Structure,energetics and properties of some molecules with potent anti-HIV activity: a theoretical study

Density functional theory (DFT; B3LYP) and Hartree–Fock (HF; 3-21G, 6-31G(d) and 6-311G(d,p)) calculations with complete geometry optimisations are carried out in the ground state on five 6-aminoquinolone derivatives, which have been proved to be highly effective in inhibiting HIV replication, to study their structures, energetics and HOMO–LUMO correlation with physiological action. The gas-phase calculations and single-point polarisable continuum model water-phase calculations show that the molecules are highly effective in inhibiting HIV replication, which is in excellent agreement with the experiment. Structural features, energies, charge densities and HOMO–LUMO correlation have been found to substantiate the experimental findings. Compound 4 (pyrazine) shows some special features in DFT calculations which are not found in HF calculations. In the present series, HF results are more reliable as expected.     

Reactivity of pyruvic acid and its derivatives towards reactive oxygen species

Pyruvic acid and its derivatives occurring in most biological systems are known to exhibit several pharmacological properties, such as anti‐inflammatory, neuroprotective or anticancer, many of which are suggested to originate from their antioxidant and free radical scavenger activity. The therapeutic potential of these compounds is a matter of particular interest, due to their mechanisms of action, particularly their possible antioxidant behaviour. Here, we report the results of a study of the effect of pyruvic acid (PA), ethyl pyruvate (EP) and sodium pyruvate (SP) on reactions generating reactive oxygen species (ROS), such as superoxide anion radicals, hydroxyl radicals and singlet oxygen, and their total antioxidant capacity. Chemiluminescence (CL) and spectrophotometry techniques were employed. The pyruvate analogues studied were found to inhibit the CL signal arising from superoxide anion radicals in a dose‐dependent manner with IC₅₀ = 0.0197 ± 0.002 mM for EP and IC₅₀ = 69.2 ± 5.2 mM for PA. These compounds exhibited a dose‐dependent decrease in the CL signal of the luminol + H₂O₂ system over the range 0.5–10 mM with IC₅₀ values of 1.71 ± 0.12 mM for PA, 3.85 ± 0.21 mM for EP and 22.91 ± 1.21 mM for SP. Furthermore, these compounds also inhibited hydroxyl radical‐dependent deoxyribose degradation in a dose‐dependent manner over the range 0.5–200 mM, with IC₅₀ values of 33.2 ± 0.3 mM for SP, 116.1 ± 6.2 mM for EP and 168.2 ± 6.2 mM for PA. All the examined compounds also showed antioxidant capacity when estimated using the ferric–ferrozine assay. The results suggest that the antioxidant activities of pyruvate derivatives may reflect a direct effect on scavenging ROS and, in part, be responsible for their pharmacological actions. Copyright © 2015 John Wiley & Sons, Ltd.     

Bonding patterns in transition metal clusters

Bonding patterns and electron counts of high-symmetry transition metal cluster compounds are discussed using a simplified Tensor Surface Harmonic (TSH) treatment. An n-vertex metal cluster has 9n valence atomic orbitals. From this set orbitals concerned in metal–ligand bonding are eliminated to yield the effective valence orbitals available to cluster bonding. Applying TSH theory to this smaller set gives a clear classification of the cluster MOs and their bonding/antibonding characteristics. Orbital mixing allowed by point group symmetry gives a final, qualitative cluster MO diagram. Results for triangular, tetrahedral and octahedral clusters are compared with other models.     

A theoretical investigation of the functional role of the axial methionine ligand of the Cu(A) site in cytochrome c oxidase

The functional roles of the amino acid residues of the Cu(A) site in bovine cytochrome c oxidase (CcO) were investigated by utilizing hybrid quantum mechanics (QM)/molecular mechanics (MM) calculations. The energy levels of the molecular orbitals (MOs) involving Cu d(zx) orbitals unexpectedly increased, as compared with those found previously with a simplified model system lacking the axial Met residue (i.e., Cu(2)S(2)N(2)). This elevation of MO energies stemmed from the formation of the anti-bonding orbitals, which are generated by hybridization between the d(zx) orbitals of Cu ions and the p-orbitals of the S and O atoms of the axial ligands. To clarify the roles of the axial Met ligand, the inner-sphere reorganization energies of the Cu(A) site were computed, with the Met residue assigned to either the QM or MM region. The reorganization energy slightly increased when the Met residue was excluded from the QM region. The existing experimental data and the present structural modeling study also suggested that the axial Met residue moderately increased the redox potential of the Cu(A) site. Thus, the role of the Met may be to regulate the electron transfer rate through the fine modulation of the electronic structure of the Cu(A) "platform", created by two Cys/His residues coordinated to the Cu ions. This regulation would provide the optimum redox potential/reorganization energy of the Cu(A) site, and thereby facilitate the subsequent cooperative reactions, such as the proton pump and the enzymatic activity, of CcO. This article is part of a Special Issue entitled: Allosteric cooperativity in respiratory proteins.     

An insight into the general relationship between the three dimensional structures of enzymes and their electronic wave functions: Implication for the prediction of functional sites of enzymes

In this study, we explored the general relationship between the three-dimensional (3D) structures of enzymes and their electronic wave functions. Furthermore, we developed a method for the prediction of their functionally important sites. For this purpose, we first performed linear-scaling molecular orbital calculations for 112 nonredundant, non-homologous enzymes with known structure and function. In consequence, we showed that the canonical molecular orbitals (MOs) of the enzymes could be classified into three groups according to the degree of electron delocalization: highly localized orbitals (Group A), highly delocalized orbitals whose electrons are distributed over almost the whole molecule (Group B), and moderately delocalized orbitals (Group C). The MOs belonging to Group A are located near the HOMO-LUMO band gap, and thereby include the frontier orbitals of a given enzyme. We inferred that the MOs of Group B play a role in stabilizing the 3D structure of the enzyme, while those of Group C contribute to constructing the covalent bond framework of the enzyme. Next, we investigated whether the frontier orbitals of enzymes could be used for identifying their potential functional sites. As a result, we found that the frontier orbitals of the 112 enzymes have a high propensity to be colocalized with the known functional sites, especially when the enzymes are hydrated. Such a propensity is shown to be remarkable when Glu or Asp is a functional site residue. On the basis of these results, we finally propose a protocol for the prediction of functional sites of enzymes.     

Density functional studies of the substituent effect on absorption and emission properties of 1, 8-naphthalimide derivatives

Using TD-PBE1PBE/6-31G* and TD-B3LYP/6-31G* approaches, we calculated the absorption and emission spectra of 1,8-naphthalmide derivatives in gas-phase. The geometric structures optimized by HF/6-31G* and B3LYP/6-31G* models and the absorption and emission maxima were in good agreement with existed experimental measurements. It was also found that the lowest singlet states corresponded mainly to the electronic transition from the highest occupied orbital (HOMO) to the lowest unoccupied molecular orbital (LUMO). Intramolecular charge transfer occurred between substituents and naphthalimic rings. Study also showed that most compounds with low absorption excitation energies had low vertical ionization potentials. Finally, the delocalization electronic energies between substituents and naphthalimic rings of isomers were investigated to obtain further sight into their stability.     

Knockdown of gene expression by antisense morpholino oligos in preimplantation mouse embryos cultured in vitro

Knockdown of gene expression by antisense morpholino oligos (MOs) is a simple and effective method for analyzing the roles of genes in mammalian cells. Here, we demonstrate the efficient delivery of MOs by Endo-Porter (EP), a special transfection reagent for MOs, into preimplantation mouse embryos cultured in vitro. A fluorescein-labeled control MO was applied for monitoring the incorporation of MOs into developing 2-cell embryos in the presence of varying amounts of EP and bovine serum albumin. In optimized conditions, fluorescence was detected in 2-cell embryos within a 3-h incubation period. In order to analyze the validity of the optimized conditions, an antisense Oct4 MO was applied for knockdown of the synthesis of OCT4 protein in developing embryos from the 2-cell stage. In blastocysts, the antisense Oct4 MO induced a decrease in the amount in OCT4 protein to less than half. An almost complete absence of OCT4-positive cells and nearly complete disappearance of the inner cell mass in the outgrowths of blastocysts were also noted. These phenotypes corresponded with those of Oct4-deficient mouse embryos. Overall, we suggest that the delivery of MOs using EP is useful for the knockdown of gene expression in preimplantation mouse embryos cultured in vitro.     

Molecular orbital theory applied to the study of nonsteroidal anti-inflammatory drug efficiency.

Using a simple quantum mechanical method, we calculated the energy of the highest-occupied molecular orbital (E(HOMO)) of three groups of anti-inflammatory compounds, and we have found correlations between E(HOMO) of these molecules and experimental data previously reported on (1) inhibition of sheep-vesicular-gland prostaglandin cyclooxygenase by phenolic compounds, (2) inhibition of prostaglandin cyclooxygenase in mouse macrophages by salicylates, benzoates and phenols, and (3) peroxyl-radical scavenging and radioprotection of a bacterial virus by NSAID drugs, including metiazinic acid, sulindac, D-penicillamine, piroxicam, indomethacin, benoxaprofen, and aspirin. Our correlations using a systematic evaluation of the HOMO energies can be of predictive value in the search for new anti-inflammatory drugs as well as for new radioprotectors.     

In pursuit of negative Fukui functions: examples where the highest occupied molecular orbital fails to dominate the chemical reactivity

In our quest to explore molecules with chemically significant regions where the Fukui function is negative, we explored reactions where the frontier orbital that indicates the sites for electrophilic attack is not the highest occupied molecular orbital. The highest occupied molecular orbital (HOMO) controls the location of the regions where the Fukui function is negative, supporting the postulate that negative values of the Fukui function are associated with orbital relaxation effects and nodal surfaces of the frontier orbitals. Significant negative values for the condensed Fukui function, however, were not observed.

Quantitative assessment of the knockdown efficiency of morpholino antisense oligonucleotides in zebrafish embryos using a luciferase assay

Despite the broad use of morpholino antisense oligonucleotides (MO) to knockdown gene function in zebrafish embryos, the efficiency of this method has not been successfully assessed, particularly in the cases of translation-blocking MOs. In our current study, we describe a luciferase assay-based system that can monitor the MO knockdown levels in zebrafish by the use of a fusion reporter construct containing the 5'-mRNA sequence of the gene of interest and the luciferase coding sequence. The decrease in luciferase activity in zebrafish embryos that have been coinjected with this reporter RNA construct and a MO that targets the gene of interest correlated well with the level of inhibition of the corresponding endogenous protein synthesis, and also with the appearance of a knockdown phenotype. This indicates the usefulness of our method. We have also found that MOs can exert considerable knockdown effects upon unintended gene targets if 15 bases or longer of contiguous homology exists between these genes and the 25-base MO in question. Our quantitative assessment method also reveals, however, that an effective and specific knockdown can be achieved when employing a strategy that takes advantage of the synergistic effect of double MOs used at low levels.     

Quantum Chemical Studies of Structures and Binding in Noncanonical RNA Base pairs: The Trans Watson-Crick:Watson-Crick Family

Abstract

The trans Watson-Crick/Watson-Crick family of base pairs represent a geometric class that play important structural and possible functional roles in the ribosome, tRNA, and other functional RNA molecules. They nucleate base triplets and quartets, participate as loop closing terminal base pairs in hair pin motifs and are also responsible for several tertiary interactions that enable sequentially distant regions to interact with each other in RNA molecules. Eleven representative examples spanning nine systems belonging to this geometric family of RNA base pairs, having widely different occurrence statistics in the PDB database, were studied at the HF/6–31G (d, p) level using Morokuma decomposition, Atoms in Molecules as well as Natural Bond Orbital methods in the optimized gas phase geometries and in their crystal structure geometries, respectively. The BSSE and deformation energy corrected interaction energy values for the optimized geometries are compared with the corresponding values in the crystal geometries of the base pairs. For non protonated base pairs in their optimized geometry, these values ranged from ?8.19 kcal/mol to ?21.84 kcal/mol and compared favorably with those of canonical base pairs. The interaction energies of these base pairs, in their respective crystal geometries, were, however, lesser to varying extents and in one case, that of A:A W:W trans, it was actually found to be positive. The variation in RMSD between the two geometries was also large and ranged from 0.32–2.19 Å. Our analysis shows that the hydrogen bonding characteristics and interaction energies obtained, correlated with the nature and type of hydrogen bonds between base pairs; but the occurrence frequencies, interaction energies, and geometric variabilities were conspicuous by the absence of any apparent correlation. Instead, the nature of local interaction energy hyperspace of different base pairs as inferred from the degree of their respective geometric variability could be correlated with the identities of free and bound hydrogen bond donor/acceptor groups present in interacting bases in conjunction with their tertiary and neighboring group interaction potentials in the global context. It also suggests that the concept of isostericity alone may not always determine covariation potentials for base pairs, particularly for those which may be important for RNA dynamics. These considerations are more important than the absolute values of the interaction energies in their respective optimized geometries in rationalizing their occurrences in functional RNAs. They highlight the importance of revising some of the existing DNA based structure analysis approaches and may have significant implications for RNA structure and dynamics, especially in the context of structure prediction algorithms.     

Quantitative structure-activity relationship studies for antioxidant hydroxybenzalacetones by quantum chemical- and 3-D-QSAR(CoMFA) analyses

Antioxidant activities for a series of hydroxybenzalacetones, OH-BZ, evaluated by their inhibitory potencies against lipid peroxidation induced by gamma-ray irradiation or t-BuOOH, were analyzed quantitatively using quantum-chemical parameters calculated by semi-empirical molecular orbital (MO) calculations. The energy of the highest occupied molecular orbital (E(HOMO)) and frontier electron densities (HOMO) on the phenolic oxygen atom (F(H,O)), together with the steric parameter (E(s)) for the substituent ortho to the phenolic oxygen, showed excellent correlations. We also performed 3D-QSAR studies by using the comparative molecular field analysis (CoMFA) model. The results were compared with the corresponding classical QSAR correlations.     

Docking on the DNA G-quadruplex: a molecular electrostatic potential study

The G-quadruplexes are four-stranded nucleic acid structures with guanine-rich sequences that play important biological roles in, for example, regulating telomerase association and activity. Recent evidence supports the hypothesis that the telomeric G-quadruplex DNA represents a target of novel anticancer drug medication. In this work, we present results of the molecular electrostatic potential (MEP), together with the HOMO and LUMO frontier orbitals, which are physical quantities of concern in the docking of compounds on the G-quadruplex. The calculations are performed in the frame of density functional theory at the B88LYP/6-31G* level of theory. Additional functionals that introduce dispersion effects were also taken into consideration. The MEP potential and electron density of the frontier molecular orbitals of the G-quadruplex exhibit topological deformations due to the coiled conformation of the compound when they are compared with the MEP and corresponding electron density of a DNA duplex with similar nucleic acid composition. The electrostatic active zone of the G-quadruplex is localized on the top part of the quadruplex structure where the MEP acquires the most negative values. Additional computations on a set of three daunomycins, a common anticancer drug for duplex DNA, indicate an electrostatic fastening between the quadruplex and the set of daunomycins. In this regard, the G-quadruplex electrostatic interactions favor the stacking of ligands. Finally, some implications on molecular drug design are briefly discussed.     

A molecular orbital selection approach for fast calculations of specific rotation with density functional theory

In this work, we describe a simple approach to select the most important molecular orbitals (MOs) to compute the optical rotation tensor through linear response (LR) Kohn-Sham density functional theory (KS-DFT). Taking advantage of the iterative nature of the algorithms commonly used to solve the LR equations, we select the MOs with contributions to the guess perturbed density that are larger than a certain threshold and solve the LR equations with the selected MOs only. We propose two criteria for the selection, and two definitions of the selection threshold. We then test the approach with two functionals (B3LYP and CAM-B3LYP) and two basis sets (aug-cc-pVDZ and aug-cc-pVTZ) on a set of 51 organic molecules with specific rotation spanning five orders of magnitude, 10⁰–10⁴ deg (dm⁻¹ (g/mL)⁻¹). We show that this approach indeed can provide very accurate values of specific rotation with estimated speedup that ranges from 2 to 8× with the most conservative selection criterion, and up to 20 to 30× with the intermediate criterion.     

Targeting gene expression in the preimplantation mouse embryo using morpholino antisense oligonucleotides

Morpholino antisense oligonucleotides act by blocking translation of their target gene products and are effective tools for down-regulating gene expression. The current study was conducted to define treatment conditions for the use of morpholino oligonucleotides (MOs) in mammalian preimplantation embryos, and to employ MOs to target genes and study gene function in the early embryo. For the first time, ethoxylated polyethylenimine (EPEI), Lipofectin or Lysolecithin delivery agents were employed in combination with a fluorescent control MO and an alpha-catenin specific MO, to down-regulate gene expression during murine preimplantation development. Experiments applied to both two- and eight-cell stage murine preimplantation embryos contrasted the efficacy of MO concentrations of 1, 2, 5, 10, and 20 microM and treatment delivery times of 3, 6, 24, and 48 hr. Continuous treatment of two-cell embryos with Lipofectin and 20 microM alpha-catenin MO for 48 hr resulted in a significant (P < 0.05) reduction in development to the blastocyst stage and was accompanied by a marked reduction in alpha-catenin protein. These results indicate that morpholino antisense oligonucleotides are effective tools for down-regulating gene expression during mammalian preimplantation development.     

Quantum chemical studies of structures and binding in noncanonical RNA base pairs: the trans Watson-Crick:Watson-Crick family

The trans Watson-Crick/Watson-Crick family of base pairs represent a geometric class that play important structural and possible functional roles in the ribosome, tRNA, and other functional RNA molecules. They nucleate base triplets and quartets, participate as loop closing terminal base pairs in hair pin motifs and are also responsible for several tertiary interactions that enable sequentially distant regions to interact with each other in RNA molecules. Eleven representative examples spanning nine systems belonging to this geometric family of RNA base pairs, having widely different occurrence statistics in the PDB database, were studied at the HF/6-31G (d, p) level using Morokuma decomposition, Atoms in Molecules as well as Natural Bond Orbital methods in the optimized gas phase geometries and in their crystal structure geometries, respectively. The BSSE and deformation energy corrected interaction energy values for the optimized geometries are compared with the corresponding values in the crystal geometries of the base pairs. For non protonated base pairs in their optimized geometry, these values ranged from -8.19 kcal/mol to -21.84 kcal/mol and compared favorably with those of canonical base pairs. The interaction energies of these base pairs, in their respective crystal geometries, were, however, lesser to varying extents and in one case, that of A:A W:W trans, it was actually found to be positive. The variation in RMSD between the two geometries was also large and ranged from 0.32-2.19 A. Our analysis shows that the hydrogen bonding characteristics and interaction energies obtained, correlated with the nature and type of hydrogen bonds between base pairs; but the occurrence frequencies, interaction energies, and geometric variabilities were conspicuous by the absence of any apparent correlation. Instead, the nature of local interaction energy hyperspace of different base pairs as inferred from the degree of their respective geometric variability could be correlated with the identities of free and bound hydrogen bond donor/acceptor groups present in interacting bases in conjunction with their tertiary and neighboring group interaction potentials in the global context. It also suggests that the concept of isostericity alone may not always determine covariation potentials for base pairs, particularly for those which may be important for RNA dynamics. These considerations are more important than the absolute values of the interaction energies in their respective optimized geometries in rationalizing their occurrences in functional RNAs. They highlight the importance of revising some of the existing DNA based structure analysis approaches and may have significant implications for RNA structure and dynamics, especially in the context of structure prediction algorithms.     

Lessons from morpholino-based screening in zebrafish

Morpholino oligonucleotides (MOs) are an effective, gene-specific antisense knockdown technology used in many model systems. Here we describe the application of MOs in zebrafish (Danio rerio) for in vivo functional characterization of gene activity. We summarize our screening experience beginning with gene target selection. We then discuss screening parameter considerations and data and database management. Finally, we emphasize the importance of off-target effect management and thorough downstream phenotypic validation. We discuss current morpholino limitations, including reduced stability when stored in aqueous solution. Advances in MO technology now provide a measure of spatiotemporal control over MO activity, presenting the opportunity for incorporating more finely tuned analyses into MO-based screening. Therefore, with careful management, MOs remain a valuable tool for discovery screening as well as individual gene knockdown analysis.     

Structure and the energy of base pairing in non-natural bases of nucleic acids: the azaguanine-cytosine and azaadenine-thymine base pairs

Watson-Crick optimized geometries and the energies of base pairing for the natural pairs of nucleic bases: adenine-thymine (AT) and guanine-cytosine (GC) have been recalculated by ab initio methods in order to compare results to those found for the non-natural azaadenine-thymine (AAT) and azaguanine-cytosine (AGC) pairs. Geometry optimizations carried out at the HF/6-31G** level and energies obtained at MP2/6-31G**, show that AAT and AGC have hydrogen bonding patterns similar to the natural AT and GC and that the interaction energies (DeltaH0int) for the former are ca. 7 kcal/mol more stable than the latter. Accordingly, the pairs based on azapurines would be favored with respect to the natural pairs. Some possible explanations why nature does not use extensively the azabases in base pairing are given.     

Theoretical study on phthalocyanine,pyrazinoporphyrazine and their complexation with Mg2+ and Zn2+

We have studied the UV–vis absorption spectra of metal-free phthalocyanine (H₂Pc), metal-free pyrazinoporphyrazine (H₂PyzPz) and their complexes with Mg^2 +  and Zn^2 +  using semiempirical Zerners intermediate neglect of differential overlap and time-dependent density functional theory methods. The predicted absorption spectra of H₂Pc and their complexes are in agreement with a previous experiment report. The calculated results show that the Q band absorption peaks of H₂PyzPz and their complexes are blue-shifted by 40 nm as compared with those of H₂Pc and their complexes, respectively. The frontier molecular orbitals (HOMO and LUMO) of H₂Pc, H₂PyzPz and their metal complexes were investigated as well. The nitrogen atoms in the pyrazine rings stabilise the HOMO more than the LUMO, and the deprotonation of the pyrrole rings induced by the metal coordination destabilises the LUMO more than the HOMO. Because of the increased band gap, the absorption bands of H₂PyzPz and the metal-coordinated compounds are blue-shifted.