Conformational analysis of the 3'-5'-cyclic dinucleotide d less than pApA greater than by means of molecular mechanics

The conformational properties of the cyclic dinucleotide d less than pApA greater than were studied by means of molecular mechanics calculations in which a multiconformation analysis was combined with minimum energy calculations. In this approach models of possible conformers are built by varying the torsion angles of the molecule systematically. These models are then subjected to energy minimization; in the present investigation use was made of the AMBER Force field. It followed that the lowest energy conformer has a pseudo-two-fold axis of symmetry. In this conformer the deoxyribose sugars adopt a N-type conformation. The conformation of the sugar-phosphate backbone is determined by the following torsion angles: alpha +, beta t, gamma +, epsilon t and zeta +. The conformation of this ringsystem corresponds to the structure derived earlier by means of NMR spectroscopy and X-ray diffraction. The observation of a preference for N-type sugar conformations in this molecule can be explained by the steric hindrance induced between opposite H3' atoms when one sugar is switched from N- to S-type puckers. The sugars can in principle switch from N- to S-type conformations, but this requires at least the transition of gamma + to gamma -. In this process the molecule obtains an extended shape in which the bases switch from a pseudo-axial to a pseudo-equatorial position. The calculations demonstrate that, apart from the results obtained for the lowest energy conformation, the 180 degrees change in the propagation direction of the phosphate backbone can be achieved by several different combinations of the backbone torsion angles. It appeared that in the low energy conformers five higher order correlations are found. The combination of torsion angles which are involved in changes in the propagation direction of the sugar-phosphate backbone in DNA-hairpin loops and in tRNA, are found in the dataset obtained for cyclic d less than pApA greater than. It turns out, that in the available examples, 180 degrees changes in the backbone direction are localized between two adjacent nucleotides.     

Determination of the backbone torsion angle ɛ in nucleic acids

Summary The multiplet structure of cross peaks in double-quantum-filtered COSY NMR spectra is analysed for those resonances that include passive heteronuclear couplings. Interestingly, the cross peak involving the sugar-ring protons H2 and H3 in nucleic acids display an E.COSY-type appearance exclusively when the backbone torsion angle (C4-C3-O3-P) adopts a gauche(-) conformation. This observation allows an unambiguous analysis of the conformation around , without the knowledge of ³J_cp coupling constants.     

The three-dimensional structure of a DNA hairpin in solution two-dimensional NMR studies and structural analysis of d(ATCCTATTTATAGGAT)

The hairpin formed by d(ATCCTATTTATAGGAT) was studied by means of two-dimensional NMR spectroscopy and conformational analysis. Almost all 1H resonances of the stem region could be assigned, while the 1H and 31P spectra of the loop region were interpreted completely; this includes the stereospecific assignment of the H5' and H5" resonances. The derivation of the detailed loop structure was carried out in a stepwise fashion including some improved and new methods for structure determination from NMR data. In the first step, the mononucleotide structures were examined. The conformational space available to the mononucleotide was scanned systematically by varying the glycosidic torsion angle and pseudorotational parameters. Each generated conformer was tested against the experimental J coupling constants and NOE parameters. In the following stage, the structures of dinucleotides and longer fragments were derived. Inter-residue distances between protons were calculated by means of a procedure in which the simulated NOEs, obtained via a relaxation-matrix approach, were fitted to the experimental NOEs without the introduction of a molecular model. In addition, the backbone torsion angles beta, gamma and epsilon were deduced from homocoupling and heterocoupling constants. These data served as constraints in the next step, in which the loop sequence was subjected to a multi-conformer generation procedure. The resulting structures were tested against the mentioned constraints and disregarded if these constraints were violated. This yielded a family of structures for the loop region, confined to a relatively narrow conformational space. A representative conformation was subsequently docked on a B-type stem which fulfilled the structural constraints (derived from the NMR experiments for the stem region) to yield the hairpin structure. Results obtained from subsequent restrained-molecular-mechanics as well as free-molecular-mechanics calculations are in accordance with those obtained by means of the analysis described above. The structure of the hairpin loop is a compactly folded conformation and the first base of the central TTTA region forms a Hoogsteen T-A pair with the fourth base. This Hoogsteen base pair is stacked upon the sixth base pair of the B-type double-helical stem. The second base of the loop is folded into the minor groove, whereas the third base of the loop is partly stacked on the first and fourth bases. The phosphate backbone exhibits a sharp turn between the third and fourth nucleotides of the loop. The peculiar structure of this hairpin loop is discussed in relation to loop folding in DNA and RNA hairpins and in relation to a general model for loop folding.     

Amblyseius bibens (Acarina: Phytoseiidae), a predator of spider mites (Tetranychidae) in Madagascar

Field observation indicated thatA. bibens is an important predator of severalTetranychus-species in parts of Madagascar. Its dependence on water for drinking was demonstrated; this feature limits its presence to more humid biotopes. Its importance in the irrigated areas of the dry southwest is stressed.A. bibens proved capable of eradicating.T. urticae on potted bean plants. Predation and oviposition of the predator were tested with various densities of adult femaleT. urticae. At the lowest densities tested, predation was limited by the availability of the prey, which also resulted in a lower oviposition rate at these densities. At more moderate densities (of around 32 prey/7 cm²) an increase in prey egg consumption made up for a decrease in adult prey consumption, indicating that spider mite eggs are suitable food for this predator. At high densities eggs were ignored and adult prey were killed and at highest densities more were killed than were actually needed for food. The effect of alternate feeding on eggs and an interference stimulation component are claimed to explain the unusual functional response curve ofA. bibens. The results imply that, under natural conditions,A. bibens feeds primarily on spider mite eggs, but shifts towards adult prey consumption with increasing prey densities.

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Résumé Des observations dans la nature montrent queA. bibens est un important prédateur de plusieurs espèces deTetranychidae dans certains secteurs à Madagascar. Son importance dans les zones irriguées du sud-ouest aride est mise en évidence, ce qui résulte des besoins pour cette espèce d'eau alimentaire, ce facteur limitant sa présence aux biotopes les plus humides. A. bibens se montre capable d'éliminerT. urticae sur les cultures de pois. Le taux de prédation et d'oviposition du prédateur ont été testés en rapport aux diverses densités de la proie,T. urticae. Aux densités les plus faibles la prédation est limitée par la disponibilité en proies, ce qui entraine également une réduction de la fécondité. Pour des densités moyennes (de l'ordre de 32 proies/7 cm²) on observe une consommation préférentielle des ufs deTetranychidae et une moindre consommation des adultes. Pour des densités élevées, par contre, les ufs sont délaissés et ce sont les adultes qui sont consommés, enfin par des densités encore plus fortes, le prédateur détruit plus de proies adultes qu'il n'en peut consommer.L'effet d'une, alimentation de substitution sur les ufs de la proie et l'interférence d'un facteur stimulant sont envisagés comme éléments explicatifs de la réponse inhabituelle d'A. bibens. Cela implique que dans les conditions naturellesA. bibens se nourrit en premier lieu sur les ufs deTetranychidae, mais transfert sa préférence sur les proies adultes quand la densité de ces derniers s'accroît.

     

Karyotypes of eight species of phytoseiid mites (Acarina: Mesostigmata) from Madagascar

Haplo-diploidy was found to be regular with eight species of phytoseiid mites occurring in Madagascar. The chromosome number in all species was n=4. A variation was found with respect to centromere position.     

Conformational analysis of a dinucleotide photodimer with the aid of the genetic algorithm.

The solution structure of the photodimer cis,syn-dUp[]dT is derived with the aid of the genetic algorithm. The conformational space available for the molecule is sampled efficiently using the computer program DENISE and tested against a set of constraints available from nmr experiments. The dominant conformation in solution found with this approach can be described by the following combinations of sugar-phosphate backbone torsion angles: epsilon(t), zeta(t), alpha(+), beta(-ac), and gamma(t). The conformation of the sugars and glycosidic torsion angles are S type and syn, respectively. The cyclobutane ring and pyrimidines are puckered. In addition, other conformations that exist in equilibrium with the first are found. It is concluded that the cyclobutane-pyrimidine system is rigid, whereas the sugar-phosphate backbone is flexible. The solution structures are compared with the crystal structure of the strongly related cyano-ethyl ester of cis,syn-dTp[]dT.     

Prediction of flight of apple sawfly, Hoplocampa testudinea, using temperature sums

A method to predict the beginning of the flight of apple sawfly, Hoplocampa testudinea (Klug) (Hymenoptera, Tenthredinidae) is presented. Adults were monitored during 8 years at the experimental orchard De Schuilenburg, Kesteren, The Netherlands, with visual sticky traps. Temperature sums, accumulated until the day of the first capture of sawfly, were calculated for different starting dates of the summations. Beginning on 1 January, variation of the temperature sums, calculated in each of 8 years, decreased when later starting dates for temperature summation were chosen, reaching a minimum on 1 April for soil temperatures (–5 cm). The soil temperature sum, accumulated from 1 April until the first capture, was on average 134 day-degrees above a threshold of 4 °C. So this was the most suitable starting date to predict the beginning of sawfly flight activity. As use of the summation method based on air temperatures is in practice more appropriate for timing of trap installation by extension services and apple growers, trap deployment is recommended at an adapted value of 157 day-degrees from 15 March.     

Time efficient detection of protein–ligand interactions with the polarization optimized PO-WaterLOGSY NMR experiment

The identification of compounds that bind to a protein of interest is of central importance in contemporary drug research. For screening of compound libraries, NMR techniques are widely used, in particular the Water-Ligand Observed via Gradient SpectroscopY (WaterLOGSY) experiment. Here we present an optimized experiment, the polarization optimized WaterLOGSY (PO-WaterLOGSY). Based on a water flip-back strategy in conjunction with model calculations and numerical simulations, the PO-WaterLOGSY is optimized for water polarization recovery. Compared to a standard setup with the conventional WaterLOGSY, time consuming relaxation delays have been considerably shortened and can even be omitted through this approach. Furthermore, the robustness of the pulse sequence in an industrial setup was increased by the use of hard pulse trains for selective water excitation and water suppression. The PO-WaterLOGSY thus yields increased time efficiency by factor of 3–5 when compared with previously published schemes. These time savings have a substantial impact in drug discovery, since significantly larger compound libraries can be tested in screening campaigns. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Aspects of the molecular structure and dynamics of neuropeptide Y.

Human neuropeptide Y (hNPY) and the Q34-->P34 mutant (P34-hNPY) have been characterized by CD spectroscopy. hNPY self-associates in aqueous solution with a dimerization constant in the micromolar range. The self-association correlates with an increase in secondary-structure content which was studied as a function of concentration, temperature and pH. The effects of temperature were measured in water (5-84 degrees C) and in ethanediol/water (2 : 1) (-90 degrees to +90 degrees C). A single-residue mutation, Q34-->P34, affects the pH, thermal and self-association properties of NPY. The CD results are correlated with photochemically induced dynamic nuclear polarization NMR experiments which show that the tyrosines at the interface between two monomer units present limited accessibility to a photoreactive dye. An equilibrium state is described, involving a PP-fold monomer form and a handshake dimer form, that accommodates the physicochemical properties of NPY.