Biomedical subjects

D L Dorset

Publications and source records attributed to D L Dorset.

At least 37 records · Page 2Linked to original sources

PubMed1995

The crystal structure of waxes.

Quantitative electron crystallographic studies have been carried out on epitaxially oriented multi-component waxes. Intensities from two paraffin-based samples, an artificial six-component medium wax (equimolar distribution of chain lengths) and a petroleum-based wax (Gaussian distribution of chain lengths) have been used to determine their crystal structures. As found earlier for binary paraffin solid solutions, differences in molecular volume are compensated by longitudinal molecular shifts within individual lamellae. Nevertheless, each lamellar surface must remain flat enough, and with enough crystallographic order intact, to nucleate the next lamella, thus accounting for the observed long-range correlation in these crystals. Recrystallized beeswax also has a layer packing somewhat similar to the paraffin waxes. However, in this case, the lamellar order is 'frustrated' so that a certain amount of 'nematically' ordered material must be present, spanning the nascent lamellar interfaces.

Animals↗

PubMed1995

Direct structure analysis in protein electron crystallography: crystallographic phases for halorhodopsin to 6-A resolution.

The crystal structure of halorhodopsin was determined in (centrosymmetric) projection to 6-A resolution by direct methods that use only the amplitudes of the electron diffraction pattern. A multisolution technique was used to generate initial 15-A-resolution basis sets, and after selection of the best phase set (by the closest match of magnitude of Eobs and magnitude of Ecalc), annealing of individual reflections was used to improve its accuracy. The Sayre equation was then used to expand the phase terms to 10 A, followed again by phase annealing. A final expansion with the Sayre equation enlarged this corrected phase set to 6 A. When the condition of density flatness was used to locate the best phase solution after each extension, a final structure could be observed that was quite similar to the one found earlier by analysis of electron micrographs.

Bacteriorhodopsins↗

PubMed1995

The Sayre equation in electron crystallography.

The Sayre equation was evaluated as a technique for phase refinement in electron crystallography. Atomic-resolution electron diffraction data from copper perchlorophthalocyanine were assigned phase values from the Fourier transforms of various experimental electron micrographs, including one at 2.3 A, containing errors due to lens astigmatism. In each case, an atomic-resolution structure could be found after Fourier refinement. In addition, it was possible to begin with a basis set derived from symbolic addition for phase extension. Such a source of phases was also found to be useful for extending zonal electron diffraction sets from six polymer crystals, even though there was considerable overlap of atomic positions in the projection down the chain axes. Other tests of the Sayre equation were made with zonal protein data sets (bacteriorhodopsin, halorhodopsin) to evaluate what difficulties are to be expected when direct phasing techniques are to be used in macromolecular electron crystallography. Comparison to known values indicated that the low-resolution range (e.g. to 6 A) was reasonably stable for phase extension from a 10-15 A resolution image. Only when a minimum in average intensity was approached (near 5 A) did the direct extension encounter serious difficulties. If this minimum was treated as a "phase node" to generate two possible solutions, a model more similar to the true phase set was found. In general, this rather simple convolutional technique for phase extension seems to be particularly suitable for a variety of electron crystallographic applications.

Crystallography↗

PubMed1995

Direct structure determination by electron crystallography: protein data sets.

Applications of the Sayre equation to the electron crystallographic analysis of protein structures are demonstrated. Starting with a lower-resolution basis phase set obtained from the Fourier-transform of an electron micrograph, it is possible, for example, to extend directly to the higher resolution of the electron diffraction pattern. Examples of such analyses include bacteriorhodopsin, halorhodopsin and the Omp F porin from the outer membrane of Escherichia coli. If a multisolution approach is taken, it may also be possible to carry out ab initio phase determinations, as demonstrated with low-resolution diffraction amplitudes from a negatively-stained membrane protein crystal.

Bacterial Outer Membrane Proteins↗

PubMed1994

Direct determination of layer packing for a phospholipid solid solution at 0.32-nm resolution.

Electron diffraction intensity data were collected from a 2:3 binary solid solution of two homologous phosphatidylethanolamines (1,2-dimyristoyl-sn-glycerophosphoethanolamine and 1,2-dipalmitoyl-sn-glycerophosphoethanolamine) epitaxially oriented by cocrystallization with naphthalene. The layer packing was determined directly by predicting the value of 12 of the 17 phases from sigma 1- and sigma 2-triplet invariants in space group P1. A reverse Fourier transform of the resulting potential maps provides estimates for three other phases and the two remaining ones were found by generating maps for the 2(2) = 4 possible phase combinations and then testing the smoothness of the potential profile of the hydrocarbon chain packing. The same phase solution can be found by translating a molecular model (based on the known x-ray crystal structure of a shorter homologue) past the unit cell origin. The solid solution is found to retain a stable polar group packing while the statistical occupancy of two terminal-chain carbons is expressed by a reduced potential profile at the nonpolar interface at the bilayer center.

Crystallography, X-Ray↗

PubMed1994

Electron crystallography of organic and biological molecules--techniques and comparison to X-ray crystallographic results.

In this review, it is shown how ab initio analyses of molecular crystal structures can be carried out with electron diffraction intensities and high-resolution, low-dose, electron micrographs, dispelling the long-held myth that such determinations are not possible. The results for small organics, polymethylene compounds, various polymers, and even intregral membrane proteins, are in good agreement with independent X-ray crystal structure determinations, which, however, were not needed to solve the crystallographic phase problem from the electron scattering data. The concept of electron crystallography benefits from the utility of electron micrographs for providing phase information, in addition to the standard 'direct' methods used nowadays by all crystallographers.

Crystallography↗

PubMed1994

Structural determination and packing analysis of a cholesteryl caprate/cholesteryl laurate solid solution.

This paper describes the X-ray crystal structure analysis of a cholesteryl ester solid solution, cholesteryl decanoate/cholesteryl laurate, grown from a bulk concentration with molar ratio 0.56/0.43. The unit cell is monoclinic with a = 12.969, b = 9.048, c = 31.137 A, and beta = 91.12 degrees and the space group P2(1) with Z = 4 (two molecules per asymmetric unit). The cell constants closely represent an average value of crystal parameters for the two pure components (hence, nearly corresponding to Vegard's law). Although the overall monolayer 1 lamellar packing is superficially similar to the earlier-studied cholesteryl undecanoate/cholesteryl laurate solid solution, a more partitioned distribution of acyl chains, i.e., a microfractionation corresponding to the observed nonideal phase behavior, is suggested. The behavior is similar to that found for n-paraffin binaries cooled below a binodal phase boundary. Although it cannot be detected conclusively in this determination (due to high thermal motion of terminal acyl chain atoms), the non-stoichiometric combination of components also requires some partial occupancy of atomic sites on the chain termini. This structural arrangement is contrasted with the alternative expression found earlier for the more ideal undecanoate/laurate solid solution, i.e., random co-packing leading to fractional atomic occupancy in an average laurate structure. The final weighted R factor for 4578 reflections is 0.138.

Cholesterol Esters↗

PubMed1992

Binary phase behavior of angiotoxic oxidized cholesterols with cholesterol.

Binary phase diagrams of oxidized cholesterols with cholesterol were constructed in order to find a possible physical mechanism for these cytotoxic sterols as disruptors of cell membranes. Two compounds, 25-hydroxycholesterol and 7-ketocholesterol, behave similarly in solids with cholesterol, i.e., continuous solubility can be demonstrated over all concentrations with some tendency for individual molecular species to cluster in the liquid state. However, the two compounds have greatly different solubilities in phospholipid bilayers. The interaction of cholesterol with cholestane 3 beta, 5 alpha,6 beta-triol differs from that of the other two binaries since these molecules form a eutectic. This compound has been found to be soluble in phospholipid bilayers. Hence, from available evidence, the three oxidized cholesterols must employ different mechanisms for disruption of cell membranes, depending on which membrane component can most easily solubilize each.

Calorimetry, Differential Scanning↗

PubMed1992

Dynamical scattering and electron crystallography--Ab initio structure analysis of copper perbromophthalocyanine.

Electron diffraction intensity data were collected at 1200 kV from thin epitaxially oriented crystals of copper perbromophthalocyanine (C32Br16CuN8) in a projection down molecular columns. Measured cell constants for the projection with cmm symmetry are d100 = 17.88 (9), b = 26.46 (15) A. The structure was determined by Fourier refinement after three heavy-atom positions were identified in an initial potential map. In addition to the copper and halogens, all light-atom positions were found. Although the final R value for all data is 0.41, n-beam dynamical calculations for crystal thicknesses corresponding to the estimated sample dimension account for the observed amplitudes that deviate most from their kinematical values.

Crystallography↗

PubMed1992

Molecular interactions in binary solids: crystal structure of a cholesteryl ester solid solution.

The x-ray crystal structure analysis of a cholesteryl ester solid solution, cholesteryl undecanoate/cholesteryl laurate in a molar ratio 0.52/0.48, is described. The unit cell is monoclinic with a = 13.005(2) A, b = 9.005(1) A, c = 31.421(3) A, and beta = 90.82(1) degrees and the space group P2(1) with Z = 4 (two molecules per asymmetric unit). Thus, the d001 spacing is almost the value predicted by Vegard's law from the values for the pure compounds. The room-temperature crystal structure is very much like that of cholesteryl laurate monolayer I packing, in the form where the esterified fatty acid chains are fully extended, with no salient sign of conformational disorder seen in this binary solid. The final R factor for 6571 observed reflections is 0.097.

Cholesterol Esters↗

PubMed1991

Electron crystallography of linear polymers: direct structure analysis of poly(epsilon-carprolactone).

Three-dimensional electron diffraction intensity data from poly(epsilon-caprolactone) are used for an ab initio phase determination based on the probabilistic estimate of three- and four-phase structure invariants in space group P212121. From the initial phase set of 30 reflections (two of which are incorrect) of the 47 in the total data set, computed electrostatic potential maps produce a recognizable structure that can be refined by Fourier techniques and atomic displacement. The final structure (R = 0.21), which is very similar to one found in an earlier fiber x-ray study, shows that the polymer chain must adopt a nonplanar conformation around the ester linkage.

Journal Article↗

PubMed1991

Electron crystallography at atomic resolution: the structure of the odd-chain paraffin n-tritriacontane.

The crystal structure of the odd-chain paraffin, n-tritriacontane, nC33H68, is determined directly by using low-dose electron microscope images and electron diffraction intensity data from epitaxially grown microcrystals. Phases of the most intense "polyethylene" reflections are determined from triplet structure-invariant relationships often used in X-ray crystallography. Low-dose electron microscopic images provide phases of the low-angle "lamellar" reflections and these can be used with one-dimensional structure-invariant relationships to determine other phases on the 00l reciprocal row. The phase set is sufficient to calculate an electrostatic potential map which is directly interpretable as a structure image at atomic resolution.

Crystallography↗

PubMed1991

Electron crystallography at atomic resolution: ab initio structure analysis of copper perchlorophthalocyanine.

High-voltage (1200 kV) electron diffraction intensities from approximately 100 A thick crystals of copper perchlorophthalocyanine are used to determine the molecular packing at atomic resolution, thus greatly exceeding the structure detail observed by electron microscopy. Initial crystallographic phases were determined by direct methods often used in X-ray crystallography, i.e., locating the positions of heavy (Cl and Cu) atoms in the structure. All other atom positions were found in subsequent Fourier refinement (final R = 0.28). Calculated bond distances and angles are similar to those found in the earlier X-ray crystal structure of the unchlorinated parent compound.

Copper↗

PubMed1991

Direct determination of crystallographic phases for diffraction data from lipid bilayers. I. Reliability and phase refinement.

Direct analysis of lipid lamellar packing based on the probabilistic estimate of sigma 1- and sigma 2-triplet phase invariants is evaluated here for a large variety of bilayer structures than examined in an original study of this problem (Dorset, D.L., 1990. Biophys. J. 58:1077-1087). Using x-ray crystal structures of five phospholipids, three glycerides and two cerebrosides, lamellar diffraction data were generated at the approximately 3 A resolution often found experimentally from oriented multilayers. For structures where no significant density occurs at the unit cell origin, the ab initio phase determination is successful for six of the ten structures. A seventh structure can be solved if a limited set of sigma 2-triples are used to determine the initial phase set based on the hierarchy of the A2 values. Bilayers, e.g., with solvent at the origin, can be analyzed if a modified criterion for accepting phase estimates for sigma 1-triples is used, as suggested by the distribution of normalized structure factors and the number of probable single-valued phase domains. In all cases, partial phase determinations can be refined effectively by density modification ("flattening") of the hydrocarbon region in real space. A figure of merit suggested by Luzzati et al. (Luzzati, V., A. Tardieu, and D. Taupin. 1972. J. Mol. Biol. 64:269-286) used to evaluate the success of such refinement can be supplemented by an evaluation of density smoothness, which can also detect the presence of near structure homomorphs not identified by the former test for density flatness.

Lipid Bilayers↗

PubMed1991

Direct determination of crystallographic phases for diffraction data from lipid bilayers. II. Refinement of phospholipid structures.

Using a systematic approach for the acceptance of crystallographic phase assignment, based on the evaluation of triplet structure invariants, electron and x-ray diffraction data from phospholipid multilamellar arrays are analyzed by direct methods. After calculation of Fourier maps with a partial set of phased structure factor magnitudes, the structure is refined in real space by flattening of the hydrocarbon region of the bilayer and an optimal solution is sought either by the calculation of [delta rho 4] suggested by Luzzati, where rho is the structure density or by a test of density smoothness [magnitude of delta rho/ delta r magnitude of], where r positions are located along the normal to the lamellar surface. Reanalyses of previously determined structures sometimes lead to new conclusions (e.g., a possible similarity of the electron density profile for DL-DMPE and L-DMPE, and a clear indication of the fatty acid adduct in the mixed L-DPPC/palmitic acid bilayer). Because of presumed secondary scattering perturbations (primarily to the least intense reflections), the refinements of the electron diffraction intensities are less easily evaluated than those carried out with x-ray diffraction data.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1990

Eutectic interactions in binary systems containing cholesterol, cholesteryl esters and triacylglycerols.

Binary phase behavior of saturated cholesterol esters with trilaurin or cholesterol, and cholesterol with trilaurin, is studied. The existence of specific molecular interactions is detected by comparing the liquidus curve of the eutectic with ideal theory of freezing point depression and correcting the theoretical curve with the Bragg-Williams model when necessary. X-ray data indicate that all eutectic solids are nearly totally fractionated. The phase diagrams are sometimes well-explained by ideal solution theory indicating that polar interactions (e.g., the hydrogen bonding of cholesterol) are much less important than van der Waals interactions between neighboring molecules. However, the hydrogen bonding networks of cholesterol can lead to nonideal solution behavior with other lipids, a phenomenon consistent with previous observations on simpler molecular binaries. An observed nonideal solution behavior of triacylglycerol with cholesterol esters, on the other hand, is unexpected since significant polar interactions are expected to be 'buried' in the predominant nonpolar volume of the molecules involved.

Chemical Phenomena↗

PubMed1990

Lamellar packing of a chiral N,N-dimethylphosphatidylethanolamine: electron diffraction. Evidence for a lecithin-type headgroup conformation.

Lamellar electron diffraction intensity data from epitaxially crystallized 1,2-dipalmitoyl-sn-glycerophospho-N,N-dimethylethanolamine were used to determine the layer packing in order to compare the chiral structure to the crystal structure of a racemic homologue. After finding the chain orientation, the structure was determined by interpretation of the Patterson function, followed by independent crystallographic phase assignments with conventional direct methods (use of three phase structure invariants). The phase determination was verified by a translational search with a molecular model based on a similar lecithin structure. The final R-value is 0.29, and this is lowered to 0.18 after a correction is made for incoherent multiple electron scattering. The layer packing is found to be very much like that of a diacyl phosphatidylcholine with the N,N-dimethylethanolamine moiety parallel to the bilayer surface rather than the perpendicular arrangement of headgroups involved in an interdigitated layer, as seen for racemic homolog.

Crystallization↗

PubMed1990

Anomeric exchange and the structure of n-alkyl D-glucopyranosides. A study of binary phase behavior.

X-Ray diffraction and calorimetric data from the alpha and beta anomers of n-alkyl D-glucopyranosides are analyzed to describe the molecular packing and co-solubility in the crystalline and liquid-crystalline phases. In the smectogenic chain-length series, the beta-glucosides are co-soluble, with almost ideal mixing in the crystalline and meso-phases for chain-length differences of two carbon atoms. The smectic phases of octyl alpha- and beta-glucosides are also co-soluble and a time sequence of phase diagrams (as well as lamellar X-ray data), as the solid obtained from the cooled melt is equilibrated, indicate that a metastable co-crystalline phase may exist until the respective hydrogen-bonding schemes are established. Lamellar spacings from a homologous series of the beta anomers indicate that both the crystal structure and smectic-layer packings involve bilayer stacking of the molecules, a result that is difficult to reconcile with the respective surface-area requirements of the molecular acyl chain and sugar moieties.

Calorimetry, Differential Scanning↗