA), and polystyrene-block-poly(methyl methacrylate) (PS-b-PMMA), are selected as nonvolatile solutes. A number of parameters are found to effectively mediate the structure formation, including the solution concentration, the interfacial interaction between the solute and the substrate, curvature and molecular effect. This simple, lithography-free route allows subsequent preparation of various metal, metal oxide, and carbon nanotube patterns with controlled spacing

  19. Use of Plastic Capillaries for Macromolecular Crystallization

    NASA Technical Reports Server (NTRS)

    Potter, Rachel R.; Hong, Young-Soo; Ciszak, Ewa M.

    2003-01-01

    Methods of crystallization of biomolecules in plastic capillaries (Nalgene 870 PFA tubing) are presented. These crystallization methods used batch, free-interface liquid- liquid diffusion alone, or a combination with vapor diffusion. Results demonstrated growth of crystals of test proteins such as thaumatin and glucose isomerase, as well as protein studied in our laboratory such dihydrolipoamide dehydrogenase. Once the solutions were loaded in capillaries, they were stored in the tubes in frozen state at cryogenic temperatures until the desired time of activation of crystallization experiments.

  20. [Structure of crambin in solution, crystal and in the trajectories of molecular dynamics simulations].

    PubMed

    Abaturov, L V; Nosova, N G

    2013-01-01

    The mechanisms of the three-dimensional crambin structure alterations in the crystalline environments and in the trajectories of the molecular dynamics simulations in the vacuum and crystal surroundings have been analyzed. In the crystalline state and in the solution the partial regrouping of remote intramolecular packing contacts, involved in the formation and stabilization of the tertiary structure of the crambin molecule, occurs in NMR structures. In the crystalline state it is initiated by the formation of the intermolecular contacts, the conformational influence of its appearance is distributed over the structure. The changes of the conformations and positions of the residues of the loop segments, where the intermolecular contacts of the crystal surroundings are preferably concentrated, are most observable. Under the influence of these contacts the principal change of the regular secondary structure of crambin is taking place: extension of the two-strand beta structure to the three-strand structure with the participation of the single last residue N46 of the C-terminal loop. In comparison with the C-terminal loop the more profound changes are observed in the conformation and the atomic positions of the backbone atoms and in the solvent accessibility of the residues of the interhelical loop. In the solution of the ensemble of the 8 NMR structures relative accessibility to the solvent differs more noticeably also in the region of the loop segments and rather markedly in the interhelical loop. In the crambin cryogenic crystal structures the positions of the atoms of the backbone and/or side chain of 14-18 of 46 residues are discretely disordered. The disorganizations of at least 8 of 14 residues occur directly in the regions of the intermolecular contacts and another 5 residues are disordered indirectly through the intramolecular contacts with the residues of the intermolecular contacts. Upon the molecular dynamics simulation in the vacuum surrounding as in the

  1. Structural determinants of protein stabilization by solutes. The important of the hairpin loop in rubredoxins.

    PubMed

    Pais, Tiago M; Lamosa, Pedro; dos Santos, Wagner; Legall, Jean; Turner, David L; Santos, Helena

    2005-02-01

    Despite their high sequence homology, rubredoxins from Desulfovibrio gigas and D. desulfuricans are stabilized to very different extents by compatible solutes such as diglycerol phosphate, the major osmolyte in the hyperthermophilic archaeon Archaeoglobus fulgidus[Lamosa P, Burke A, Peist R, Huber R, Liu M Y, Silva G, Rodrigues-Pousada C, LeGall J, Maycock C and Santos H (2000) Appl Environ Microbiol66, 1974-1979]. The principal structural difference between these two proteins is the absence of the hairpin loop in the rubredoxin from D. desulfuricans. Therefore, mutants of D. gigas rubredoxin bearing deletions in the loop region were constructed to investigate the importance of this structural feature on protein intrinsic stability, as well as on its capacity to undergo stabilization by compatible solutes. The three-dimensional structure of the mutant bearing the largest deletion, Delta17/29, was determined by 1H-NMR, demonstrating that, despite the drastic deletion, the main structural features were preserved. The dependence of the NH chemical shifts on temperature and solute concentration (diglycerol phosphate or mannosylglycerate) provide evidence of subtle conformational changes induced by the solute. The kinetic stability (as assessed from the absorption decay at 494 nm) of six mutant rubredoxins was determined at 90 degrees C and the stabilizing effect exerted by both solutes was assessed. The extent of protection conferred by each solute was highly dependent on the specific mutant examined: while the half-life for iron release in the wild-type D. gigas rubredoxin increased threefold in the presence of 0.1 M diglycerol phosphate, mutant Delta23/29 was destabilized. This study provides evidence for solute-induced compaction of the protein structure and occurrence of weak, specific interactions with the protein surface. The relevance of these findings to our understanding of the molecular basis for protein stabilization is discussed.

  2. Ice crystallization in ultrafine water-salt aerosols: nucleation, ice-solution equilibrium, and internal structure.

    PubMed

    Hudait, Arpa; Molinero, Valeria

    2014-06-01

    Atmospheric aerosols have a strong influence on Earth's climate. Elucidating the physical state and internal structure of atmospheric aqueous aerosols is essential to predict their gas and water uptake, and the locus and rate of atmospherically important heterogeneous reactions. Ultrafine aerosols with sizes between 3 and 15 nm have been detected in large numbers in the troposphere and tropopause. Nanoscopic aerosols arising from bubble bursting of natural and artificial seawater have been identified in laboratory and field experiments. The internal structure and phase state of these aerosols, however, cannot yet be determined in experiments. Here we use molecular simulations to investigate the phase behavior and internal structure of liquid, vitrified, and crystallized water-salt ultrafine aerosols with radii from 2.5 to 9.5 nm and with up to 10% moles of ions. We find that both ice crystallization and vitrification of the nanodroplets lead to demixing of pure water from the solutions. Vitrification of aqueous nanodroplets yields nanodomains of pure low-density amorphous ice in coexistence with vitrified solute rich aqueous glass. The melting temperature of ice in the aerosols decreases monotonically with an increase of solute fraction and decrease of radius. The simulations reveal that nucleation of ice occurs homogeneously at the subsurface of the water-salt nanoparticles. Subsequent ice growth yields phase-segregated, internally mixed, aerosols with two phases in equilibrium: a concentrated water-salt amorphous mixture and a spherical cap-like ice nanophase. The surface of the crystallized aerosols is heterogeneous, with ice and solution exposed to the vapor. Free energy calculations indicate that as the concentration of salt in the particles, the advance of the crystallization, or the size of the particles increase, the stability of the spherical cap structure increases with respect to the alternative structure in which a core of ice is fully surrounded by

  3. Ice crystallization in ultrafine water-salt aerosols: nucleation, ice-solution equilibrium, and internal structure.

    PubMed

    Hudait, Arpa; Molinero, Valeria

    2014-06-01

    Atmospheric aerosols have a strong influence on Earth's climate. Elucidating the physical state and internal structure of atmospheric aqueous aerosols is essential to predict their gas and water uptake, and the locus and rate of atmospherically important heterogeneous reactions. Ultrafine aerosols with sizes between 3 and 15 nm have been detected in large numbers in the troposphere and tropopause. Nanoscopic aerosols arising from bubble bursting of natural and artificial seawater have been identified in laboratory and field experiments. The internal structure and phase state of these aerosols, however, cannot yet be determined in experiments. Here we use molecular simulations to investigate the phase behavior and internal structure of liquid, vitrified, and crystallized water-salt ultrafine aerosols with radii from 2.5 to 9.5 nm and with up to 10% moles of ions. We find that both ice crystallization and vitrification of the nanodroplets lead to demixing of pure water from the solutions. Vitrification of aqueous nanodroplets yields nanodomains of pure low-density amorphous ice in coexistence with vitrified solute rich aqueous glass. The melting temperature of ice in the aerosols decreases monotonically with an increase of solute fraction and decrease of radius. The simulations reveal that nucleation of ice occurs homogeneously at the subsurface of the water-salt nanoparticles. Subsequent ice growth yields phase-segregated, internally mixed, aerosols with two phases in equilibrium: a concentrated water-salt amorphous mixture and a spherical cap-like ice nanophase. The surface of the crystallized aerosols is heterogeneous, with ice and solution exposed to the vapor. Free energy calculations indicate that as the concentration of salt in the particles, the advance of the crystallization, or the size of the particles increase, the stability of the spherical cap structure increases with respect to the alternative structure in which a core of ice is fully surrounded by

  4. Atomic detail brownian dynamics simulations of concentrated protein solutions with a mean field treatment of hydrodynamic interactions.

    SciTech Connect

    Mereghetti, Paolo; Wade, Rebecca C.

    2012-07-26

    High macromolecular concentrations are a distinguishing feature of living organisms. Understanding how the high concentration of solutes affects the dynamic properties of biological macromolecules is fundamental for the comprehension of biological processes in living systems. In this paper, we describe the implementation of mean field models of translational and rotational hydrodynamic interactions into an atomically detailed many-protein brownian dynamics simulation method. Concentrated solutions (30-40% volume fraction) of myoglobin, hemoglobin A, and sickle cell hemoglobin S were simulated, and static structure factors, oligomer formation, and translational and rotational self-diffusion coefficients were computed. Good agreement of computed properties with available experimental data was obtained. The results show the importance of both solvent mediated interactions and weak protein-protein interactions for accurately describing the dynamics and the association properties of concentrated protein solutions. Specifically, they show a qualitative difference in the translational and rotational dynamics of the systems studied. Although the translational diffusion coefficient is controlled by macromolecular shape and hydrodynamic interactions, the rotational diffusion coefficient is affected by macromolecular shape, direct intermolecular interactions, and both translational and rotational hydrodynamic interactions.

  5. Mathieu function solutions for photoacoustic waves in sinusoidal one-dimensional structures.

    PubMed

    Wu, Binbin; Diebold, Gerald J

    2012-07-01

    The photoacoustic effect for a one-dimensional structure, the sound speed of which varies sinusoidally in space, is shown to be governed by an inhomogeneous Mathieu equation with the forcing term dependent on the spatial and temporal properties of the exciting optical radiation. New orthogonality relations, traveling wave Mathieu functions, and solutions to the inhomogeneous Mathieu equation are found, which are used to determine the character of photoacoustic waves in infinite and finite length phononic structures. Floquet solutions to the Mathieu equation give the positions of the band gaps, the damping of the acoustic waves within the band gaps, and the dispersion relation for photoacoustic waves. The solutions to the Mathieu equation give the photoacoustic response of the structure, show the space equivalent of subharmonic generation and acoustic confinement when waves are excited within band gaps.

  6. Evidence for water structuring forces between surfaces

    SciTech Connect

    Stanley, Christopher B; Rau, Dr. Donald

    2011-01-01

    Structured water on apposing surfaces can generate significant energies due to reorganization and displacement as the surfaces encounter each other. Force measurements on a multitude of biological structures using the osmotic stress technique have elucidated commonalities that point toward an underlying hydration force. In this review, the forces of two contrasting systems are considered in detail: highly charged DNA and nonpolar, uncharged hydroxypropyl cellulose. Conditions for both net repulsion and attraction, along with the measured exclusion of chemically different solutes from these macromolecular surfaces, are explored and demonstrate features consistent with a hydration force origin. Specifically, the observed interaction forces can be reduced to the effects of perturbing structured surface water.

  7. On the variability of experimental data in macromolecular crystallography.

    PubMed

    Pozharski, Edwin

    2012-09-01

    Experimental errors as determined by data-processing algorithms in macromolecular crystallography are compared with the direct error estimates obtained by a multiple crystal data-collection protocol. It is found that several-fold error inflation is necessary to account for crystal-to-crystal variation. It is shown that similar error inflation is observed for data collected from multiple sections of the same crystal, indicating non-uniform crystal growth as one of the likely sources of additional data variation. Other potential sources of error inflation include differential X-ray absorption for different reflections and variation of unit-cell parameters. The underestimation of the experimental errors is more severe in lower resolution shells and for reflections characterized by a higher signal-to-noise ratio. These observations partially account for the gap between the expected and the observed R values in macromolecular crystallography.

  8. Stochastic reaction–diffusion algorithms for macromolecular crowding

    NASA Astrophysics Data System (ADS)

    Sturrock, Marc

    2016-06-01

    Compartment-based (lattice-based) reaction–diffusion algorithms are often used for studying complex stochastic spatio-temporal processes inside cells. In this paper the influence of macromolecular crowding on stochastic reaction–diffusion simulations is investigated. Reaction–diffusion processes are considered on two different kinds of compartmental lattice, a cubic lattice and a hexagonal close packed lattice, and solved using two different algorithms, the stochastic simulation algorithm and the spatiocyte algorithm (Arjunan and Tomita 2010 Syst. Synth. Biol. 4, 35–53). Obstacles (modelling macromolecular crowding) are shown to have substantial effects on the mean squared displacement and average number of molecules in the domain but the nature of these effects is dependent on the choice of lattice, with the cubic lattice being more susceptible to the effects of the obstacles. Finally, improvements for both algorithms are presented.

  9. Solution structure of a hydrophobic analogue of the winter flounder antifreeze protein.

    PubMed

    Liepinsh, Edvards; Otting, Gottfried; Harding, Margaret M; Ward, Leanne G; Mackay, Joel P; Haymet, A D J

    2002-02-01

    The solution structure of a synthetic mutant type I antifreeze protein (AFP I) was determined in aqueous solution at pH 7.0 using nuclear magnetic resonance (NMR) spectroscopy. The mutations comprised the replacement of the four Thr residues by Val and the introduction of two additional Lys-Glu salt bridges. The antifreeze activity of this mutant peptide, VVVV2KE, has been previously shown to be similar to that of the wild type protein, HPLC6 (defined here as TTTT). The solution structure reveals an alphahelix bent in the same direction as the more bent conformer of the published crystal structure of TTTT, while the side chain chi1 rotamers of VVVV2KE are similar to those of the straighter conformer in the crystal of TTTT. The Val side chains of VVVV2KE assume the same orientations as the Thr side chains of TTTT, confirming the conservative nature of this mutation. The combined data suggest that AFP I undergoes an equilibrium between straight and bent helices in solution, combined with independent equilibria between different side chain rotamers for some of the amino acid residues. The present study presents the first complete sequence-specific resonance assignments and the first complete solution structure determination by NMR of any AFP I protein.

  10. New insights into structural alteration of enamel apatite induced by citric acid and sodium fluoride solutions.

    PubMed

    Wang, Xiaojie; Klocke, Arndt; Mihailova, Boriana; Tosheva, Lubomira; Bismayer, Ulrich

    2008-07-24

    Attenuated total reflectance infrared spectroscopy and complementary scanning electron microscopy were applied to analyze the surface structure of enamel apatite exposed to citric acid and to investigate the protective potential of fluorine-containing reagents against citric acid-induced erosion. Enamel and, for comparison, geological hydroxylapatite samples were treated with aqueous solutions of citric acid and sodium fluoride of different concentrations, ranging from 0.01 to 0.5 mol/L for citric acid solutions and from 0.5 to 2.0% for fluoride solutions. The two solutions were applied either simultaneously or consecutively. The citric acid-induced structural modification of apatite increases with the increase in the citric acid concentration and the number of treatments. The application of sodium fluoride alone does not suppress the atomic level changes in apatite exposed to acidic agents. The addition of sodium fluoride to citric acid solutions leads to formation of surface CaF2 and considerably reduces the changes in the apatite P-O-Ca framework. However, the CaF2 globules deposited on the enamel surface seem to be insufficient to prevent the alteration of the apatite structure upon further exposure to acidic agents. No evidence for fluorine-induced recovery of the apatite structure was found.

  11. Impact of synchrotron radiation on macromolecular crystallography: a personal view

    PubMed Central

    Dauter, Zbigniew; Jaskolski, Mariusz; Wlodawer, Alexander

    2010-01-01

    The introduction of synchrotron radiation sources almost four decades ago has led to a revolutionary change in the way that diffraction data from macromolecular crystals are being collected. Here a brief history of the development of methodologies that took advantage of the availability of synchrotron sources are presented, and some personal experiences with the utilization of synchrotrons in the early days are recalled. PMID:20567074

  12. A 3D cellular context for the macromolecular world

    PubMed Central

    Patwardhan, Ardan; Ashton, Alun; Brandt, Robert; Butcher, Sarah; Carzaniga, Raffaella; Chiu, Wah; Collinson, Lucy; Doux, Pascal; Duke, Elizabeth; Ellisman, Mark H; Franken, Erik; Grünewald, Kay; Heriche, Jean-Karim; Koster, Abraham; Kühlbrandt, Werner; Lagerstedt, Ingvar; Larabell, Carolyn; Lawson, Catherine L; Saibil, Helen R; Sanz-García, Eduardo; Subramaniam, Sriram; Verkade, Paul; Swedlow, Jason R; Kleywegt, Gerard J

    2015-01-01

    We report the outcomes of the discussion initiated at the workshop entitled A 3D Cellular Context for the Macromolecular World and propose how data from emerging three-dimensional (3D) cellular imaging techniques—such as electron tomography, 3D scanning electron microscopy and soft X-ray tomography—should be archived, curated, validated and disseminated, to enable their interpretation and reuse by the biomedical community. PMID:25289590

  13. Thermodynamics and Statistical Mechanics of Macromolecular Systems

    NASA Astrophysics Data System (ADS)

    Bachmann, Michael

    2014-04-01

    Preface and outline; 1. Introduction; 2. Statistical mechanics: a modern review; 3. The complexity of minimalistic lattice models for protein folding; 4. Monte Carlo and chain growth methods for molecular simulations; 5. First insights to freezing and collapse of flexible polymers; 6. Crystallization of elastic polymers; 7. Structural phases of semiflexible polymers; 8. Generic tertiary folding properties of proteins in mesoscopic scales; 9. Protein folding channels and kinetics of two-state folding; 10. Inducing generic secondary structures by constraints; 11. Statistical analyses of aggregation processes; 12. Hierarchical nature of phase transitions; 13. Adsorption of polymers at solid substrates; 14. Hybrid protein-substrate interfaces; 15. Concluding remarks and outlook; Notes; References; Index.

  14. Solution Structure of the PhoP DNA-Binding Domain from Mycobacterium tuberculosis

    PubMed Central

    Macdonald, Ramsay; Sarkar, Dibyendu; Amer, Brendan R.; Clubb, Robert T.

    2015-01-01

    Summary Tuberculosis caused by Mycobacterium tuberculosis is a leading cause of death world-wide. The PhoP protein is required for virulence and is part of the PhoPR two-component system that regulates gene expression. The NMR-derived solution structure of the PhoP C-terminal DNA-binding domain is reported. Residues 150 to 246 form a structured domain that contains a winged helix-turn-helix motif. We provide evidence that the transactivation loop postulated to contact RNA polymerase is partially disordered in solution, and that the polypeptide that connects the DNA-binding domain to the regulatory domain is unstructured. PMID:26209027

  15. Frequency modelling and solution of fluid-structure interaction in complex pipelines

    NASA Astrophysics Data System (ADS)

    Xu, Yuanzhi; Johnston, D. Nigel; Jiao, Zongxia; Plummer, Andrew R.

    2014-05-01

    Complex pipelines may have various structural supports and boundary conditions, as well as branches. To analyse the vibrational characteristics of piping systems, frequency modelling and solution methods considering complex constraints are developed here. A fourteen-equation model and Transfer Matrix Method (TMM) are employed to describe Fluid-Structure Interaction (FSI) in liquid-filled pipes. A general solution for the multi-branch pipe is proposed in this paper, offering a methodology to predict frequency responses of the complex piping system. Some branched pipe systems are built for the purpose of validation, indicating good agreement with calculated results.

  16. A Solution Adaptive Structured/Unstructured Overset Grid Flow Solver with Applications to Helicopter Rotor Flows

    NASA Technical Reports Server (NTRS)

    Duque, Earl P. N.; Biswas, Rupak; Strawn, Roger C.

    1995-01-01

    This paper summarizes a method that solves both the three dimensional thin-layer Navier-Stokes equations and the Euler equations using overset structured and solution adaptive unstructured grids with applications to helicopter rotor flowfields. The overset structured grids use an implicit finite-difference method to solve the thin-layer Navier-Stokes/Euler equations while the unstructured grid uses an explicit finite-volume method to solve the Euler equations. Solutions on a helicopter rotor in hover show the ability to accurately convect the rotor wake. However, isotropic subdivision of the tetrahedral mesh rapidly increases the overall problem size.

  17. Macromolecular Assemblage in the Design of a Synthetic AIDS Vaccine

    NASA Astrophysics Data System (ADS)

    Defoort, Jean-Philippe; Nardelli, Bernardetta; Huang, Wolin; Ho, David D.; Tam, James P.

    1992-05-01

    We describe a peptide vaccine model based on the mimicry of surface coat protein of a pathogen. This model used a macromolecular assemblage approach to amplify peptide antigens in liposomes or micelles. The key components of the model consisted of an oligomeric lysine scaffolding to amplify peptide antigens covalently 4-fold and a lipophilic membrane-anchoring group to further amplify noncovalently the antigens many-fold in liposomal or micellar form. A peptide antigen derived from the third variable domain of glycoprotein gp120 of human immunodeficiency virus type 1 (HIV-1), consisting of neutralizing, T-helper, and T-cytotoxic epitopes, was used in a macromolecular assemblage model (HIV-1 linear peptide amino acid sequence 308-331 in a tetravalent multiple antigen peptide system linked to tripalmitoyl-S-glycerylcysteine). The latter complex, in liposome or micelle, was used to immunize mice and guinea pigs without any adjuvant and found to induce gp120-specific antibodies that neutralize virus infectivity in vitro, elicit cytokine production, and prime CD8^+ cytotoxic T lymphocytes in vivo. Our results show that the macromolecular assemblage approach bears immunological mimicry of the gp120 of HIV virus and may lead to useful vaccines against HIV infection.

  18. Folate-mediated delivery of macromolecular anticancer therapeutic agents.

    PubMed

    Lu, Yingjuan; Low, Philip S

    2002-09-13

    The receptor for folic acid constitutes a useful target for tumor-specific drug delivery, primarily because: (1) it is upregulated in many human cancers, including malignancies of the ovary, brain, kidney, breast, myeloid cells and lung, (2) access to the folate receptor in those normal tissues that express it can be severely limited due to its location on the apical (externally-facing) membrane of polarized epithelia, and (3) folate receptor density appears to increase as the stage/grade of the cancer worsens. Thus, cancers that are most difficult to treat by classical methods may be most easily targeted with folate-linked therapeutics. To exploit these peculiarities of folate receptor expression, folic acid has been linked to both low molecular weight drugs and macromolecular complexes as a means of targeting the attached molecules to malignant cells. Conjugation of folic acid to macromolecules has been shown to enhance their delivery to folate receptor-expressing cancer cells in vitro in almost all situations tested. Folate-mediated macromolecular targeting in vivo has, however, yielded only mixed results, largely because of problems with macromolecule penetration of solid tumors. Nevertheless, prominent examples do exist where folate targeting has significantly improved the outcome of a macromolecule-based therapy, leading to complete cures of established tumors in many cases. This review presents a brief mechanistic background of folate-targeted macromolecular therapeutics and then summarizes the successes and failures observed with each major application of the technology.

  19. Path Similarity Analysis: A Method for Quantifying Macromolecular Pathways

    PubMed Central

    Seyler, Sean L.; Kumar, Avishek; Thorpe, M. F.; Beckstein, Oliver

    2015-01-01

    Diverse classes of proteins function through large-scale conformational changes and various sophisticated computational algorithms have been proposed to enhance sampling of these macromolecular transition paths. Because such paths are curves in a high-dimensional space, it has been difficult to quantitatively compare multiple paths, a necessary prerequisite to, for instance, assess the quality of different algorithms. We introduce a method named Path Similarity Analysis (PSA) that enables us to quantify the similarity between two arbitrary paths and extract the atomic-scale determinants responsible for their differences. PSA utilizes the full information available in 3N-dimensional configuration space trajectories by employing the Hausdorff or Fréchet metrics (adopted from computational geometry) to quantify the degree of similarity between piecewise-linear curves. It thus completely avoids relying on projections into low dimensional spaces, as used in traditional approaches. To elucidate the principles of PSA, we quantified the effect of path roughness induced by thermal fluctuations using a toy model system. Using, as an example, the closed-to-open transitions of the enzyme adenylate kinase (AdK) in its substrate-free form, we compared a range of protein transition path-generating algorithms. Molecular dynamics-based dynamic importance sampling (DIMS) MD and targeted MD (TMD) and the purely geometric FRODA (Framework Rigidity Optimized Dynamics Algorithm) were tested along with seven other methods publicly available on servers, including several based on the popular elastic network model (ENM). PSA with clustering revealed that paths produced by a given method are more similar to each other than to those from another method and, for instance, that the ENM-based methods produced relatively similar paths. PSA applied to ensembles of DIMS MD and FRODA trajectories of the conformational transition of diphtheria toxin, a particularly challenging example, showed that

  20. A theory for water and macromolecular transport in the pulmonary artery wall with a detailed comparison to the aorta.

    PubMed

    Zeng, Zhongqing; Jan, Kung-Ming; Rumschitzki, David S

    2012-04-15

    The pulmonary artery (PA) wall, which has much higher hydraulic conductivity and albumin void space and approximately one-sixth the normal transmural pressure of systemic arteries (e.g, aorta, carotid arteries), is rarely atherosclerotic, except under pulmonary hypertension. This study constructs a detailed, two-dimensional, wall-structure-based filtration and macromolecular transport model for the PA to investigate differences in prelesion transport processes between the disease-susceptible aorta and the relatively resistant PA. The PA and aorta models are similar in wall structure, but very different in parameter values, many of which have been measured (and therefore modified) since the original aorta model of Huang et al. (23). Both PA and aortic model simulations fit experimental data on transwall LDL concentration profiles and on the growth of isolated endothelial (horseradish peroxidase) tracer spots with circulation time very well. They reveal that lipid entering the aorta attains a much higher intima than media concentration but distributes better between these regions in the PA than aorta and that tracer in both regions contributes to observed tracer spots. Solutions show why both the overall transmural water flow and spot growth rates are similar in these vessels despite very different material transport parameters. Since early lipid accumulation occurs in the subendothelial intima and since (matrix binding) reaction kinetics depend on reactant concentrations, the lower intima lipid concentrations in the PA vs. aorta likely lead to slower accumulation of bound lipid in the PA. These findings may be relevant to understanding the different atherosusceptibilities of these vessels. PMID:22198178

  1. A theory for water and macromolecular transport in the pulmonary artery wall with a detailed comparison to the aorta

    PubMed Central

    Zeng, Zhongqing; Jan, Kung-Ming

    2012-01-01

    The pulmonary artery (PA) wall, which has much higher hydraulic conductivity and albumin void space and approximately one-sixth the normal transmural pressure of systemic arteries (e.g, aorta, carotid arteries), is rarely atherosclerotic, except under pulmonary hypertension. This study constructs a detailed, two-dimensional, wall-structure-based filtration and macromolecular transport model for the PA to investigate differences in prelesion transport processes between the disease-susceptible aorta and the relatively resistant PA. The PA and aorta models are similar in wall structure, but very different in parameter values, many of which have been measured (and therefore modified) since the original aorta model of Huang et al. (23). Both PA and aortic model simulations fit experimental data on transwall LDL concentration profiles and on the growth of isolated endothelial (horseradish peroxidase) tracer spots with circulation time very well. They reveal that lipid entering the aorta attains a much higher intima than media concentration but distributes better between these regions in the PA than aorta and that tracer in both regions contributes to observed tracer spots. Solutions show why both the overall transmural water flow and spot growth rates are similar in these vessels despite very different material transport parameters. Since early lipid accumulation occurs in the subendothelial intima and since (matrix binding) reaction kinetics depend on reactant concentrations, the lower intima lipid concentrations in the PA vs. aorta likely lead to slower accumulation of bound lipid in the PA. These findings may be relevant to understanding the different atherosusceptibilities of these vessels. PMID:22198178

  2. Rasta silanes: new silyl resins with novel macromolecular architecture via living free radical polymerization

    PubMed

    Lindsley; Hodges; Filzen; Watson; Geyer

    2000-09-01

    Heating TEMPO-methyl resin with dialkylsilane styrenes affords larger resin beads via living free radical polymerization. The new silyl resins prepared by this solvent-free suspension polymerization protocol have been coined "Rasta silanes". Rasta silanes have a novel macromolecular architecture typified by long straight chain polymers bearing the silanes which emanate from the phenyl rings of a cross-linked polystyrene core. By careful selection of comonomers during the polymerization step, loading capacity, silane spacing, and the relative distance of the silane moieties from the resin core can be controlled. The consistently high-loading Rasta silane resins produced can be easily converted into either a reactive silyl chloride or triflate to subsequently anchor alcohols and phenols to the solid phase. Cleavage from the resin can be mediated by treatment with HF.pyridine, TFA solutions, or TBAF.

  3. Polarizable Atomic Multipole X-Ray Refinement: Particle Mesh Ewald Electrostatics for Macromolecular Crystals.

    PubMed

    Schnieders, Michael J; Fenn, Timothy D; Pande, Vijay S

    2011-04-12

    Refinement of macromolecular models from X-ray crystallography experiments benefits from prior chemical knowledge at all resolutions. As the quality of the prior chemical knowledge from quantum or classical molecular physics improves, in principle so will resulting structural models. Due to limitations in computer performance and electrostatic algorithms, commonly used macromolecules X-ray crystallography refinement protocols have had limited support for rigorous molecular physics in the past. For example, electrostatics is often neglected in favor of nonbonded interactions based on a purely repulsive van der Waals potential. In this work we present advanced algorithms for desktop workstations that open the door to X-ray refinement of even the most challenging macromolecular data sets using state-of-the-art classical molecular physics. First we describe theory for particle mesh Ewald (PME) summation that consistently handles the symmetry of all 230 space groups, replicates of the unit cell such that the minimum image convention can be used with a real space cutoff of any size and the combination of space group symmetry with replicates. An implementation of symmetry accelerated PME for the polarizable atomic multipole optimized energetics for biomolecular applications (AMOEBA) force field is presented. Relative to a single CPU core performing calculations on a P1 unit cell, our AMOEBA engine called Force Field X (FFX) accelerates energy evaluations by more than a factor of 24 on an 8-core workstation with a Tesla GPU coprocessor for 30 structures that contain 240 000 atoms on average in the unit cell. The benefit of AMOEBA electrostatics evaluated with PME for macromolecular X-ray crystallography refinement is demonstrated via rerefinement of 10 crystallographic data sets that range in resolution from 1.7 to 4.5 Å. Beginning from structures obtained by local optimization without electrostatics, further optimization using AMOEBA with PME electrostatics improved

  4. Crystal and solution structures of a superantigen from Yersinia pseudotuberculosis reveal a jelly-roll fold.

    PubMed

    Donadini, Roberta; Liew, Chu Wai; Kwan, Ann H Y; Mackay, Joel P; Fields, Barry A

    2004-01-01

    Superantigens are a class of microbial proteins with the ability to excessively activate T cells by binding to the T cell receptor. The staphylococcal and streptococcal superantigens are closely related in structure and possess an N-terminal domain that resembles an OB fold and a C-terminal domain similar to a beta-grasp fold. Yersinia pseudotuberculosis produces superantigens, YPMa, YPMb, and YPMc, which have no significant amino acid similarity to other proteins. We have determined the crystal and solution structures of YPMa, which show that the protein has a jelly-roll fold. The closest structural neighbors to YPMa are viral capsid proteins and members of the tumor necrosis factor superfamily. In the crystal structure, YPMa packs as a trimer, another feature shared with viral capsid proteins and TNF superfamily proteins. However, in solution YPMa behaves as a monomer, and any functional relevance of the trimer observed in the crystals is yet to be established.

  5. Structural characterization of NaOH aqueous solution in the glass and liquid states

    NASA Astrophysics Data System (ADS)

    Bruni, F.; Ricci, M. A.; Soper, A. K.

    2001-05-01

    Using the technique of hydrogen and deuterium substitution, the structure of water in concentrated NaOH solution (10 M) is explored. It is found that major changes in water structure occur both in the liquid phase at T=300 K and in the glassy phase at T=173 K. In particular the 4.4 Å peak in the OO pair correlation function of pure water, which is normally viewed as indicating tetrahedral short-range coordination in water, is totally absent in the NaOH solution at room temperature, and shows up only as a small feature in the NaOH solution in the glassy state. Corresponding changes occur in the OH and HH correlation functions: The hydrogen bond peak position is shifted from 1.85 Å in pure water to 1.65 Å for both the liquid and glassy NaOH, with a reduced number of hydrogen bonds in the glassy phase. The intramolecular HH distance, 1.5 Å, of the water molecule is unaffected by the presence of the solute, but the positions of the peaks in the HH function at 2.4 and 3.8 Å, due to the orientational correlation between neighboring pure water molecules, are respectively, shifted to 2.15 and 3.5 Å. The above findings indicate that ions in aqueous solutions induce a change in water structure equivalent to the application of high pressures.

  6. Comparison of shock structure solutions using independent continuum and kinetic theory approaches

    NASA Technical Reports Server (NTRS)

    Fiscko, Kurt A.; Chapman, Dean R.

    1988-01-01

    A vehicle traversing the atmosphere will experience flight regimes at high altitudes in which the thickness of a hypersonic shock wave is not small compared to the shock standoff distance from the hard body. When this occurs, it is essential to compute accurate flow field solutions within the shock structure. In this paper, one-dimensional shock structure is investigated for various monatomic gases from Mach 1.4 to Mach 35. Kinetic theory solutions are computed using the Direct Simulation Monte Carlo method. Steady-state solutions of the Navier-Stokes equations and of a slightly truncated form of the Burnett equations are determined by relaxation to a steady state of the time-dependent continuum equations. Monte Carlo results are in excellent agreement with published experimental data and are used as bases of comparison for continuum solutions. For a Maxwellian gas, the truncated Burnett equations are shown to produce far more accurate solutions of shock structure than the Navier-Stokes equations.

  7. Molecular Effects of Concentrated Solutes on Protein Hydration, Dynamics, and Electrostatics.

    PubMed

    Abriata, Luciano A; Spiga, Enrico; Peraro, Matteo Dal

    2016-08-23

    Most studies of protein structure and function are performed in dilute conditions, but proteins typically experience high solute concentrations in their physiological scenarios and biotechnological applications. High solute concentrations have well-known effects on coarse protein traits like stability, diffusion, and shape, but likely also perturb other traits through finer effects pertinent at the residue and atomic levels. Here, NMR and molecular dynamics investigations on ubiquitin disclose variable interactions with concentrated solutes that lead to localized perturbations of the protein's surface, hydration, electrostatics, and dynamics, all dependent on solute size and chemical properties. Most strikingly, small polar uncharged molecules are sticky on the protein surface, whereas charged small molecules are not, but the latter still perturb the internal protein electrostatics as they diffuse nearby. Meanwhile, interactions with macromolecular crowders are favored mainly through hydrophobic, but not through polar, surface patches. All the tested small solutes strongly slow down water exchange at the protein surface, whereas macromolecular crowders do not exert such strong perturbation. Finally, molecular dynamics simulations predict that unspecific interactions slow down microsecond- to millisecond-timescale protein dynamics despite having only mild effects on pico- to nanosecond fluctuations as corroborated by NMR. We discuss our results in the light of recent advances in understanding proteins inside living cells, focusing on the physical chemistry of quinary structure and cellular organization, and we reinforce the idea that proteins should be studied in native-like media to achieve a faithful description of their function. PMID:27558718

  8. Structure and properties of regenerated Antheraea pernyi silk fibroin in aqueous solution.

    PubMed

    Tao, Wei; Li, Mingzhong; Zhao, Chunxia

    2007-04-10

    Antheraea pernyi silk fibroin fibers were dissolved by aqueous lithium thiocyanate to obtain regenerated A. pernyi silk fibroin solution. By means of circular dichroism, (13)C NMR and Raman spectroscopy, the molecular conformation of regenerated A. pernyi silk fibroin in aqueous solution was investigated. The relationship of environmental factors and sol-gel transformation behavior of regenerated A. pernyi silk fibroin was also studied. The molecular conformations of regenerated A. pernyi silk fibroin mainly were alpha-helix and random coil in solution. There also existed a little beta-sheet conformation. It was obviously different with Bombyx mori silk fibroin, whose molecular conformation in solution was only random coil but no alpha-helix existence. With the increase of temperature and solution concentration and with the decrease of solution pH value, the gelation velocity of regenerated A. pernyi silk fibroin solution increased. Especially, it showed that A. pernyi silk fibroin was more sensitive to temperature than B. mori silk fibroin during the sol-gel transformation. The velocity increased obviously when the temperature was above 30 degrees C. During the sol-gel transformation, the molecular conformation of regenerated A. pernyi silk fibroin changed from random coil to beta-sheet structure. The results of these studies provided important insight into the preparation of new biomaterials by silk fibroin protein.

  9. Combining crystallography and EPR: crystal and solution structures of the multidomain cochaperone DnaJ

    SciTech Connect

    Barends, Thomas R. M.; Brosi, Richard W. W.; Steinmetz, Andrea; Scherer, Anna; Hartmann, Elisabeth; Eschenbach, Jessica; Lorenz, Thorsten; Seidel, Ralf; Shoeman, Robert L.; Zimmermann, Sabine; Bittl, Robert; Schlichting, Ilme; Reinstein, Jochen

    2013-08-01

    The crystal structure of the N-terminal part of T. thermophilus DnaJ unexpectedly showed an ordered GF domain and guided the design of a construct enabling the first structure determination of a complete DnaJ cochaperone molecule. By combining the crystal structures with spin-labelling EPR and cross-linking in solution, a dynamic view of this flexible molecule was developed. Hsp70 chaperones assist in a large variety of protein-folding processes in the cell. Crucial for these activities is the regulation of Hsp70 by Hsp40 cochaperones. DnaJ, the bacterial homologue of Hsp40, stimulates ATP hydrolysis by DnaK (Hsp70) and thus mediates capture of substrate protein, but is also known to possess chaperone activity of its own. The first structure of a complete functional dimeric DnaJ was determined and the mobility of its individual domains in solution was investigated. Crystal structures of the complete molecular cochaperone DnaJ from Thermus thermophilus comprising the J, GF and C-terminal domains and of the J and GF domains alone showed an ordered GF domain interacting with the J domain. Structure-based EPR spin-labelling studies as well as cross-linking results showed the existence of multiple states of DnaJ in solution with different arrangements of the various domains, which has implications for the function of DnaJ.

  10. Local structure in the disordered solid solution of cis- and trans-perinones.

    PubMed

    Teteruk, Jaroslav L; Glinnemann, Jürgen; Heyse, Winfried; Johansson, Kristoffer E; van de Streek, Jacco; Schmidt, Martin U

    2016-06-01

    The cis- and trans-isomers of the polycyclic aromatic compound perinone, C26H12N4O2, form a solid solution (Vat Red 14). This solid solution is isotypic to the crystal structures of cis-perinone (Pigment Red 194) and trans-perinone (Pigment Orange 34) and exhibits a combined positional and orientational disorder: In the crystal, each molecular position is occupied by either a cis- or trans-perinone molecule, both of which have two possible molecular orientations. The structure of cis-perinone exhibits a twofold orientational disorder, whereas the structure of trans-perinone is ordered. The crystal structure of the solid solution was determined by single-crystal X-ray analysis. Extensive lattice-energy minimizations with force-field and DFT-D methods were carried out on combinatorially complete sets of ordered models. For the disordered systems, local structures were calculated, including preferred local arrangements, ordering lengths, and probabilities for the arrangement of neighbouring molecules. The superposition of the atomic positions of all energetically favourable calculated models corresponds well with the experimentally determined crystal structures, explaining not only the atomic positions, but also the site occupancies and anisotropic displacement parameters. PMID:27240774

  11. A decade of user operation on the macromolecular crystallography MAD beamline ID14-4 at the ESRF.

    PubMed

    McCarthy, Andrew A; Brockhauser, Sandor; Nurizzo, Didier; Theveneau, Pascal; Mairs, Trevor; Spruce, Darren; Guijarro, Matias; Lesourd, Marc; Ravelli, Raimond B G; McSweeney, Sean

    2009-11-01

    ID14-4 at the ESRF is the first tunable undulator-based macromolecular crystallography beamline that can celebrate a decade of user service. During this time ID14-4 has not only been instrumental in the determination of the structures of biologically important molecules but has also contributed significantly to the development of various instruments, novel data collection schemes and pioneering radiation damage studies on biological samples. Here, the evolution of ID14-4 over the last decade is presented, and some of the major improvements that were carried out in order to maintain its status as one of the most productive macromolecular crystallography beamlines are highlighted. The experimental hutch has been upgraded to accommodate a high-precision diffractometer, a sample changer and a large CCD detector. More recently, the optical hutch has been refurbished in order to improve the X-ray beam quality on ID14-4 and to incorporate the most modern and robust optical elements used at other ESRF beamlines. These new optical elements will be described and their effect on beam stability discussed. These studies may be useful in the design, construction and maintenance of future X-ray beamlines for macromolecular crystallography and indeed other applications, such as those planned for the ESRF upgrade.

  12. Pitfalls in assessing microvascular endothelial barrier function: impedance-based devices versus the classic macromolecular tracer assay

    PubMed Central

    Bischoff, Iris; Hornburger, Michael C.; Mayer, Bettina A.; Beyerle, Andrea; Wegener, Joachim; Fürst, Robert

    2016-01-01

    The most frequently used parameters to describe the barrier properties of endothelial cells (ECs) in vitro are (i) the macromolecular permeability, indicating the flux of a macromolecular tracer across the endothelium, and (ii) electrical impedance of ECs grown on gold-film electrodes reporting on the cell layer’s tightness for ion flow. Due to the experimental differences between these approaches, inconsistent observations have been described. Here, we present the first direct comparison of these assays applied to one single cell type (human microvascular ECs) under the same experimental conditions. The impact of different pharmacological tools (histamine, forskolin, Y-27632, blebbistatin, TRAP) on endothelial barrier function was analyzed by Transwell® tracer assays and two commercial impedance devices (xCELLigence®, ECIS®). The two impedance techniques provided very similar results for all compounds, whereas macromolecular permeability readings were found to be partly inconsistent with impedance. Possible reasons for these discrepancies are discussed. We conclude that the complementary combination of both approaches is highly recommended to overcome the restrictions of each assay. Since the nature of the growth support may contribute to the observed differences, structure-function relationships should be based on cells that are consistently grown on either permeable or impermeable growth supports in all experiments. PMID:27025965

  13. Neutron reflectivity study of the swollen structure of polyzwitterion and polyeletrolyte brushes in aqueous solution.

    PubMed

    Kobayashi, Motoyasu; Ishihara, Kazuhiko; Takahara, Atsushi

    2014-01-01

    The swollen brush structures of polycation and zwitterionic polymer brushes, such as poly(2-methacryloyloxyethyltrimethylammonium chloride) (PMTAC), poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC), and poly[3-(N-2-methacryloyloxyethyl-N,N-dimethyl)ammonatopropanesulfonate] (PMAPS), in aqueous solutions of various ionic strengths were characterized by neutron reflectivity (NR) measurements. A series of the polyelectrolyte brushes were prepared by surface-initiated controlled radical polymerization on silicon substrates. A high-graft-density PMTAC brush in salt-free water (D2O) adopted a two-region step-like structure consisting of a shrunk region near the Si substrate surface and a diffuse brush region with a relatively stretched chain structure at the solution interface. The diffuse region of PMTAC was reduced with increase in salt (NaCl) concentration. The PMAPS brush in D2O formed a collapsed structure due to the strong molecular interaction between betaine groups, while significant increase in the swollen thickness was observed in salt aqueous solution. In contrast, no change was observed in the depth profile of the swollen PMPC brush in D2O with various salt concentrations. The unique solution behaviors of zwitterionic polymer brushes were described. PMID:25178564

  14. Parallel-vector solution of large-scale structural analysis problems on supercomputers

    NASA Technical Reports Server (NTRS)

    Storaasli, Olaf O.; Nguyen, Duc T.; Agarwal, Tarun K.

    1989-01-01

    A direct linear equation solution method based on the Choleski factorization procedure is presented which exploits both parallel and vector features of supercomputers. The new equation solver is described, and its performance is evaluated by solving structural analysis problems on three high-performance computers. The method has been implemented using Force, a generic parallel FORTRAN language.

  15. Comparing Multiple Solutions in the Structured Problem Solving: Deconstructing Japanese Lessons from Learner's Perspective

    ERIC Educational Resources Information Center

    Hino, Keiko

    2015-01-01

    The purpose of this study is to enhance our understanding of how students listen and attend to multiple solutions proposed by their classmates during the activity of comparison. This study examines ten consecutive lessons in each of the two eighth-grade classrooms in Tokyo that are organized in the style of "structured problem solving".…

  16. Decision-making in structure solution using Bayesian estimates of map quality: the PHENIX autosol wizard

    SciTech Connect

    Terwilliger, Thomas C; Adams, Paul D; Read, Randy J; Mccoy, Airlie J

    2008-01-01

    Ten measures of experimental electron-density-map quality are examined and the skewness of electron density is found to be the best indicator of actual map quality. A Bayesian approach to estimating map quality is developed and used in the PHENIX AutoSol wizard to make decisions during automated structure solution.

  17. Protein folding, protein structure and the origin of life: Theoretical methods and solutions of dynamical problems

    NASA Technical Reports Server (NTRS)

    Weaver, D. L.

    1982-01-01

    Theoretical methods and solutions of the dynamics of protein folding, protein aggregation, protein structure, and the origin of life are discussed. The elements of a dynamic model representing the initial stages of protein folding are presented. The calculation and experimental determination of the model parameters are discussed. The use of computer simulation for modeling protein folding is considered.

  18. Structure and Interactions of Fish Type III Antifreeze Protein in Solution

    PubMed Central

    Salvay, Andrés G.; Gabel, Frank; Pucci, Bernard; Santos, Javier; Howard, Eduardo I.; Ebel, Christine

    2010-01-01

    Abstract It has been suggested that above a critical protein concentration, fish Type III antifreeze protein (AFP III) self-assembles to form micelle-like structures that may play a key role in antifreeze activity. To understand the complex activity of AFP III, a comprehensive description of its association state and structural organization in solution is necessary. We used analytical ultracentrifugation, analytical size-exclusion chromatography, and dynamic light scattering to characterize the interactions and homogeneity of AFP III in solution. Small-angle neutron scattering was used to determine the low-resolution structure in solution. Our results clearly show that at concentrations up to 20 mg mL−1 and at temperatures of 20°C, 6°C, and 4°C, AFP III is monomeric in solution and adopts a structure compatible with that determined by crystallography. Surface tension measurements show a propensity of AFP III to localize at the air/water interface, but this surface activity is not correlated with any aggregation in the bulk. These results support the hypothesis that each AFP III molecule acts independently of the others, and that specific intermolecular interactions between monomers are not required for binding to ice. The lack of attractive interactions between monomers may be functionally important, allowing for more efficient binding and covering of the ice surface. PMID:20643081

  19. Structure and interactions of fish type III antifreeze protein in solution.

    PubMed

    Salvay, Andrés G; Gabel, Frank; Pucci, Bernard; Santos, Javier; Howard, Eduardo I; Ebel, Christine

    2010-07-21

    It has been suggested that above a critical protein concentration, fish Type III antifreeze protein (AFP III) self-assembles to form micelle-like structures that may play a key role in antifreeze activity. To understand the complex activity of AFP III, a comprehensive description of its association state and structural organization in solution is necessary. We used analytical ultracentrifugation, analytical size-exclusion chromatography, and dynamic light scattering to characterize the interactions and homogeneity of AFP III in solution. Small-angle neutron scattering was used to determine the low-resolution structure in solution. Our results clearly show that at concentrations up to 20 mg mL(-1) and at temperatures of 20 degrees C, 6 degrees C, and 4 degrees C, AFP III is monomeric in solution and adopts a structure compatible with that determined by crystallography. Surface tension measurements show a propensity of AFP III to localize at the air/water interface, but this surface activity is not correlated with any aggregation in the bulk. These results support the hypothesis that each AFP III molecule acts independently of the others, and that specific intermolecular interactions between monomers are not required for binding to ice. The lack of attractive interactions between monomers may be functionally important, allowing for more efficient binding and covering of the ice surface.

  20. Reformulation of Maxwell's equations to incorporate near-solute solvent structure.

    PubMed

    Yang, Pei-Kun; Lim, Carmay

    2008-09-01

    Maxwell's equations, which treat electromagnetic interactions between macroscopic charged objects in materials, have explained many phenomena and contributed to many applications in our lives. Derived in 1861 when no methods were available to determine the atomic structure of macromolecules, Maxwell's equations assume the solvent to be a structureless continuum. However, near-solute solvent molecules are highly structured, unlike far-solute bulk solvent molecules. Current methods cannot treat both the near-solute solvent structure and time-dependent electromagnetic interactions in a macroscopic system. Here, we derive "microscopic" electrodynamics equations that can treat macroscopic time-dependent electromagnetic field problems like Maxwell's equations and reproduce the solvent molecular and dipole density distributions observed in molecular dynamics simulations. These equations greatly reduce computational expense by not having to include explicit solvent molecules, yet they treat the solvent electrostatic and van der Waals effects more accurately than continuum models. They provide a foundation to study electromagnetic interactions between molecules in a macroscopic system that are ubiquitous in biology, bioelectromagnetism, and nanotechnology. The general strategy presented herein to incorporate the near-solute solvent structure would enable studies on how complex cellular protein-ligand interactions are affected by electromagnetic radiation, which could help to prevent harmful electromagnetic spectra or find potential therapeutic applications. PMID:18698705

  1. The solvent component of macromolecular crystals

    PubMed Central

    Weichenberger, Christian X.; Afonine, Pavel V.; Kantardjieff, Katherine; Rupp, Bernhard

    2015-01-01

    The mother liquor from which a biomolecular crystal is grown will contain water, buffer molecules, native ligands and cofactors, crystallization precipitants and additives, various metal ions, and often small-molecule ligands or inhibitors. On average, about half the volume of a biomolecular crystal consists of this mother liquor, whose components form the disordered bulk solvent. Its scattering contributions can be exploited in initial phasing and must be included in crystal structure refinement as a bulk-solvent model. Concomitantly, distinct electron density originating from ordered solvent components must be correctly identified and represented as part of the atomic crystal structure model. Herein, are reviewed (i) probabilistic bulk-solvent content estimates, (ii) the use of bulk-solvent density modification in phase improvement, (iii) bulk-solvent models and refinement of bulk-solvent contributions and (iv) modelling and validation of ordered solvent constituents. A brief summary is provided of current tools for bulk-solvent analysis and refinement, as well as of modelling, refinement and analysis of ordered solvent components, including small-molecule ligands. PMID:25945568

  2. Molecular dynamics simulations of large macromolecular complexes

    PubMed Central

    Perilla, Juan R.; Goh, Boon Chong; Cassidy, C. Keith; Liu, Bo; Bernardi, Rafael C.; Rudack, Till; Yu, Hang; Wu, Zhe; Schulten, Klaus

    2015-01-01

    Connecting dynamics to structural data from diverse experimental sources, molecular dynamics simulations permit the exploration of biological phenomena in unparalleled detail. Advances in simulations are moving the atomic resolution descriptions of biological systems into the million-to-billion atom regime, in which numerous cell functions reside. In this opinion, we review the progress, driven by large-scale molecular dynamics simulations, in the study of viruses, ribosomes, bioenergetic systems, and other diverse applications. These examples highlight the utility of molecular dynamics simulations in the critical task of relating atomic detail to the function of supramolecular complexes, a task that cannot be achieved by smaller-scale simulations or existing experimental approaches alone. PMID:25845770

  3. Macromolecular assemblies in reduced gravity environments

    NASA Technical Reports Server (NTRS)

    Moos, Philip J.; Hayes, James W.; Stodieck, Louis S.; Luttges, Marvin W.

    1990-01-01

    The assembly of protein macro molecules into structures commonly produced within biological systems was achieved using in vitro techniques carried out in nominal as well as reduced gravity environments. Appropriate hardware was designed and fabricated to support such studies. Experimental protocols were matched to the available reduced gravity test opportunities. In evaluations of tubulin, fibrin and collagen assembly products the influence of differing gravity test conditions are apparent. Product homogeneity and organization were characteristic enhancements documented in reduced gravity samples. These differences can be related to the fluid flow conditions that exist during in vitro product formation. Reduced gravity environments may provide a robust opportunity for directing the products formed in a variety of bioprocessing applications.

  4. Solution NMR structure of a designed metalloprotein and complementary molecular dynamics refinement.

    PubMed

    Calhoun, Jennifer R; Liu, Weixia; Spiegel, Katrin; Dal Peraro, Matteo; Klein, Michael L; Valentine, Kathleen G; Wand, A Joshua; DeGrado, William F

    2008-02-01

    We report the solution NMR structure of a designed dimetal-binding protein, di-Zn(II) DFsc, along with a secondary refinement step employing molecular dynamics techniques. Calculation of the initial NMR structural ensemble by standard methods led to distortions in the metal-ligand geometries at the active site. Unrestrained molecular dynamics using a nonbonded force field for the metal shell, followed by quantum mechanical/molecular mechanical dynamics of DFsc, were used to relax local frustrations at the dimetal site that were apparent in the initial NMR structure and provide a more realistic description of the structure. The MD model is consistent with NMR restraints, and in good agreement with the structural and functional properties expected for DF proteins. This work demonstrates that NMR structures of metalloproteins can be further refined using classical and first-principles molecular dynamics methods in the presence of explicit solvent to provide otherwise unavailable insight into the geometry of the metal center.

  5. Refined solution structure of the oligomerization domain of the tumour suppressor p53.

    PubMed

    Clore, G M; Ernst, J; Clubb, R; Omichinski, J G; Kennedy, W M; Sakaguchi, K; Appella, E; Gronenborn, A M

    1995-04-01

    The NMR solution structure of the oligomerization domain of the tumour suppressor p53 (residues 319-360) has been refined. The structure comprises a dimer of dimers, oriented in an approximately orthogonal manner. The present structure determination is based on 4,472 experimental NMR restraints which represents a three and half fold increase over our previous work in the number of NOE restraints at the tetramerization interface. A comparison with the recently solved 1.7 A resolution X-ray structure shows that the structures are very similar and that the average angular root-mean-square difference in the interhelical angles is about 1 degree. The results of recent extensive mutagenesis data and the possible effects of mutations which have been identified in human cancers are discussed in the light of the present structure. PMID:7796267

  6. Solution Structure and Dynamics of Human Hemoglobin in the Carbonmonoxy Form

    PubMed Central

    Fan, Jing-Song; Zheng, Yu; Choy, Wing-Yiu; Simplaceanu, Virgil; Ho, Nancy T.; Ho, Chien; Yang, Daiwen

    2014-01-01

    The solution structure of HbCO A was refined using stereospecifically assigned methyl groups and residual dipolar couplings based on our previous NMR structure. The tertiary structures of individual chains were found to be very similar to the X-ray structures, while the quaternary structures in solution with low-salt resembled the X-ray R structure more than the R2 structure. On the basis of chemical shift perturbation by inositol hexaphosphate (IHP) titration and docking, we identified five possible IHP binding sites in HbCO A. Amide-water proton exchange experiments demonstrated that αThr38 located in the α1β2 interface and several loop regions in both α- and β-chains were dynamic on the sub-second timescale. Side-chain methyl dynamics revealed that methyl groups in the α1β2 interface were dynamic, but those in the α1β1 interface were quite rigid on the ns-ps and ms-µs timescales. All the data strongly suggest a dynamic α1β2 interface that allows conformational changes among different forms (like T, R, and R2) easily in solution. Binding of IHP to HbCO A induced small structural and dynamics changes in the α1β2 interface and the regions around the hemes, but did not increase the conformational entropy of HbCO A. The binding also caused conformational changes on the ms timescale, very likely arising from the relative motion of the α1β1 dimer with respect to the α2β2 dimer. Heterotropic effectors like IHP may change the oxygen affinity of Hb through modulating the relative motion of the two dimers and then further altering the structure of heme binding regions. PMID:23901897

  7. Structure and dynamics of a polysaccharide matrix: aqueous solutions of bacterial levan.

    PubMed

    Benigar, Elizabeta; Dogsa, Iztok; Stopar, David; Jamnik, Andrej; Kralj Cigić, Irena; Tomšič, Matija

    2014-04-15

    The polysaccharide levan is a homopolymer of fructose and appears in nature as an important structural component of some bacterial biofilms. This paper reports the structural and dynamic properties of aqueous solutions of levan of various origin obtained from dynamic rheological, small-angle X-ray scattering, static and dynamic light scattering, as well as density and sound velocity measurements, determination of polymer branching after per-O-methylation, and microscopy. Besides samples of commercially available levan from Zymomonas mobilis and Erwinia herbicola, we also isolated, purified, and studied a levan sample from the biofilm of Bacillus subtilis. The results of dynamic rheological and light scattering measurements revealed very interesting viscoelastic properties of levan solutions even at very low polymer concentrations. The findings were complemented by small-angle X-ray scattering data that revealed some important differences in the structure of the aqueous levan solutions at the molecular level. Besides presenting detailed dynamic and structural results on the polysaccharide systems of various levans, one of the essential goals of this work was to point out the level of structural information that may be obtained for such polymer systems by combining basic physicochemical, rheological, and various light scattering techniques. PMID:24654746

  8. Solution structure of CEH-37 homeodomain of the nematode Caenorhabditis elegans

    SciTech Connect

    Moon, Sunjin; Lee, Yong Woo; Kim, Woo Taek; Lee, Weontae

    2014-01-10

    Highlights: •We have determined solution structures of CEH-37 homedomain. •CEH-37 HD has a compact α-helical structure with HTH DNA binding motif. •Solution structure of CEH-37 HD shares its molecular topology with that of the homeodomain proteins. •Residues in the N-terminal region and HTH motif are important in binding to Caenorhabditis elegans telomeric DNA. •CEH-37 could play an important role in telomere function via DNA binding. -- Abstract: The nematode Caenorhabditis elegans protein CEH-37 belongs to the paired OTD/OTX family of homeobox-containing homeodomain proteins. CEH-37 shares sequence similarity with homeodomain proteins, although it specifically binds to double-stranded C. elegans telomeric DNA, which is unusual to homeodomain proteins. Here, we report the solution structure of CEH-37 homeodomain and molecular interaction with double-stranded C. elegans telomeric DNA using nuclear magnetic resonance (NMR) spectroscopy. NMR structure shows that CEH-37 homeodomain is composed of a flexible N-terminal region and three α-helices with a helix-turn-helix (HTH) DNA binding motif. Data from size-exclusion chromatography and fluorescence spectroscopy reveal that CEH-37 homeodomain interacts strongly with double-stranded C. elegans telomeric DNA. NMR titration experiments identified residues responsible for specific binding to nematode double-stranded telomeric DNA. These results suggest that C. elegans homeodomain protein, CEH-37 could play an important role in telomere function via DNA binding.

  9. Grain boundary structure and solute segregation in titanium-doped sapphire bicrystals

    SciTech Connect

    Taylor, Seth T.

    2002-05-17

    Solute segregation to ceramic grain boundaries governs material processing and microstructure evolution, and can strongly influence material properties critical to engineering performance. Understanding the evolution and implications of grain boundary chemistry is a vital component in the greater effort to engineer ceramics with controlled microstructures. This study examines solute segregation to engineered grain boundaries in titanium-doped sapphire (Al2O3) bicrystals, and explores relationships between grain boundary structure and chemistry at the nanometer scale using spectroscopic and imaging techniques in the transmission electron microscope (TEM). Results demonstrate dramatic changes in solute segregation stemming from small fluctuations in grain boundary plane and structure. Titanium and silicon solute species exhibit strong tendencies to segregate to non-basal and basal grain boundary planes, respectively. Evidence suggests that grain boundary faceting occurs in low-angle twis t boundaries to accommodate nonequilibrium solute segregation related to slow specimen cooling rates, while faceting of tilt grain boundaries often occurs to expose special planes of the coincidence site lattice (CSL). Moreover, quantitative analysis of grain boundary chemistry indicates preferential segregation of charged defects to grain boundary dislocations. These results offer direct proof that static dislocations in ionic materials can assume a net charge, and emphasize the importance of interactions between charged point, line, and planar defects in ionic materials. Efforts to understand grain boundary chemistry in terms of space charge theory, elastic misfit and nonequilibrium segregation are discussed for the Al2O3 system.

  10. Structural Properties and Phase Behavior of Crosslinked Networks in Polymer Solutions

    PubMed Central

    Benmouna, Farida; Zemmour, Samira; Benmouna, Mustapha

    2016-01-01

    ABSTRACT Structural properties and phase behavior of crosslinked networks embedded in polymer solutions are theoretically investigated. The partial structure factor of the network is calculated using a matrix formulation of the random phase approximation and the forward scattering limit is correlated with the phase behavior. Swelling and deswelling processes are analyzed in terms of the polymer concentration, the mismatch of solvent quality with respect to polymer and network, the polymers incompatibility and their characteristic sizes. Most studies reported so far in the literature have focussed on the swelling of crosslinked networks and gels in pure solvents but the correlation of the structural properties with the phase behavior in the presence of high molecular weight polymers in solution has not been given sufficient attention. The present work is intended to fill this gap in view of the current efforts to develop novel drug encapsulating and targeted delivery devices. PMID:27134310

  11. Solution structure of a soluble fragment derived from a membrane protein by shotgun proteolysis

    PubMed Central

    Allen, Mark D.; Christie, Mary; Jones, Peter; Porebski, Benjamin T.; Roome, Brendan; Freund, Stefan M.V.; Buckle, Ashley M.; Bycroft, Mark; Christ, Daniel

    2015-01-01

    We have previously reported a phage display method for the identification of protein domains on a genome-wide scale (shotgun proteolysis). Here we present the solution structure of a fragment of the Escherichia coli membrane protein yrfF, as identified by shotgun proteolysis, and determined by NMR spectroscopy. Despite the absence of computational predictions, the fragment formed a well-defined beta-barrel structure, distantly falling within the OB-fold classification. Our results highlight the potential of high-throughput experimental approaches for the identification of protein domains for structural studies. PMID:25877662

  12. Assessment of solvent effects: do weak alignment media affect the structure of the solute?

    PubMed

    Shahkhatuni, Astghik A; Shahkhatuni, Aleksan G; Panosyan, Henry A; Sahakyan, Aleksandr B; Byeon, In-Ja L; Gronenborn, Angela M

    2007-07-01

    Alignment media used for measuring residual dipolar couplings, such as solutions of filamentous phages, phospholipid mixtures, polyacrylamide gels and various lyotropic liquid crystalline systems were investigated with respect to solvent effects on molecular structure. Structural parameters of the small rigid model compound 13C-acetonitrile were calculated from dipolar couplings and variations from expectation values were used for assessment of solvent effects. Only minor solvent effects were observed for most of the media employed and the measured structural data are in good agreement with microwave data and theoretical predictions. PMID:17534883

  13. Structural and Electronic Properties of a Wide-Gap Quaternary Solid Solution: \\(Zn, Mg\\) \\(S, Se\\)

    NASA Astrophysics Data System (ADS)

    Saitta, A. M.; de Gironcoli, S.; Baroni, S.

    1998-06-01

    The structural properties of the (Zn, Mg) (S, Se) solid solutions are determined by a combination of the computational alchemy and the cluster expansion methods with Monte Carlo simulations. We determine the phase diagram of the alloy and show that the homogeneous phase is characterized by a large amount of short-range order occurring among first-nearest neighbors. Electronic-structure calculations performed using the special quasirandom structure approach indicate that the energy gap of the alloy is rather sensitive to this short-range order.

  14. Biophotonic probing of macromolecular transformations during apoptosis

    PubMed Central

    Pliss, Artem; Kuzmin, Andrey N.; Kachynski, Aliaksandr V.; Prasad, Paras N.

    2010-01-01

    We introduce here multiplex nonlinear optical imaging as a powerful tool for studying the molecular organization and its transformation in cellular processes, with the specific example of apoptosis. Apoptosis is a process of self-initiated cell death, critically important for physiological regulation and elimination of genetic disorders. Nonlinear optical microscopy, combining the coherent anti-Stokes Raman scattering (CARS) microscopy and two-photon excited fluorescence (TPEF), has been used for analysis of spatial distribution of major types of biomolecules: proteins, lipids, and nucleic acids in the cells while monitoring their changes during apoptosis. CARS imaging revealed that in the nuclei of proliferating cells, the proteins are distributed nearly uniformly, with local accumulations in several nuclear structures. We have found that this distribution is abruptly disrupted at the onset of apoptosis and is transformed to a progressively irregular pattern. Fluorescence recovery after photobleaching (FRAP) studies indicate that pronounced aggregation of proteins in the nucleoplasm of apoptotic cells coincides with a gradual reduction in their mobility. PMID:20615987

  15. Structure and intermolecular interactions in selected binary solutions studied by X-ray methods

    NASA Astrophysics Data System (ADS)

    Drozdowski, Henryk; Romaniuk, Anna; Błaszczak, Zdzisław

    2013-12-01

    The results of X-ray structural studies of liquid chloroanisole C6H4OCH3Cl and 10% solutions of chloroanisole in 1,4-dimethylbenzene C8H10 are presented. It is the first paper on an X-ray diffraction study of the liquid solutions of chloroanisole. The X-ray measurements were made at 293 K for the scattering angle range 2Θ varying from 6° to 120°. Averaged scattered X-ray angular distributions I¯(S) were determined. The angular distributions of the intensity of X-ray scattered by 10% solutions of chloroanisole in 1,4-dimethylbenzene were compared to the angular distributions obtained for liquid ortho-, meta- and para-chloroanisole. The differential radial distribution functions of electron density 4πr∑j,knK[ρk(r)-ρ0] were numerically found using the Fourier analysis from a modified Warren, Krutter and Morningstar equation. To the maxima of DRDFs, interatomic and intermolecular distances were assigned. The use of short-wave radiation from an X-ray tube with a molybdenum anode permitted determination of the spheres of intermolecular ordering in the studied liquids and their solutions. The experimental results were used to plot models of the most highly probable mutual disposition of the molecules in liquid chloroanisole and their solutions. The benzene rings of two molecules are situated in parallel plane what results in antiparallel setting of the dipole moments of the chloroanisole molecules. X-ray structural analysis was applied to determine the packing coefficients of chloroanisole molecules. The results obtained in this paper confirm the specific structural properties of the solutions studied.

  16. Liquid-Structure Forces and Electrostatic Modulation of Biomolecular Interactions in Solution

    PubMed Central

    Hassan, Sergio A.

    2008-01-01

    Molecular interactions in solution are controlled by the bulk medium and by the forces originating in the structured region of the solvent close to the solutes. In this paper, a model of electrostatic and liquid-structure forces for dynamics simulations of biomolecules is presented. The model introduces information on the microscopic nature of the liquid in the vicinity of polar and charged groups and the associated non-pairwise character of the forces, thus improving upon conventional continuum representations. The solvent is treated as a polar and polarizable medium, with dielectric properties described by an inhomogeneous version of the Onsager theory. This treatment leads to an effective position-dependent dielectric permittivity that incorporates saturation effects of the electric field and the spatial variation of the liquid density. The non-pairwise additivity of the liquid-structure forces is represented by centers of force located at specific points in the liquid phase. These out-of-the-solute centers are positioned at the peaks of liquid density and exert local, external forces on the atoms of the solute. The density is calculated from a barometric law, using a Lennard-Jones-type solute–liquid effective interaction potential. The conceptual aspects of the model and its exact numerical solutions are discussed for single alkali and halide ions and for ion-pair interactions. The practical aspects of the model and the simplifications introduced for efficient computation of forces in molecular solutes are discussed in the context of polar and charged amino acid dimers. The model reproduces the contact and solvent-separated minima and the desolvation barriers of intermolecular potentials of mean force of amino acid dimers, as observed in atomistic dynamics simulations. Possible refinements based on an improved treatment of molecular correlations are discussed. PMID:17201447

  17. In vitro studies on the structure and properties of silk fibroin aqueous solutions in silkworm.

    PubMed

    Jin, Yuan; Hang, Yichun; Luo, Jie; Zhang, Yaopeng; Shao, Huili; Hu, Xuechao

    2013-11-01

    The spinning process of silkworm in vivo attracts great attentions. In this work, the structures and properties of the silk fibroin (SF) aqueous solutions from different divisions of silk glands of silkworms were investigated by using polarized microscope, rotational rheometer, Raman spectrometer and dynamic laser light scattering instrument. It was found that only the anterior (A) division and the anterior part of middle division (MA) of silk gland showed birefringence. With flowing from the posterior part (MP) to the MA part in the middle division of silk gland, the SF aqueous solutions was gradually transformed from random coil/α-helix to β-sheet conformation. Meantime, the elastic and viscous nature of the SF aqueous solution changed, and the mean diameter of SF aggregates increased from 118 nm to 331 nm. It could be concluded that the structures and properties of the SF aqueous solutions were gradually changed along the silk gland and the liquid crystal structure was initially formed in the MA part of silk gland. PMID:23994738

  18. Structural properties and adsorption capacity of holocellulose aerogels synthesized from an alkali hydroxide-urea solution

    NASA Astrophysics Data System (ADS)

    Kwon, Gu-Joong; Kim, Dae-Young; Hwang, Jae-Hyun; Kang, Joo-Hyon

    2014-05-01

    A tulip tree was used to synthesize a holocellulose aerogel from an aqueous alkali hydroxide-urea solution with the substitution of an organic solvent followed by freeze-drying. For comparison, the synthesized holocellulose aerogels were divided into two groups according to the source of the hydrogel, an upper suspended layer and a bottom concentrated layer of the centrifuged solution of cellulose and NaOH/urea solvents. We investigated the effects of the temperature of the pre-cooled NaOH/urea solution ( i.e., dissolution temperature) on the pore structure and the adsorption capacity of the holocellulose aerogel. A nano-fibrillar network structure of the holocellulose aerogel was observed, with little morphological difference in pore structure for different dissolution temperatures. Both micropores and mesopores were observed in the holocellulose aerogel. The specific surface area of the holocellulose aerogel was generally greater at lower dissolution temperatures. In a series of adsorption tests using methylene blue, the holocellulose aerogel showed the greatest adsorption capacity at the lowest dissolution temperature tested (-2°C). However, the dissolution temperature generally had little effect on the adsorption capacity. The holocellulose aerogel produced from the upper suspended layer of the centrifuged hydrogel solution showed a greater porosity and adsorption capacity than the one produced from the bottom concentrated layer. Overall, the aerogel made by utilizing a delignified tulip tree display a high surface area and a high adsorption property, indicating its possible application in eco-friendly adsorption materials.

  19. Implementation and performance of SIBYLS: a dual endstation small-angle X-ray scattering and macromolecular crystallography beamline at the Advanced Light Source.

    PubMed

    Classen, Scott; Hura, Greg L; Holton, James M; Rambo, Robert P; Rodic, Ivan; McGuire, Patrick J; Dyer, Kevin; Hammel, Michal; Meigs, George; Frankel, Kenneth A; Tainer, John A

    2013-02-01

    The SIBYLS beamline (12.3.1) of the Advanced Light Source at Lawrence Berkeley National Laboratory, supported by the US Department of Energy and the National Institutes of Health, is optimized for both small-angle X-ray scattering (SAXS) and macromolecular crystallography (MX), making it unique among the world's mostly SAXS or MX dedicated beamlines. Since SIBYLS was commissioned, assessments of the limitations and advantages of a combined SAXS and MX beamline have suggested new strategies for integration and optimal data collection methods and have led to additional hardware and software enhancements. Features described include a dual mode monochromator [containing both Si(111) crystals and Mo/B(4)C multilayer elements], rapid beamline optics conversion between SAXS and MX modes, active beam stabilization, sample-loading robotics, and mail-in and remote data collection. These features allow users to gain valuable insights from both dynamic solution scattering and high-resolution atomic diffraction experiments performed at a single synchrotron beamline. Key practical issues considered for data collection and analysis include radiation damage, structural ensembles, alternative conformers and flexibility. SIBYLS develops and applies efficient combined MX and SAXS methods that deliver high-impact results by providing robust cost-effective routes to connect structures to biology and by performing experiments that aid beamline designs for next generation light sources. PMID:23396808

  20. Implementation and performance of SIBYLS: a dual endstation small-angle X-ray scattering and macromolecular crystallography beamline at the Advanced Light Source

    PubMed Central

    Classen, Scott; Hura, Greg L.; Holton, James M.; Rambo, Robert P.; Rodic, Ivan; McGuire, Patrick J.; Dyer, Kevin; Hammel, Michal; Meigs, George; Frankel, Kenneth A.; Tainer, John A.

    2013-01-01

    The SIBYLS beamline (12.3.1) of the Advanced Light Source at Lawrence Berkeley National Laboratory, supported by the US Department of Energy and the National Institutes of Health, is optimized for both small-angle X-ray scattering (SAXS) and macromolecular crystallography (MX), making it unique among the world’s mostly SAXS or MX dedicated beamlines. Since SIBYLS was commissioned, assessments of the limitations and advantages of a combined SAXS and MX beamline have suggested new strategies for integration and optimal data collection methods and have led to additional hardware and software enhancements. Features described include a dual mode monochromator [containing both Si(111) crystals and Mo/B4C multilayer elements], rapid beamline optics conversion between SAXS and MX modes, active beam stabilization, sample-loading robotics, and mail-in and remote data collection. These features allow users to gain valuable insights from both dynamic solution scattering and high-resolution atomic diffraction experiments performed at a single synchrotron beamline. Key practical issues considered for data collection and analysis include radiation damage, structural ensembles, alternative conformers and flexibility. SIBYLS develops and applies efficient combined MX and SAXS methods that deliver high-impact results by providing robust cost-effective routes to connect structures to biology and by performing experiments that aid beamline designs for next generation light sources. PMID:23396808