A structural and free energy analysis of Ag complexes to five small peptides

The complexes of Ag⁺ with the peptides MetGly, ProGly, GlyPro, GlyHis and GlyProAla were investigated using hybrid density functional theory at the B3LYP/DZVP level. The silver ion binding free energies at 298 K to each of these peptides was calculated to be 60.8, 52.0, 54.3, 71.2 and 63.3 kcal mol⁻¹, respectively. Structural information and relative free energies are presented for several isomers for each of the five complexes. Each of the global minima found for the five complexes is a charge-solvated ion. An important finding is that the Ag⁺-ProGly is the only complex where a salt bridge structure is energetically favored occurring at 4.0 kcal mol⁻¹ higher in free energy than the global minimum. The Ag⁺ ion in this salt bridge structure is attached to the carboxylate anion of zwitterionic ProGly in which the terminal amino nitrogen is protonated. For all the other complexes studied, the salt bridge structure occurs at much higher energies. All the dipeptide complexes with Ag⁺, but one, exhibit a di- or tri-coordinate metal where the sites of attachment are amino and carbonyl groups. However, the highest coordination numbers are not always the global minima due to steric costs. The global minimum of the Ag⁺-GlyProAla complex is the only structure found in this study where the metal is tetra-coordinated, binding to the terminal amino nitrogen and all three carbonyl oxygen atoms. Silver binding to sulphur and imidazole nitrogen atoms of MetGly and GlyHis, respectively, are present in the three most energetically favored species in each of these cases.     

Fibre X-Ray and Conformational Study of the Binding of Metal Ions on DNA

Abstract

Results obtained from X-ray diffraction as well as from conformational analysis of Ag-DNA fibres are presented. For small percentages of Ag⁺ bound and high humidity, the B-DNA form is maintained. As the percentage of Ag⁺ is increased, the helical parameters of the B-DNA are modified. These modifications are directly related to the percentage of G—C bases. The periodicity of the DNA fibres are perturbed as Ag⁺ is mainly bound to G—C pairs and, thus, only the equatorial diffracted intensities can be compared to values calculated from molecular models. It is shown, by this way, that the first binding site is located on N7 of G. A second site is situated between N3 and N1 of the G—C pair, at the place of a hydrogen bond. A molecular model of the Ag-DNA complex is proposed and shown to be in agreement with experimental data. Results obtained allow to get some information on the binding of other ions such as Cu²⁺ and Hg²⁺ which give very little modification of the fibre X-ray patterns.     

Induction of helicity in polyuridylic acid and polyinosinic acid by silver ions

Silver ions binding to poly(U) and poly(I) produce highly ordered multistranded helices under conditions which would otherwise lead to random coils. Evidence for helicity comes from the hypochromicity and high ellipticity generated in the polymers by Ag⁺ binding, as well as from x-ray studies and from the cooperativity of the Ag⁺ complexing reaction. Continuous variation studies show that both polymers form 1:1 and 2:1 polymer–Ag⁺ complexes. Low pH favors the 1:1 complex with poly(U) and the 2:1 complex with poly(I); the reverse is true at high pH. Ag⁺ binding and proton-release experiments make it clear that at low pH, unprotonated electron-donor groups are complexed preferentially, but that at high pH, Ag⁺ readily displaces H⁺ from protonated groups. In poly(I) the unprotonated donor is N(7), leading at low pH to a 2:1 complex containing N(7)-Ag-N(7) bonds; at high pH, proton release from N(1) leads to a 1:1 complex containing N(1)-Ag-O bonds. In poly(U) there is no unprotonated donor; the low-pH 1:1 complex involves deprotonation of only one N(3) per bound Ag⁺, leading to N₃-Ag-O bonding, while high pH causes deprotonation of two N(3) per Ag⁺ and a 2:1 N(3)-Ag-N(3) complex. Thus silver ions react with the nucleotide bases in chemically predictable ways, and the formation of different Ag–nucleotide bonds leads to different multiple-helix structures.     

Novel Hoogsteen-like bases for configurational recognition of the T-A base pair by DNA triplex formation

Effective sequence-specific recognition of duplex DNA is possible by triplex formation with natural oligonucleotides via Hoogsteen H-bonding. However, triplex formation is in practice limited to pyrimidine oligonucleotides binding duplex A-T or G-C base-pair DNA sequences specifically at homopurine sites in the major groove as T·A-T and C⁺·G-C triplets. Here we report the successful modeling of novel unnatural nucleosides that recognize the T-A DNA base pair by Hoogsteen interaction. Since the DNA triplex can be considered to assume an A-type or B-type conformation, these novel Hoogsteen nucleotides are tested within model A-type and B-type conformation triplex structures. A triplet consisting of the T-A base pair and one of the novel Hoogsteen nucleotides replaces the central T·A-T triplet in the triplex using the same deoxyribose-phosphodiester and base-deoxyribose dihedral angle configuration. The entire triplex is energy minimized and the presence of any structural or energetic perturbations due to the central triplet is assessed with respect to the unmodified energy-minimized (T·A-T)₁₁ proposed starting structures. Incorporation of these novel triplets into both A-type and B-type natural triplex structures provokes minimal change in the configuration of the central and adjacent triplets. The plan is to produce a series of Hoogsteen-like bases that preferentially bind the T-A major groove in either an A-type or B-type conformation. Selective recognition of the T-A major groove with respect to the G-C major groove, which presents similar keto and amine placement, is also assessed with configurational preference. Evaluation of the triplex solution structure by using these unnatural bases as binding conformational probes is a prerequisite to the further design of triplet forming bases. © 1996 John Wiley & Sons, Inc.     

DNA binding of a spermine derivative: Spectroscopic study of anthracene-9-carbonyl-n1-spermine with poly[d(G-C)·(d(G-C))] and poly[d(A-T) · d(A-T)]

The binding of polyamines, including spermidine ( 1 ) and spermine ( 2 ), to poly[d(G-C) · d(G-C) ] was probed using spectroscopic studies of anthracene-9-carbonyl-N¹-spermine ( 3 ); data from normal absorption, linear dichroism (LD), and circular dichroism (CD) are reported. Ligand LD and CD for transitions located in the DNA region of the spectrum were used. The data show that 3 binds to DNA in a manner characteristic of both its amine and polycyclic aromatic parts. With poly [(dG-dC) · (dG-dC)], binding modes are occupied sequentially and different modes correspond to different structural perturbations of the DNA. The most stable binding mode for 3 with poly[d(G-C) · d(G-C)] has a site size of 6 ± 1 bases, and an equilibrium binding constant of (2.2 ± 1.1) × 10⁷ M^?1 with the anthracene moiety intercalated. It dominates the spectra from mixing ratios of approximately 133:1 until 6:1 DNA phosphate: 3 is reached. The analogous data for poly [d(A-T) · d(A-T)] between mixing ratios 36:1 and 7:1 indicates a site size of 8.3 ± 1.1 bases and an equilibrium binding constant of (6.6 ± 3.3) × 10⁵ M^?1. Thus, 3 binds preferentially to poly [d(G-C) · d(G-C)] at these concentrations. © 1994 John Wiley & Sons, Inc.     

DNA Logic Gate Based on Metallo-Toehold Strand Displacement

DNA is increasingly being used as an ideal material for the construction of nanoscale structures, circuits, and machines. Toehold-mediated DNA strand displacement reactions play a very important role in these enzyme-free constructions. In this study, the concept of metallo-toehold was utilized to further develop a mechanism for strand displacement driven by Ag⁺ ions, in which the intercalation of cytosine–cytosine mismatched base pairs on the toeholds provides additional control by varying of the concentration of Ag⁺ ions. The characteristics of displacement reaction in response to different concentration of Ag⁺ ions are investigated by fluorescence spectral and non-denaturing polyacrylamide gel electrophoresis. The reaction can successfully occur when the concentration of Ag⁺ ions is suitabe; excess Ag⁺ ions block the reaction. Furthermore, the displacement reaction can be tuned and controlled most efficiently under the condition of two C:C mismatched base pairs placed on the six-nt toehold. Based on our research, a mechanism was developed to construct Boolean logic gate AND and OR by employing strand displacement reaction as a tool, Ag⁺ and Hg²⁺ as input.     

Isolation and characterization of satellite DNA from mustard seedlings

The DNA from mustard (Sinapis alba L.) seedlings was examined by neutral CsCl and Ag⁺/Cs₂SO₄ density gradient centrifugation. Different satellite fractions were revealed by these two methods. The satellite fractions obtained from the Ag⁺/Cs₂SO₄ density gradient could not be generally correlated with satellite DNA fractions observed in CsCl. In CsCl density gradient centrifugation, a main band at density 1,695 g/cm³ and a heavy shoulder at density 1,703 g/cm³ are found. By preparative CsCl gradient centrifugation the heavy shoulder can be enriched but not completely separated from the main band DNA.—Gradient centrifugation by complexing the DNA with Ag⁺ rf. 0.25 to DNA phosphate reveals three distinct fractions which are further characterized: The heavy satelite DNA fraction revealed by Ag⁺/Cs₂SO₄ gradient centrifugation has the same density in a CsCl gradient and the same Tm value as the main band, but differs from main band DNA in the details of its melting profile and in its renaturation kinetics. The light Ag⁺/Cs₂SO₄ satellite DNA fraction had a higher melting temperature corresponding to a GC-rich base composition. Differences between these 3 fractions are observed in thermal denaturation and renaturation profiles, hybridization in situ with ribosomal RNA, and their response to restriction endonuclease digestion. The light satellite fraction from the Ag⁺/Cs₂SO₄ gradient, rich in ribosomal cistrons corresponds to the heavy shoulder DNA of neutral CsCl gradients which also is rich in ribosomal cistrons. The heavy satellite fraction from Ag⁺/Cs₂SO₄ gradient which contains highly repetitive short nucleotide sequences could not be revealed by the classical CsCl gradient centrifugation technique.     

The ground-state Na+ affinities of a series of substituted acetophenones: a DFT study

A detailed study of Na⁺ affinities of a series of para-substituted acetophenones and their O–Na⁺ counterparts was performed using density functional theory [Becke, Lee, Yang and Parr (B3LYP)] method using 6-311G(d,p) basis sets with complete geometry optimisation. The gas-phase O–Na⁺ complex formation turns out to be an exothermic case and the local stereochemical disposition of Na⁺ is found to be almost the same in each case. The presence of the para-substituent is seen to cause very little change in the Na⁺ affinity relative to the unsubstituted acetophenones. Electron-releasing p-substituents increase it by 0.0105 hartree and electron-withdrawing p-substituents decrease it by 0.011 hartree. Computed Na⁺ affinities are sought to be correlated with a number of computed system parameters such as the net charge on the Na⁺ and the carbonyl oxygen of the Na⁺ complexes and the net charge on the carbonyl oxygen of the free bases. The energetics, structural and electronic properties of the complexes indicate that the interaction between the Na⁺ ion and a carbonyl base is predominantly an ion–dipole attraction and the ion-induced dipole interaction as well rather than a covalent interaction.     

Glucocorticoid receptor-steroid complex binding to DNA. Competition between DNA and DNA-cellulose.

The binding of the glucocorticoid receptor-steroid complex from a line of rat hepatoma tissue culture (HTC) cells to DNA has been examined. An equilibrium competition assay involving a constant, low total amount of double-stranded DNA was developed to compare the complex binding ability of DNA free in solution and bound to cellulose. This binding ability is lowered by a factor of five when DNA is associated with cellulose. Similar studies with HTC cell, calf-thymus, and Escherichia coli DNA revealed no difference in the relative number or affinity of binding sites for receptor-steroid complex in each DNA. The synthetic DNA molecules poly[d(A-T)-d(A-T)] and poly[d(G-C)-d(G-C)] bound complexes equally well but less than the three "natural" DNA molecules. This appears to be due to differences in acceptor site affinity and suggests that nucleotide complexity and/or sequence influences the affinity of HTC cell receptor-glucocorticoid complexes for DNA.     

Mutagenic SOS repair of site-specific psoralen damage in plasmid pBR322

The mutagenic repair of psoralen damage was examined by transforming Escherichia coli with psoralen-treated pBR322. Plasmid DNA randomly reacted with psoralen was repaired only when the E. coli was uvrA⁺ and recA⁺, and only when the cells were pre-irradiated with far-ultraviolet light. The recA dependence and requirement for pre-irradiation are characteristics of SOS repair.Psoralens were placed specifically near the BamHI site, in the tetracycline-resistance gene of pBR322, using a sulfhydryl-containing psoralen derivative. Repair of this damage also required pre-irradiation of the host cells. This repair was accompanied by a 4% frequency of mutagenesis to a tetraeycline-sensitive phenotype. Sequence analysis of these mutant plasmids revealed that 75% had mutations within the targeted region, while 25% had no sequence changes within 100 bases of the BamHI site. In up to five independent isolates only one kind of mutation was observed at each site, suggesting that mutagenic SOS repair is influenced by DNA structure at the site of the psoralen. Most mutations were transitions, primarily G-C to A-T changes. Some transitions occurred at sites where psoralen crosslinks could not have formed, and these may have arisen from the repair of psoralen monoadducts.     

Mutagenic role of Watson-Crick protons in alkylated DNA bases: A theoretical study

Loss of Watson-Crick protons following DNA base alkylation has been proposed as a key event which confers mutation-inducing properties on to alkylated DNA bases. In this theoretical study, the promutagenic O⁶-guanine and O⁴-thymine sites are clearly distinguished from the nonmutagenic N⁷-guanine site on the basis of calculated values of mechanistic indicators for Watson-Crick proton acidity following alkylation at these respective sites. The degree of acidity predicted for these protons for each type of alkylated base accords well with the presence or absence of mutagenicity observed experimentally in each case.     

Interactions des ions métalliques avec le DNA. III. Stabilité et configuration des complexes Ag–DNA

The thermal stabilization and the configurational changes of DNA were studied during the binding process of silver ion. Melting curves of silver–DNA complexes were analyzed according to the method of Felsenfeld. An increase of T_m occurred during the formation of the first complex with a preferential stabilization of G–C pairs. In the case of DNA from H. influenzae, the transforming power was maintained after heating in presence of silver at a temperature where AT pairs were dissociated but G-C pairs were not. Shape modifications of the molecule were followed by light scattering, both with native and ultrasonic degraded DNA. In the case of native DNA, the binding of positive ions modified the electrostatic potential and increased the flexibility of the molecule. In the case of rod shaped fragments of DNA, the progressive formation of a kind of polyampholyte was accompanied by aggregation phenomena. The formation of the second complex, with proton release, induced changes of secondary structure and possibly a tilt of the plane of the base.     

Thermodynamic and structural properties of the specific binding between Ag ion and C:C mismatched base pair in duplex DNA to form C–Ag–C metal-mediated base pair

Metal ion-nucleic acid interactions have attracted considerable interest for their involvement in structure formation and catalytic activity of nucleic acids. Although interactions between metal ion and mismatched base pair duplex are important to understand mechanism of gene mutations related to heavy metal ions, they have not been well-characterized. We recently found that the Ag⁺ ion stabilized a C:C mismatched base pair duplex DNA. A C–Ag–C metal-mediated base pair was supposed to be formed by the binding between the Ag⁺ ion and the C:C mismatched base pair to stabilize the duplex. Here, we examined specificity, thermodynamics and structure of possible C–Ag–C metal-mediated base pair. UV melting indicated that only the duplex with the C:C mismatched base pair, and not of the duplexes with the perfectly matched and other mismatched base pairs, was specifically stabilized on adding the Ag⁺ ion. Isothermal titration calorimetry demonstrated that the Ag⁺ ion specifically bound with the C:C base pair at 1:1 molar ratio with a binding constant of 10⁶ M⁻¹, which was significantly larger than those for nonspecific metal ion-DNA interactions. Electrospray ionization mass spectrometry also supported the specific 1:1 binding between the Ag⁺ ion and the C:C base pair. Circular dichroism spectroscopy and NMR revealed that the Ag⁺ ion may bind with the N3 positions of the C:C base pair without distorting the higher-order structure of the duplex. We conclude that the specific formation of C–Ag–C base pair with large binding affinity would provide a binding mode of metal ion-DNA interactions, similar to that of the previously reported T-Hg-T base pair. The C–Ag–C base pair may be useful not only for understanding of molecular mechanism of gene mutations related to heavy metal ions but also for wide variety of potential applications of metal-mediated base pairs in various fields, such as material, life and environmental sciences.     

Chiral Discrimination of Ofloxacin Enantiomers Using DNA Double Helix Regulated by Metal Ions

DNA‐based chiral selectors are constructed to discriminate ofloxacin enantiomers through metal‐ion anchoring on a special DNA double helix that contains successive GC pairs. The effects of metal ions involving Mg²⁺, Ni²⁺, Cu²⁺, Ag⁺, and Pt²⁺ were studied on the regulation of DNA chiral discrimination towards ofloxacin enantiomers. It is shown that DNA‐Cu(II) complexes exhibit the highest enantioselectivities at the [Cu²⁺]/base ratio of 0.1. The enantiomeric excess can reach 59% in R‐enantiomer after being adsorbed by the RET‐Cu(II) complex. Stereoselective recognition of ofloxacin enantiomers on the double helix is tunable via external stimulus, providing a programmable desorption process to regenerate DNA. This DNA‐based chiral selector exhibits excellent reusability without apparent loss of enantioselectivity after three cycles of adsorption and desorption. Chirality 26:249–254, 2014. © 2014 Wiley Periodicals, Inc.     

DNA nicking by HinP1I endonuclease: bending, base flipping and minor groove expansion

HinP1I recognizes and cleaves the palindromic tetranucleotide sequence G↓CGC in DNA. We report three structures of HinP1I–DNA complexes: in the presence of Ca²⁺ (pre-reactive complex), in the absence of metal ion (binary complex) and in the presence of Mg²⁺ (post-reactive complex). HinP1I forms a back-to-back dimer with two active sites and two DNA duplexes bound on the outer surfaces of the dimer facing away from each other. The 10 bp DNA duplexes undergo protein-induced distortions exhibiting features of A-, B- and Z-conformations: bending on one side (by intercalation of a phenylalanine side chain into the major groove), base flipping on the other side of the recognition site (by expanding the step rise distance of the local base pair to Z-form) and a local A-form conformation between the two central C:G base pairs of the recognition site (by binding of the N-terminal helix in the minor groove). In the pre- and post-reactive complexes, two metals (Ca²⁺ or Mg²⁺) are found in the active site. The enzyme appears to cleave DNA sequentially, hydrolyzing first one DNA strand, as seen in the post-reactive complex in the crystalline state, and then the other, as supported by the observation that, in solution, a nicked DNA intermediate accumulates before linearization.     

Structure–function correlation for the EcoRV restriction enzyme: from non-specific binding to specific DNA cleavage

The EcoRV restriction endonuclease cleaves DNA at its recognition sequence at least a million times faster than at any other DNA sequence. The only cofactor it requires for activity is Mg²⁺: but in binding to DNA in the absence of Mg²⁺, the EcoRV enzyme shows no specificity for its recognition site. Instead, the reason why EcoRV cuts one DNA sequence faster than any other is that the rate of cleavage is controlled by the binding of Mg²⁺ to EcoRV-DNA complexes: the complex at the recognition site has a high affinity for Mg²⁺, while the complexes at other DNA sequences have low affinities for Mg²⁺. The structures of the EcoRV endonuclease, and of its complexes with either 8pecific or non-specific DNA, have been solved by X-ray crystallography. In the specific complex, the protein interacts with the bases in the recognition sequence and the DNA takes up a highly distorted structure. In the non-specific complex with an unrelated DNA sequence, there are virtually no interactions with the bases and the DNA retains a B-like structure. Since the free energy changes for the formation of specific and non-specific complexes are the same, the energy from the specific interactions balances that required for the distortion of the DNA. The distortion inserts the phosphate at the scissile bond into the active site of the enzyme, where it forms part of the binding site for Mg²⁺. Without this distortion, the EcoRV–DNA complex would be unable to bind Mg²⁺ and thus unable to cleave DNA. The specificity of the EcoRV restriction enzyme is therefore governed, not by DNA binding as such, but by its ability to organize the structure of the DNA to which it is bound.     

In vitro recognition of DNA base pairs by histones in histone-DNA complexes and reconstituted core particles: an ultraviolet resonance Raman study.

Resonance Raman spectra of complexes between DNA and the four core histones, alone or associated, have been investigated in vitro using excitations at 300 and 257 nm, which give complementary informations about the DNA bases. H2A and H2B fractions recognize the G-C base pairs, while H3 and H4 (arginine rich fractions) recognize the A-T base pairs. The associated fractions form complexes with DNA which yield about the same DNA spectral modifications as the DNA-H4 complexes. This reveals the important role of the arginine rich fractions in the core particle formation and confirms the preferential in vitro assembly of nucleosome cores on A-T rich regions of DNA (25).     

DNA conformational change in Gal repressor-operator complex: involvement of central G-C base pair(s) of dyad symmetry.

Gal repressor dimer binds to two gal operator sites, OE and OI, which are 16 bp long similar sequences with hyphenated dyad symmetries (11,12). Repressor occupation hinders the reactivity of the N7 atoms in the major groups of guanines, located at positions 1, 3 and 8, and the rotational 1', 3' and 8' of the symmetries. We have shown that Gal repressor binding to OE or OI DNA fragments increases the circular dichroism (CD) spectral peak in the 270 to 300 nm range. The CD change is similar to that observed for Lac repressor binding to its operator site (14). It is consistent with a DNA conformational change during complex formation between Gal repressor and OE and OI DNA. The CD spectral change was not observed when the central 8,8' G-C base pairs in the DNA-protein complex were replaced by A-T base pairs, whereas substitution of the 1,1' G-C base pairs do show the accompanying increase in the spectra during repressor binding. The absence of CD change of the Gal repressor complex with DNA mutated at the 8,8' base pairs suggest that the central G-C base pairs are required for the repressor induced conformational change.     

Solution structure of actinomycin-DNA complexes: Drug intercalation at isolated G-C sites

Summary The actinomycin-D-d(A1-A2-A3-G4-C5-T6-T7-T8) complex (1 drug per duplex) has been generated in aqueous solution and its structure characterized by a combined application of two-dimensional NMR experiments and molecular dynamics calculations. We have assigned the exchangeable and nonexchangeable proton resonances of Act and d(A₃GCT₃) in the complex and identified the intermolecular proton-proton NOES that define the alignment of the antitumor agent at its binding site on duplex DNA. The molecular dynamics calculations were guided by 70 intermolecular distance constraints between Act and nucleic acid protons in the complex. The phenoxazone chromophore of Act intercalates at the (G-C)_I·(G-C)_II step in the d(A₃GCT₃) duplex with the phenoxazone ring stacking selectively with the G4_I and G4_II purine bases but not with C4_I and C4_II pyrimidine bases at the intercalation site. There is a pronounced unwinding between the A3·T6 and G4·C5 base pairs which are the next steps located in either direction from the intercalation site in the Act-d(A₃GCT₃) complex. The Act cyclic pentapeptide ring conformations in the complex are similar to those for free Act in the crystal except for a change in orientation of the ester linkage connecting meVal and Thr residues. The cyclic pentapeptide rings are positioned in the minor groove with the established G-C sequence specificity of binding associated with intermolecular hydrogen bonds between the Thr backbone CO and NH groups to the NH₂-2 and N3 positions of guanosine, respectively. Complex formation is also stabilized by van der Waals interactions between nonpolar groups on the cyclic pentapeptide rings and the sugar residues and base pair edges lining the widened minor groove of the (A3-G4-C5-T6)_I·(A3-G4-C5-T6)_II binding site segment of the DNA helix.Dedicated to the memory of Professor V.F. Bystrov     

Covalent modification of guanine bases in double-stranded DNA. The 1.2-A Z-DNA structure of d(CGCGCG) in the presence of CuCl2

We have solved the single crystal structure to 1.2-A resolution of the Z-DNA sequence d(CGCGCG) soaked with copper(II) chloride. This structure allows us to elucidate the structural properties of copper in a model that mimics a physiologically relevant environment. A copper(II) cation was observed to form a covalent coordinate bond to N-7 of each guanine base along the hexamer duplex. The occurrence of copper bound at each site was dependent on the exposure of the bases and the packing of the hexamers in the crystal. The copper at the highest occupied site was observed to form a regular octahedral complex, with four water ligands in the equatorial plane and a fifth water along with N-7 of the purine base at the axial positions. All other copper complexes appear to be variations of this structure. By using the octahedral complex as the prototype for copper(II) binding to guanine bases in the Z-DNA crystal, model structures were built showing that duplex B-DNA can accommodate octahedral copper(II) complexes at the guanine bases as well as copper complexes bridged at adjacent guanine residues by a reactive dioxygen species. The increased susceptibility to oxidative DNA cleavage induced by copper(II) ions in solution of the bases located 5' to one or more adjacent guanine residues can thus be explained in terms of the cation and DNA structures described by these models.