Calibration of ring-current effects in proteins and nucleic acids

Summary Density functional chemical shielding calculations are reported for methane molecules placed in a variety of positions near aromatic rings of the type found in proteins and nucleic acids. The results are compared to empirical formulas that relate these intermolecular shielding effects to magnetic anisotropy (ring-current) effects and to electrostatic polarization of the C–H bonds. Good agreement is found between the empirical formulas and the quantum chemistry results, allowing a reassessment of the ring-current intensity factors for aromatic amino acids and nucleic acid bases. Electrostatic interactions contribute significantly to the computed chemical shift dispersion. Prospects for using this information in the analysis of chemical shifts in proteins and nucleic acids are discussed.     

Specificity in DNA interactions: an nmr investigation of the interaction of propidium with oligodeoxyribonucleotides containing normal and G-T base pairs

The interaction of propidium with three self-complementary oligodeoxyribonucleotides has been investigated by ¹H- (base-pair imino proton assigned by 1D NOE and saturation transfer methods) and ³¹P-nmr as a function of ratio of propidium to oligomer (from zero to saturation) and temperature. The three oligomers are dTATATGCGCATATA (1), dTATATGTGCATATA (2), which has the same sequence as 1 except for the mismatched base pair at position 7, and dTATGTGCATA (3), which is a shortened version of 2. The imino proton chemical-shift changes of 1 on titration with propidium can be explained by the effects of the ring-current anisotropy of propidium at intercalation (3.4 Å) and next-neighbor sites (6.8 Å). The results indicate that propidium binds with neighbor exclusion but with no significant specificity for any intercalation site in the sequence of 1. The addition of propidium to 1 results in general downfield shifts of all ³¹P signals, as expected for a nonspecific intercalator. Imino and ³¹P-nmr spectra for 2 indicate that this oligomer forms a hydrogen-bonded G · T base pair at position 7 with little change in base pairing and stacking of base pairs 1–6 compared to 1. The results for addition of propidium to 2 and 3 are quite different than with 1. At low ratio only secondary shifts (6.8 Å) are seen for the G and T imino protons of base-pair 7 on addition of propidium. At higher ratios of propidium, the signals for these G and T protons are lost in 2 and severely broadened in 3, even at low temperature. The other potential intercalation sites in 2 and 3 appear to bind propidium strongly and without significant specificity as with 1. ³¹P spectra of 2 in the presence of propidium show the expected downfield shifts and broadening. Thus, the minor differences in local helix geometry in 1, and in 2 and 3, away from the G · T base pair do not significantly affect propidium intercalation specificity. Having one or two G · T base pairs at a site, however, makes intercalation in the standard manner significantly less favorable.     

Electrical and hydrodynamical properties of polypeptides in solution. I. Poly(L-glutamic acid) in methanol-water mixtures

The electric birefringence of poly(L -glutamic acid) (PLGA) in methanol–water mixtures has been measured by the use of the rectangular pulse technique at 25°C. The permanent dipole moment, the anisotropy of electrical polarizability, and the optical anisotropy factor of PLGA in solution were obtained from the dependence of the steady-state birefringence on the electric field strength. Further, the mean length of PLGA in solution was calculated by a parameter method developed for analyzing the decay curve of electric birefringence. The permanent dipole moment per unit length obtained from these studies was 2.9₆, 2.4₈, 2.3₀, 2.6₆ D/Å in pure methanol, 10, 30, and 50 vol-% water, respectively. The increase of water content caused the decrease of the mean length and broadened the length distribution of PLGA. These results are discussed in relation to the viscosity and the electrical conductivity of PLGA solutions.     

Metal—phenoxyalkanoic acid interactions. Part 14. The crystal and molecular structures of diaquabis(4-fluorophenoxyacetato)copper(II) bis(4-fluorophenoxyacetic acid)dihydrate and tetra-μ-(4-fluorophenoxyacetato-0,0′)-bis[(2-aminopyrimidine)copper(II)]

The crystal structures of two copper(II) complexes of 4-fluorophenoxyacetic acid (4-FPAH) have been determined by X-ray diffraction. [Cu(4-FPA)₂(H₂O)₂]·2(4-FPAH)·2H₂O (1) is triclinic, space group P1 with Z = 1 in a cell of dimensions a = 14.808(2), b = 9.832(2), c = 6.847(2) Å, α = 87.77(2), β = 98.41(2), γ = 112.33(2)° and was refined to a residual of 0.038 for 1697 ‘observed’ reflections. The coordination sphere in this complex is tetragonally distorted octahedral comprising two waters [CuO, 1.940(3) Å], two unidentate carboxylate oxygens [CuO, 1.942(2) Å] and two ether oxygens [CuO, 2.471(2) Å]. Two adducted [4-FPAH] acid molecules are linked to the un-coordinated oxygens of the acid ligands by hydrogen bonds [2.547(4) Å]. [Cu₂(4-FPA)₄(2-aminopyrimidine)₂] (2) is triclinic, space group P1 with Z = 1 in a cell of dimensions a = 12.688(2), b = 11.422(2), c = 7.951(1) Å, α = 78.74(1), β = 107.51(1), γ = 75.78(1)°, and was refined to a residual of 0.042 for 2683 ‘observed’ reflections. (2) is a centrosymmetric tetracarboxylate bridged dimer with four similar CuO (equatorial) distances [1.967–1.987 Å; 1.977(3) Å mean] and the axial position occupied by the hetero nitrogen of the 2-aminopyrimidine ligand [CuN, 2.176(3) Å]. The Cu---Cu separation is 2.710(1) Å. Crystal data are also presented which confirm the isostructurality of complex (2) with [Cu₂(phenoxyacetate)₄(2-aminopyrimidine)₂], the Co^II, Mg^II and Mn^II4-fluorophenoxyacetate complexes with their phenoxyacetic and 4-chlorophenoxyacetic acid analogues, and of Cd^II4-fluorophenoxyacetate with Cd^II and Zn^II phenoxyacetates.     

Structural components of alginic acid. II. The crystalline structure of poly-alpha-L-guluronic acid. Results of x-ray diffraction and polarized infrared studies

A structural investigation of the marine algal polysaccharide poly-α-L -guluronic acid is described. The molecular chains consist of 1 → 4 diaxially linked L -guluronic acid residues in the 1C chair conformation and are stabilized in a twofold helix conformation by an intra-molecular O(2)H … O(6)D hydrogen-bond. The X-ray fiber diffraction photograph has been indexed to an orthorhombic unit cell in which a = 8.6 Å, b (fiber axis) = 8.7 Å, c = 10.7 Å. A structure corresponding to the space group P2₁2₁2₁ is proposed, in which all intermolecular hydrogen bonds interact with water molecules and in which all oxygen atoms except for the inaccessible bridge oxygens are involed. The relationship between the shape and structure of the polyguluronic acid molecule and its biological function is discussed.     

Structural and mass spectral studies on some multiply bonded diosmium tetracarboxylate compounds

The compounds Os₂(O₂CCH₃)₄Cl₂,1 and Os(O₂CC₂H₅)₄Cl₂, 2, have been structurally characterized. Both compounds crystallize in space group P2₁/n. For 1 the unit cell parameters are a = 6.546(1) Å, b = 8.950(1) Å, c = 12.533(1) Å, β = 90.17(1)° and Z = 2; for 2 they are a = 6.792(2) Å, b = 10.519(3) Å, c = 13.372(4) Å, β = 89.27(3)° and Z = 2. Both molecules have approximate D_4th symmetry with important dimensions as follows: for 1, Os≡Os 2.314(1) Å, OsCl 2.448(2) Å; for 2, Os≡Os 2.316(2) Å, OsCl 2.430(5) Å. The mass spectra of these compounds as well as that of Os₂(O₂CC₃H₇)₄Cl₂,3, are reported and discussed.     

Structural components of alginic acid. I. The crystalline structure of poly-beta-D-mannuronic acid. Results of x-ray diffraction and polarized infrared studies

A Structure determination of the naturally occuring marine algal polysaccharide poly-β-D -mannuronic acid is described. The structure consists of 1e → 4e linked D -mannuronic acid chains with the monosaccharide units in the C1 chair conformation. The X-ray fiber diffraction photograph obtained from bundles of fibers prepared from Fucus vesiculosus has been indexed to an orthorhombic unit cell in which a =7.6 Å, b (fiber axis) = 10.4 Å, c = 8.6 Å, the unit cell containing two disaccharide chain segments with space group P2₁2₁2₁. A sheet-like structure involving one intra-chain, one intra-sheet, and one inter-sheet hydrogen bond per monosaccharide is proposed. Features of the chain-packing arrangement are compared with mannan.     

The crystal structure of penicillopepsin at 6Åresolution

A 6Åresolution electron density map of crystals of penicillopepsin, an acid protease from Penicillium janthinellum, has been computed from multiple isomorphous replacement phases determined from two heavy metal derivatives, K₂PtCl₆ and UO₂Cl₂. The mean figure of merit of the map is 0.939. The boundaries of the molecules, of which there are four per unit cell, are readily discernible. The molecule is highly asymmetric with approximate dimensions 60Å× 40Å× 30Å. The molecule consists of two distinct lobes separated by a deep cleft, which is probably the extended substrate binding site.     

Crystallization and preliminary diffraction studies of thioesterase II from rat mammary gland

Thioesterase II from rat mammary gland has been crystallized in the presence of decanoic acid by the vapor diffusion method. The crystals belong to the orthorhombic space group P2₁2₁2₁, and have cell dimensions, a = 52.7 Å, b = 78.0 Å, and c = 133.6 Å. The asymmetric unit likely consists of two protein monomers based on predictions from its calculated Matthews coefficient. Crystals typically diffract to at least 2.5 Å resolution and are suitable for X-ray crystallographic analysis. © 1995 Wiley-Liss, Inc.     

Synthesis,characterization and crystal structure of 2-dimethylacetal-4-chloro-6-formylphenol and aquabis(2-dimethylacetal-4-chloro-6-formylphenolato)dioxouranium(VI)

The ligand 2-dimethylacetal-4-chloro-6-formylphenol, H(ALAC), prepared by boiling 2,6-diformyl-4-chlorophenol, H(DIAL), in methanol, was reacted with uranyl acetate to obtain the complex [UO₂(ALAC)₂(H₂O)]. The ligand and the uranyl complex were characterized by X-ray crystallography, infrared, ¹H NMR and electronic spectroscopy. Thermogravimetric and mass spectrometry data are also reported. In acid media H(ALAC) transforms easily into H(DIAL). H(ALAC) is monoclinic, P2₁/n, with a=13.951(5), b=7.902(5), c=9.465(5) Å, β= 91.33(3)°. The structure was refined to R=3.9%, based on 1657 observed reflexions. [UO₂(ALAC)₂(H₂O)] is tetragonal, P4₃2₁2, with a=11.147(5) and c=19.150(4) Å. The structure was refined to R=4.0%, based on 2938 observed reflexions. Four ligand oxygens and one water molecule are equatorially bonded to the uranyl group in this compound. Uranium and water oxygen lie in special positions on a crystallographic twofold axis so that the two halves of this molecule are symmetrically related. Selected bond distances for [UO₂(ALAC)₂(H₂O)] are: UO (charged) 2.28(2) Å, UO (neutral) 2.45(2) Å, UO (uranyl) 1.77(2) Å, UO (water) 2.44(4) Å.     

Structure determination of feline panleukopenia virus empty particles

Various crystal forms of the single-stranded DNA, feline panleukopenia virus (FPV), a parvovirus, have been grown of both full virions and empty particles. The structure of empty particles crystallized in an orthorhombic space group P2₁2₁2₁, with unit cell dimensions a = 380.1 Å, b = 379.3 Å, and c = 350.9 Å, has been determined to 3.3 Å resolution. The data were collected using oscillation photography with synchrotron radiation. The orientations of the empty capsids in the unit cell were determined using a self-rotation function and their positions were obtained with an R-factor search using canine parvovirus (CPV) as a model. Phases were then calculated, based on the CPV model, to 6.0 Å resolution and gradually extended to 3.3 Å resolution by molecular replacement electron density averaging. The resultant electron density was readily interpreted in terms of the known amino acid sequence. The structure is contrasted to that of CPV in terms of host range, neutralization by antibodies, hemagglutination properties, and binding of genomic DNA. © Wiley-Liss, Inc.     

Conformations within soluble oligomers and insoluble aggregates revealed by resonance energy transfer

A fluorescently labeled 20‐residue polyglutamic acid (polyE) peptide 20 amino acid length polyglutamic acid (E₂₀) was used to study structural changes which occur in E₂₀ as it co‐aggregates with other unlabeled polyE peptides. Resonance energy transfer (RET) was performed using an o‐aminobenzamide donor at the N‐terminus and 3‐nitrotyrosine acceptor at the C‐terminus of E₂₀. PolyE aggregates were not defined as amyloid, as they were nonfibrillar and did not bind congo red. Circular dichroism measurements indicate that polyE aggregation involves a transition from α‐helical monomers to aggregated β‐sheets. Soluble oligomers are also produced along with aggregates in the reaction, as determined through size exclusion chromatography. Time‐resolved and steady‐state RET measurements reveal four dominant E₂₀ conformations: (1) a partially collapsed conformation (24 Å donor–acceptor distance) in monomers, (2) an extended conformation in soluble oligomers (>29 Å donor–acceptor distance), (3) a minor partially collapsed conformation (22 Å donor‐acceptor distance) in aggregates, and (4) a major highly collapsed conformation (13 Å donor–acceptor distance) in aggregates. These findings demonstrate the use of RET as a means of determining angstrom‐level structural details of soluble oligomer and aggregated states of proteins. © 2009 Wiley Periodicals, Inc. Biopolymers 93: 299–317, 2010. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com     

Comparative structural studies of the first row early transition metal(III) chloride tetrahydrofuran solvates

Two compounds of empirical formula MCl₃- (THF)₃, M = V and Cr, have been characterized by single crystal X-ray studies. The VCl₃(THF)₃ molecule, which has a mer octahedral stereochemistry, crystallizes in the monoclinic space group P2₁/c with a= 8.847(2),b= 12.861(5),c= 15.134(3) Å, β = 91.94(2)°, V = 1721(1) ų and Z = 4. The V-Ci(1) and V-CI(2) distances have a mean value of 2.330 [3] Å while V-CI(3) = 2.297(2) Å, The VO(1) and VO(2) distances have a mean value of 2.061[8] Å while V-O(3) = 2.102(3) Å cis ClVCl angles average 92.0[5]° and cis OVO angles average 86.2[2]° . The isostmctural complex, CrCl₃(THF)₃, has a crystal structure made up of discrete octahedral mer-CrCl₃(THF)₃ molecules with the following unit cell dimensions (space group P2₁/c): a = 8.715(1), b= 12.786(3), c = 15.122(3) Å, β = 92.15(1)°, V = 1684(1) ų and Z = 4. The CrCl(1) and CrCl(2) distances have a mean value of 2.310131 Å while CrCl(3) = 2.283(2) Å. The CrO(1) and CrO(2) distances have a mean value of 2.0101171 Å while CrO(3) = 2.077(4) Å. cis ClCrCl angles average 90.9[4]° and cis OCrO angles average 86.1 [2]°. The structures of these two octahedral complexes and those previously reported for ScCl₃(THF)₃ and TiCl₃(THF)₃ are compared and certain general trends are discussed.     

Crystallization,molecular replacement solution,and refinement of tetrameric β-amylase from sweet potato

Sweet potato β-amylase is a tetramer of identical subunits, which are arranged to exhibit 222 molecular symmetry. Its subunit consists of 498 amino acid residues (Mr 55,880). It has been crystallized at room temperature using polyethylene glycol 1500 as precipitant. The crystals, growing to dimensions of 0.4 mm × 0.4 mm × 1.0 mm within 2 weeks, belong to the tetragonal space group P4₂2₁2 with unit cell dimensions of a = b = 129.63 Å and c = 68.42 Å. The asymmetric unit contains 1 subunit of β-amylase, with a crystal volume per protein mass (V_M) of 2.57 ų/Da and a solvent content of 52% by volume. The three-dimensional structure of the tetrameric β-amylase from sweet potato has been determined by molecular replacement methods using the monomeric structure of soybean enzyme as the starting model. The refined subunit model contains 3,863 nonhydrogen protein atoms (488 amino acid residues) and 319 water oxygen atoms. The current R-value is 20.3% for data in the resolution range of 8–2.3 Å (with 2 σ cut-off) with good stereochemistry. The subunit structure of sweet potato β-amylase (crystallized in the absence of α-cyclodextrin) is very similar to that of soybean β-amylase (complexed with α-cyclodextrin). The root-mean-square (RMS) difference for 487 equivalent Cα atoms of the two β-amylases is 0.96 Å. Each subunit of sweet potato β-amylase is composed of a large (α/β)₈ core domain, a small one made up of three long loops [L3 (residues 91–150), LA (residues 183–258), and L5 (residues 300–327)], and a long C-terminal loop formed by residues 445–493. Conserved Glu 187, believed to play an important role in catalysis, is located at the cleft between the (α/β)₈ barrel core and a small domain made up of three long loops (L3, L4, and L5). Conserved Cys 96, important in the inactivation of enzyme activity by sulfhydryl reagents, is located at the entrance of the (α/β)₈ barrel. © 1995 Wiley-Liss, Inc.     

Full-matrix least-squares refinement of lysozymes and analysis of anisotropic thermal motion

Crystal structures of turkey egg lysozyme (TEL) and human lysozyme (HL) were refined by full-matrix least-squares method using anisotropic temperature factors. The refinement converged at the conventional R-values of 0.104 (TEL) and 0.115 (HL) for reflections with F_o > 0 to the resolution of 1.12 Å and 1.15 Å, respectively. The estimated r.m.s. coordinate errors for protein atoms were 0.031 Å (TEL) and 0.034 Å (HL). The introduction of anisotropic temperature factors markedly reduced the R-value but did not significantly affect the main chain coordinates. The degree of anisotropy of atomic thermal motion has strong positive correlation with the square of distance from the molecular centroid. The ratio of the radial component of thermal ellipsoid to the r.m.s. magnitude of three principal components has negative correlation with the distance from the molecular centroid, suggesting the domination of libration rather than breathing motion. The TLS model was applied to elucidate the characteristics of the rigid-body motion. The TLS tensors were determined by the least-squares fit to observed temperature factors. The profile of the magnitude of reproduced temperature factors by the TLS method well fitted to that of observed B_eqv. However, considerable disagreement was observed in the shape and orientation of thermal ellipsoid for atoms with large temperature factors, indicating the large contribution of local motion. The upper estimate of the external motion, 67% (TEL) and 61% (HL) of B_eqv, was deduced from the plot of the magnitude of TLS tensors determined for main chain atoms which were grouped into shells according to the distance from the center of libration. In the external motion, the translational portion is predominant and the contribution of libration and screw motion is relatively small. The internal motion, estimated by subtracting the upper estimate of the external motion from the observed temperature factor, is very similar between TEL and HL in spite of the difference in 54 of 130 amino acid residues and in crystal packing, being suggested to reflect the intrinsic internal motion of chicken-type lysozymes. Proteins 30:232–243, 1998. © 1998 Wiley-Liss, Inc.     

Assignment of the proton Nmr chemical shifts of the T-N3H and G-N1H proton resonances in isolated AT and GC Watson-Crick base pairs in double-stranded deoxy oligonucleotides in aqueous solution

The 300-MHz proton nmr spectra (between 11 and 14 ppm) of a series of double-stranded deoxy oligonucleotides of known sequence have been recorded in H₂O solution. These resonances have been assigned to the G? N₁H and T? N₃H protons of specific base pairs from an evaluation of the temperature dependence of the ring NH linewidths and from the selective ring NH chemical shift changes on actinomycin-D binding. The deoxy oligonucleotides exist predominantly in the DNA-B conformation as evaluated from antibiotic binding studies. Ring-current calculations have been utilized to evaluate the up-field shifts of the G? N₁H and T? N₃H protons in Watson-Crick base pairs due to the ring currents from the pyrimidine and purine rings of nearest neighbor base pairs in regular DNA-B- and RNA-A-type helices. The perturbations on these up-field ring-current contributions that arise from twisting and tilting a base pair adjacent to the ring NH under study have been evaluated and found to change the calculated chemical shift by ±0.6 ppm for twist and tilt distortions of <30°C in a single adjacent base pair. A knowledge of the experimentally assigned ring NH chemical shifts of specific base pairs in known sequences of double-stranded deoxy oligonucleotides coupled with the ring-current tables for the DNA-B helical structure permit the assignment of 13.6 ± 0.1 ppm and 14.6 ± 0.2 ppm for the G? N₁H proton of an isolated GC base pair and the T? N₃H proton of an isolated AT base pair, respectively.     

1H- and 13C-nmr studies on caffeine and its interaction with nucleic acids

The relative orientation of caffeine in stacks formed by self-association in aqueous solution has been evaluated by both ¹H- and ¹³C-nmr spectroscopy. The data, which were interpreted through the calculation of ring-current and atomic diamagnetic anisotropic effects of the caffeine molecule, suggest two caffeine bases may stack in a nearly orthogonal manner. The interactions between caffeine and adenylyl-3′,5′-adenosine, polyadenylic acid, and ribo-(A-A-G-C-U-U)₂ helix were also studied by nmr at different caffeine/base ratios and at varying temperatures. The results show that caffeine tends to stack on the top of the terminal purine bases or to insert (the single-stranded) or to intercalate (the double-stranded) between purine bases.     

Eight-coordination. Synthesis and structure of the schiff-base chelate bis(N,N′-disalicylidene-1,2-phenylenediamino)cerium(IV)

In an effort to elucidate the factors that determine the geometry of eight-coordinate complexes we synthesized and structurally characterizes the complex bis(N,N′-disalicylidene-1,2-phenylenediamino)Cerium(IV), Ce(salophen)₂. It is a square antiprismatic species with each chelating ligand spanning sss edges. The CeO bonds average 2.214 Å, appreciably shorter than CeN distance of 2.606 Å. The tow ligands have the sandwich orientation and the trapezoidal planes intersect at 88.2°. The δ angles are 7.0°, 2.6°, 56.5° and 55.9° and the ? angles are 30.0° and 28.6°. The complex crystallized in space group P2₁/C with Z = 4, a = 11.090(2) Å, b = 16.332(3) Å, c = 18.916(3) Å, β = 70.10(2)° and d_calc = 1.585 g/cm₃.     

Chlorotriphenyl(quinolinium-2-carboxylato)tin(IV) monohydrate

The crystal structure of chlorotriphenyl(quinolinium-2-carboxylato)tin(IV) monohydrate is reported. The crystals are monoclinic, space group C2/c with cell parameters a = 20.048(3) Å, b = 11.724(1) Å, c = 23.291(3) Å, ]gb = 113.42(1), Z = 8, refined to R_F = 0.034 on 3331 observed reflections. The tin(IV) atom is five-coordinate, being found to three phenyl groups, the chlorine atom and an oxygen from the quinaldic acid. The geometry around the tin atom is trigonal bipyramidal, with the three phenyl groups occupying the equatorial positions, and the chlorine and quinaldic acid oxygen, the apical ones. The acidic proton of quinaldic acid has shifted position in the complex, and is bound to the heterocyclic nitrogen atom.The acid is thus coordinated in the form of a zwitterion. These trigonal bipyramidal units are linked together as dimers by pairs of water molecules, each of which hydrogen-bonds to the non-coordinated carboxylate oxygen atoms of both quinaldic acid molecules, plus the heterocyclic nitrogen atom of one quinaldic acid molecule. For complex formation with the protonated acid, the heterocyclic nitrogen should be alpha to the carboxylic acid group.     

Three-dimensional structure of actinoxanthin. IV. A 2.5-Å resolution

The protein actinoxanthin (isolated from Actinomyces globisporus—molecular weight, 10,300; 107 amino acid residues) crystallizes in space group P2₁2₁2₁ with cell dimensions: a = 30.9 Å, b = 48.8 Å, c = 64.1 Å, and Z = 4. The three-dimensional structure of actinoxanthin was determined by the x-ray multiple isomorphous replacement method at 2.5-Å resolution. The molecule is kidney-shaped and has a well-defined cavity. Its characteristic features are the absence of α-helices and the presence of enhanced content of antiparallel β-structure (～55%). A cylinder-shaped formation of seven antiparallel β-strands comprises the main part of the protein structure. The established β-supersecondary structure is characterized by a three-dimensional topology similar to that of immunoglobulin domains, superoxide dismutase subunits, and azurin and plastocyanin proteins.