Density Modification Method Research Articles

Remediation of DNAPL-contaminated aquifers using density modification method with colloidal liquid aphrons

Entrapped and pooled dense non-aqueous phase liquid (DNAPL) often persists in aquifers and serves as a long-term source of groundwater contamination. Despite vigorous research efforts over the last two decades, all current DNAPL pool remediation strategies suffer from a combination of inefficiency, enhanced risk of contaminant spreading due to uncontrolled mobilization, and or high treatment costs. In the present contribution, we report a novel strategy that makes the density of the representative DNAPL of trichloroethene (TCE) reduction utilizing colloidal liquid aphrons (CLAs) in the remediation process. The effects of the electrolyte (AlCl 3) concentrations, the volume fraction of the dispersed oil phase ( n-octane) and temperature on the demulsification behavior of CLA dispersion were discussed. Steady-state flow and dynamic viscoelasticity were measured by using a controlled stress rheometer HAAKE RS 6000 with cone-plate geometry in an attempt to reveal the rheological properties of CLAs. A series of batch experiments was conducted to evaluate the recovery efficiency of DNAPL from sand pack under different volumes, flow rates of CLAs and surfactant (Tergitol 15-S-9) flushing solutions. Experimental results suggest that CLAs can be destabilized by a low concentration of Al 3+ in the continuous phase. The flow behavior of CLAs presents the typical non-Newtonian liquids with strong shear thinning properties and it could be well described by the Hershel–Bulkley model. Above the phase volume ratio (PVR = V org/ V aq) of 8, the systems display viscoelastic behavior. Relative to water, the density reversal of TCE occurs at an electrolyte concentration of 0.05 M, which effectively prevents TCE downward migration during surfactant flushing. As much as 94% of the TCE entrapped in the sand column is removed under the conditions of amounts of 1 PV (pore volume) for CLAs, 4 PV and 1.5 mL/min for Tergitol solution. This approach holds great promise for manipulating DNAPL densities prior to or during remediation processes.

Comparative Structural Biology of Eubacterial and Archaeal Oligosaccharyltransferases

Oligosaccharyltransferase (OST) catalyzes the transfer of an oligosaccharide from a lipid donor to an asparagine residue in nascent polypeptide chains. In the bacterium Campylobacter jejuni, a single-subunit membrane protein, PglB, catalyzes N-glycosylation. We report the 2.8 A resolution crystal structure of the C-terminal globular domain of PglB and its comparison with the previously determined structure from the archaeon Pyrococcus AglB. The two distantly related oligosaccharyltransferases share unexpected structural similarity beyond that expected from the sequence comparison. The common architecture of the putative catalytic sites revealed a new catalytic motif in PglB. Site-directed mutagenesis analyses confirmed the contribution of this motif to the catalytic function. Bacterial PglB and archaeal AglB constitute a protein family of the catalytic subunit of OST along with STT3 from eukaryotes. A structure-aided multiple sequence alignment of the STT3/PglB/AglB protein family revealed three types of OST catalytic centers. This novel classification will provide a useful framework for understanding the enzymatic properties of the OST enzymes from Eukarya, Archaea, and Bacteria.

Comparison of phasing methods for sulfur-SAD using in-house chromium radiation: case studies for standard proteins and a 69 kDa protein

Phasing of the crystal structures of four standard proteins (lysozyme, trypsin, glucose isomerase and thaumatin) and a novel 69 kDa protein from Thermus thermophilus, TT0570, was performed using the single-wavelength anomalous diffraction of S atoms intrinsically present within the native protein molecules. To utilize the sulfur anomalous diffraction, the data sets were collected using the loopless data-collection method with chromium Kalpha X-rays of wavelength 2.29 A. Three phasing methods, MLPHARE, SHARP and OASIS-2004, were tested in combination with the DM or SOLOMON density-modification method. The results showed that the solvent contents are still an important factor for phasing with the S-SAD method, even when longer wavelength Cr Kalpha radiation is used. Of the three procedures, the improved direct phasing of OASIS-2004 with its implemented fragment feedback to the direct-method probability calculation gave the best results in determining the initial phases. For all five proteins, almost the entire models could be built automatically.

Ab initiocrystal structure determination of spherical viruses that exhibit a centrosymmetric location in the unit cell

Ab initio phasing by non-crystallographic symmetry averaging coupled with solvent flattening has previously been used to determine the structure of canine parvovirus. As CPV particles were located at general positions, initial phases were generated with a spherical shell deviating from the centre of symmetry. In many virus crystals, the viral particles are located at positions with a centre of symmetry in the unit cell. Thus, the initial phases calculated with a spherical shell model have a centre of symmetry. The inherent difficulty in structural determination was breaking the centre of symmetry in the initial phase. The centric nature of the initial phases of the rice dwarf virus crystal, however, was successfully broken at low resolution by iteration of the density-modification method described by Tsao et al. (1992). In this study, ab initio phasing was tested for seven viruses ranging from 82 to 344 A in radius. Although the crystal structures of the initial spherical shell models for each virus had a centre of symmetry, the centric natures of the initial phases were successfully broken by non-crystallographic symmetry averaging coupled with solvent flattening at a low resolution; these phases were then successfully extended to high-resolution measurements. A novel procedure of ab initio phasing for spherical virus crystals is proposed.

Multi-wavelength anomalous diffraction method for I and Xe atoms using ultra-high-energy X-rays from SPring-8

The first successful multi-wavelength anomalous diffraction (MAD) experiments using ultra-high-energy X-rays (∼35 keV) were performed for iodine and xenon derivatives of hen egg-white lysozyme crystals. The beamline BL41XU of SPring-8 enabled the collection of high-quality MAD data, which led to the calculation of anomalous or dispersive difference Patterson maps that determined the positions of iodine and xenon atoms. The electron density maps obtained by the density modification method for both cases proved to be of sufficient quality for building molecular models. I-MAD and Xe-MAD phasing are now available at SPring-8, and the utilization of ultra-high-energy X-rays will make a significant contribution to the solution of the phase problem in protein crystallography.

Ab Initio Structure Solution of Icosahedral Cd-Yb Quasicrystals by a Density Modification Method

The six-dimensional (6D) low density elimination method has been applied to solve an icosahedral Cd-Yb quasicrystal. It shows that i-Cd-Yb is composed of similar atomic clusters that exist in the cubic approximant crystals, but the inner most shell is highly disordered. A construction of the 6D cluster model of i-Cd-Yb is outlined.

Multidimensional Histograms for Density Modification

Publisher Summary Density modification improves the quality of an approximate electron-density map by imposing physical constraints based on some conserved features of the correct electron density map. Density modification methods generally require an initial map with substantial phase information. In most cases, these phases are obtained from multiple isomorphous replacement (MIR) or multiwavelength anomalous dispersion (MAD), and these phases are of pivotal importance when the experimental source of phase information is bimodal as it is in single-wavelength anomalous scattering (SAD) and single isomorphous replacement methods (SIR). The most commonly used density modification method is solvent flattening that exploits the observation that the solvent region of an electron density map is featureless at medium resolution because of the high thermal motion and disorder of the solvent molecules. Xiang and Carter proposed extending the density histogram to include relationships between neighboring density values via multidimensional histograms defined as joint probabilities of the density values and their higher-order derivatives. In the conventional one-dimensional (1D) electron density histogram matching method, a one-to-one mapping is made on the original electron density to the new electron density so that the density histogram of the modified map matches that of the ideal histogram. The two-dimensional (2D) histogram specifies not only the probability of the electron density value for a given grid point in the map but also the local environment around that grid point as reflected by the density gradients that arise from chemical bonding. The strategy of using alternating 1D histogram matching to achieve 2D histogram matching can be generalized to the matching of higher dimension histograms.

General concepts underlying ACORN, a computer program for the solution of protein structures

ACORN has been shown to be a successful program for the solution of small and medium-size protein structures, when data at atomic resolution or better are available. At the heart of ACORN lies a density modification method called DDM (Dynamic Density Modification). DDM operates on any density map containing a minimum of information. Its action is to progressively enhance positive features of the map and to reduce, at the same time, undesired noise and bias towards very small values. This is indeed a feature claimed by other density modification procedures. The effectiveness of the procedure arises from the fact that the shape of DDM's modification curve is one of a restricted class of functions, whose properties will be described. The importance of a proper weighting scheme will also be considered. Tests on a few structures will help to clarify the theoretical concepts involved as well as to make clear the potential applications and limits of the program.

Ab initio structure determination of quasicrystals by density modification method

A novel density modification method has been applied to phase reconstruction of X-ray single crystal data of quasicrystals. This is based on the low electron density elimination (LDE) method that is an ab initio structure determination method for crystal in the real space. The location, size and shape of the occupation domains for quasicrystals in the n-dimensional ( nD) unit cell are obtained without any model structure. Therefore, the LDE method can be a substitute for the direct method for quasicrystals as nD crystals. The structure solution can easily be found in the trial sets (normally 100 sets) and is confirmed by the subsequent analysis. The result is used as a crude starting model for constructing a detailed structure model of quasicrystals. The solutions of several quasicrystals (i-AlPdMn, i-ZnMgHo and i-CdYb) are exemplified. The reliability and limitations of the algorithm to retrieve the quasicrystalline structures will also be discussed.

密度修正法による準結晶構造解析の研究

A novel density modification method has been applied to phase reconstruction of X-ray single crystal data of quasicrystals. This is based on the low density elimination (LDE) method that is an ab initio structure determination method for crystals in the real space. The location, size andshape of the occupation domains of quasicrystals in the n-dimensional (nD) unit cell are obtained without a prioi model structure. Therefore, the LDE method can be a substitute for the direct method for quasicrystals that are described as nD crystals. The structure solution can easily be found in the trial sets (normally 100 sets) and is confirmed by the subsequent analysis. The result is used as a starting model for constructing a detailed structure model of quasicrystals. The solution of several qusicrystalline structures including a model quasicrystal structure are exemplified.

Structure of icosahedral Zn-Mg-Ho quasicrystals determined by a density modification method

The structure of icosahedral Zn60Mg31Ho9 quasicrystal has been determined, by means of ab initio structure determination, by applying a novel density modification method (low density elimination method) within a framework of 6D description. The suggested Ho sites in the 3D structure obtained as a section of resulting 6D electron density are consistent with the results of magnetic diffuse scattering.

Ab initio structure solutions of quasicrystals by a density modification method

A unique and powerful density modification method for solving multi-dimensional crystal structures has been developed. The method is an extension of the Low Density Elimination (LDE) method1,2,3. The LDE method was made applicable to multi-dimensional crystal structures and applied to several quasicrystals including an unknown structure. Tests on the X-ray single crystal data of icosahedral AlPdMn4 and synthetic data of decagonal AlNiCo quasicrystals suggests that the multi-dimensional LDE method can easily solve quasicrystal structures starting from completely random phases. The procedure has also been applied to an unknown structure of icosahedral ZnMgHo quasicrystal. Multi-solution strategy was employed and 28 resultant maps clearly showed the positions and shapes of occupation domains. We were convinced by analyzing the map that the 28 results were the solutions.

Ab initio structure determination of icosahedral Zn-Mg-Ho quasicrystals by density modification method.

A novel density modification method is applied for the first time to phase reconstruction of x-ray single crystal data of quasicrystals. The structure of icosahedral Zn-Mg-Ho quasicrystals has been determined by means of this ab initio structure determination within a framework of a 6D description. The location, size, and shape of the occupation domains are deduced. The suggested Ho sites in the 3D structure are consistent with the results of magnetic diffuse scattering [T. J. Sato et al., Phys. Rev. Lett. 81, 2364 (1998)].

Ab initio phasing using molecular envelope from solution X-ray scattering.

Solving the phase problem is the crucial and quite often the most difficult and time-consuming step in crystallographic structure determination. The traditional methods of isomorphous replacement (MIR or SIR) and molecular replacement require the availability of an isomorphous heavy-atom derivative or the structure of a homologous protein, respectively. Here, a method is presented which utilizes the low-resolution molecular shape determined from solution X-ray scattering data for the molecular search. The molecular shape of a protein is an important structural property and can be determined directly by the small-angle scattering technique. The idea of locating this molecular shape in the crystallographic unit cell has been tested with experimental diffraction data from nitrite reductase (NiR). The conventional Patterson search proved to be unsuccessful, as the intra-envelope vectors are uniformly distributed and do not match those of intra-molecular (atom-to-atom) vectors. A direct real-space search for orientation and translation was then performed. A self-rotation function using 2.8 A crystallographic data yielded the polar angles of the non-crystallographic threefold axis. Knowledge of the orientation of this axis reduces the potential six-dimensional search to four (Eulerian angle gamma and three translational parameters). The direct four-dimensional search within the unit cell produced a clear solution. The electron-density map based on this solution agrees well with the known structure, and the phase error calculated from the map was 61 degrees within 20 A resolution. It is anticipated that the low-resolution envelope can be used as a starting model for phase extension by the maximum-entropy and density-modification method.

The two-dimensional histogram as a constraint for protein phase improvement.

The joint distribution of electron density and its gradient in a protein electron-density map was examined. This joint distribution was represented by a two-dimensional histogram (2D histogram) of electron-density values and the modulus of the gradient. 16 structures representing distinct protein-fold families were selected to study the dependence of the 2D histogram on resolution, overall temperature factor, structural conformation and phase error. The similarity between the histograms for a pair of structures was measured by correlation coefficient, and the residual provided a measure of the difference. The 2D histogram was found to vary with resolution and overall temperature factor, but was found to be insensitive to structure conformation. The average correlation coefficient between pairs of 2D histograms at three different resolutions examined was 0.90 with a standard deviation of 0.04. The average residual for the same condition was 0.13 with a standard deviation of 0.03. The 2D histogram was also found to be sensitive to phase error. The average correlation coefficient and residual between 2D histograms with 10 degrees phase difference are 0.71 and 0.18, respectively. The variation of the 2D histogram resulting from structure-conformation changes was estimated to be equivalent to that of a 4 degrees phase error. This establishes the minimal phase error that a 2D histogram-matching method could achieve. The conservation of the 2D histogram with respect to structure conformation enables the prediction of the ideal 2D histogram for unknown structures. The sensitivity of the 2D histogram to phase error suggests that it could be used as a target for the density-modification method and also could be used as a figure of merit for phase selection in ab initio phasing.

Procedure to calculate the molecular envelope from a partial model.

Density-modification algorithms have gained, in recent years, a widespread use in the early stages of protein structure determination, especially in combination with the single isomorphous replacement, the multiple isomorphous replacement and the multiple-wavelength anomalous dispersion methods, where density modification usually leads to a significant improvement in the quality and interpretability of the initial electron-density map. The current computer programs which are used to perform this task combine several approaches, an important component of which is the solvent-flattening procedure. The latter procedure depends crucially on the correct determination of the molecular envelope. The solvent-flattening procedure has also been applied to the electron-density maps calculated from partial mod- els obtained from the molecular replacement method. In such case the envelope calculated in the standard way does not always encompass entirely the missing part. It has been found that the standard application of the density-modification method (as implemented by programs SQUASH and DM) to a map calculated from a molecular replacement model containing approximately 60% of the molecule, led to little improvement in the map interpretability. Here, it is shown that a significant improvement of the map can be achieved when a better envelope is used in the procedure. Various methods of calculating the molecular envelope have been evaluated, the effect of the shape of the envelope on the modified electron-density map has been investigated and an improved procedure to calculate the envelope from a partial molecular replacement model is proposed.

Use of Harker sections in a density modification method for direct solutions of crystal structures

Use of Harker sections in a density modification method for direct solutions of crystal structures

Deconvoluting Laue multiple diffraction spots by the density modification method

Deconvoluting Laue multiple diffraction spots by the density modification method

A Metropolis-like algorithm to improve phases: some preliminary results

A procedure to improve phases by a Metropolis-like algorithm (MLA) in reciprocal space is presented. It uses a density modification method (DMM) in a MLA to evaluate progress and to suggest changes to phase angles. First, in the neighborhood of the trial values derived from a DMM, new options for the phase angles are generated. Then, a DMM is used to evaluate and improve compliance with the rule of nonnegativity of direct methods. The procedure of DMM~ phase change~ DMM-~ phase change is iterated several times. For a centrosymmetric case in which phases may not be changed by a DMM, compliance with the non-negativity rule is judged by the ratio of observed to calculated structure-factor amplitudes, and new options for the phase angles are generated by adding 180 °. For a non-centrosymmetric case both the amplitude and the phase values of the structure factors before and after a DMM may differ. However, only the phase values are used to judge the compliance with the non-negativity rule, and to decide phase shifts in a subsequent phase change step. This procedure is shown to improve the phases determined ab initio by the consistent electron density approach [Bhat (1984). Acta Cryst. A40, C15; (1985). Am. Crystallogr. Assoc. Annu. Meet., Stanford, California. Abstr. HI]. The mean phase errors for the phases obtained by the procedure proposed here ranged from 55 to 25 ° in different tests, significantly better than those obtained from a DMM under similar conditions.

A density-modification method for the improvement of poorly resolved protein electron-density maps

An efficient computer procedure has been developed for the extraction of regions of contiguous, well-connected high density from a three-dimensional electron-density map. This procedure may be used to generate an extended model volume, from a smaller volume based on a starting atomic model. The starting model may, for example, only include main-chain atoms for a protein, or may omit uninterpretable segments of chain. The procedure may also be used to extract regions of contiguous density where no model exists. The extended model volume is used to produce a properly scaled model electron density. Calculated structure factors are obtained from the scaled model electron density by fast Fourier transform, and combined, with appropriate weights, with the existing phase information to give improved phase angles. Calculation of a new electron-density map with the new phase angles initiates the next step of a cyclic procedure which converges rapidly. The procedure has been applied to the structure determination of tyrosyl-tRNA synthetase. It has led to identification of most of the available amino-acid sequence in the electron density, and a revised tracing of the main polypeptide chain. Evidence for improvement in phase angles is obtained from electron-density difference maps for substrate and inhibitor binding, in which a reduction in background density is observed.