Biomolecular Simulations under Realistic Macroscopic Salt Conditions

Biomolecular simulations are typically performed in an aqueous environment where the number of ions remains fixed for the duration of the simulation, generally with either a minimally neutralizing ion environment or a number of salt pairs intended to match the macroscopic salt concentration. In contrast, real biomolecules experience local ion environments where the salt concentration is dynamic and may differ from bulk. The degree of salt concentration variability and average deviation from the macroscopic concentration remains, as yet, unknown. Here, we describe the theory and implementation of a Monte Carloosmostatthat can be added to explicit solvent molecular dynamics or Monte Carlo simulations to sample from a semigrand canonical ensemble in which the number of salt pairs fluctuates dynamically during the simulation. The osmostat reproduce the correct equilibrium statistics for a simulation volume that can exchange ions with a large reservoir at a defined macroscopic salt concentration. To achieve useful Monte Carlo acceptance rates, the method makes use of nonequilibrium candidate Monte Carlo (NCMC) moves in which monovalent ions and water molecules are alchemically transmuted using short nonequilibrium trajectories, with a modified Metropolis-Hastings criterion ensuring correct equilibrium statistics for an (Δµ, N, p, T) ensemble. We demonstrate how typical protein (DHFR and the tyrosine kinase Src) and nucleic acid (Drew-Dickerson B-DNA dodecamer) systems exhibit salt concentration distributions that significantly differ from fixed-salt bulk simulations and display fluctuations that are on the same order of magnitude as the average.

Synthesis, spectroscopic studies of novel N-substituted phthalimides and evaluation of their antibacterial, antioxidant, DNA binding and molecular docking studies

<p class="Abstract">A new series of N-substituted phthalimide derivatives were prepared by condensation of appropriate amount of n-amino tetrachlorophthalimide with respective aldehyde in glacial acetic acid. The structural investigation of the synthesized compounds was done by spectroscopic methods (UV-Vis., IR, ¹H and ¹³C NMR) and elemental analysis. The antibacterial screening of these compounds was performed against Escherichia coli and Staphylococcus mutans. The synthesized compounds were evaluated for their antioxidant potential using 2,2-diphenyl-1-picrylhydrazyl (DPPH) as a scavenging agent. The interaction ability of most promising compounds (3a and 3b) with native calf thymus DNA (Ct-DNA) was also studied by means of UV-Vis., circular dichroism (CD), viscosity measurements and thermal studies. The intrinsic binding constants (K_b) of 3a and 3b with Ct-DNA obtained from UV-Vis. absorption studies were 8 × 10⁴ and 1 × 10⁵, respectively. Molecular docking of target compounds (3a and 3b) against DNA dodecamer d(CGCGAATTCGCG)₂ has been carried out. The test compounds exhibited remarkable antibacterial, antioxidant and DNA binding activities.</p><p> </p>

Terminal lipophilization of a unique DNA dodecamer by various nucleolipid headgroups: Their incorporation into artificial lipid bilayers and hydrodynamic properties

A series of six cyanine-5-labeled oligonucleotides (LONs 10–15), each terminally lipophilized with different nucleolipid head groups, were synthesized using the recently prepared phosphoramidites 4b–9b. The insertion of the LONs within an artificial lipid bilayer, composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE), was studied by single molecule fluorescence spectroscopy and microscopy with the help of an optically transparent microfluidic sample carrier with perfusion capabilities. The incorporation of the lipo-oligonucleotides into the bilayer was studied with respect to efficiency (maximal bilayer brightness) as well as stability against perfusion (final stable bilayer brightness). Attempts to correlate these parameters with the log P values of the corresponding nucleolipid head groups failed, a result which clearly demonstrates that not only the lipophilicity but mainly the chemical structure and topology of the head group is of decisive importance for the optimal interaction of a lipo-oligonucleotide with an artificial lipid bilayer. Moreover, fluorescence half-live and diffusion time values were measured to determine the diffusion coefficients of the lipo-oligonucleotides.

Structural flexibility revealed by ultra-high resolution data

Not many macromolecular crystals diffract X-rays to ultra-high resolution, defined usually as higher than 0.8 Å, and in the Protein Data Bank there are currently 43 such submissions. These structures range in size from antibiotics of about a hundred atoms to proteins with more than 3,000 independent atoms in the asymmetric unit of the crystal cell. The unprecedented data resolution reveals a great wealth of structural details, which cannot be visualized by analyses at lower resolution. The accuracy of the refined stereochemical and geometrical parameters is then comparable with values typical for small-molecular crystallography and exceeds the accuracy of the library of the standard restraint target values, routinely used in refinement of proteins and nucleotides. Somewhat unexpectedly, the very high resolution diffraction does not necessarily relates to extreme stability of the crystallized molecules, so that the obtained electron density maps reveal significant parts of the atomic models existing in multiple conformations, slightly differing from each other. For example, about 1/3 of the protein chain in the 0.65 Å structure of lysozyme [1] and majority of phosphate groups in the 0.75 Å structure of Z-DNA dodecamer [2] could be modeled in double conformations.

Phosphates in the Z-DNA dodecamer are flexible, but their P-SAD signal is sufficient for structure solution

A large number of Z-DNA hexamer duplex structures and a few oligomers of different lengths are available, but here the first crystal structure of the d(CGCGCGCGCGCG)2dodecameric duplex is presented. Two synchrotron data sets were collected; one was used to solve the structure by the single-wavelength anomalous dispersion (SAD) approach based on the anomalous signal of P atoms, the other set, extending to an ultrahigh resolution of 0.75 Å, served to refine the atomic model to anRfactor of 12.2% and anRfreeof 13.4%. The structure consists of parallel duplexes arranged into practically infinitely long helices packed in a hexagonal fashion, analogous to all other known structures of Z-DNA oligomers. However, the dodecamer molecule shows a high level of flexibility, especially of the backbone phosphate groups, with six out of 11 phosphates modeled in double orientations corresponding to the two previously observed Z-DNA conformations: ZI, with the phosphate groups inclined towards the inside of the helix, and ZII, with the phosphate groups rotated towards the outside of the helix.