G. Ravishanker

A 5-NANOSECOND MOLECULAR DYNAMICS TRAJECTORY FOR B-DNA : ANALYSIS OF STRUCTURE, MOTIONS, AND SOLVATION

We report the results of four new molecular dynamics (MD) simulations on the DNA duplex of sequence d(CGCGAATTCGCG)2, including explicit consideration of solvent water, and a sufficient number of Na+ counterions to provide electroneutrality to the system. Our simulations are configured particularly to characterize the latest MD models of DNA, and to provide a basis for examining the sensitivity of MD results to the treatment of boundary conditions, electrostatics, initial placement of solvent, and run lengths. The trajectories employ the AMBER 4.1 force field. The simulations use particle mesh Ewald summation for boundary conditions, and range in length from 500 ps to 5.0 ns. Analysis of the results is carried out by means of time series for conformationalm, helicoidal parameters, newly developed indices of DNA axis bending, and groove widths. The results support a dynamically stable model of B-DNA for d(CGCGAATTCGCG)2 over the entire length of the trajectory. The MD results are compared with corresponding crystallographic and NMR studies on the d(CGCGAATTCGCG)2 duplex, and placed in the context of observed behavior of B-DNA by comparisons with the complete crystallographic data base of B-form structures. The calculated distributions of mobile solvent molecules, both water and counterions, are displayed. The calculated solvent structure of the primary solvation shell is compared with the location of ordered solvent positions in the corresponding crystal structure. The results indicate that ordered solvent positions in crystals are roughly twice as structured as bulk water. Detailed analysis of the solvent dynamics reveals evidence of the incorporation of ions in the primary solvation of the minor groove B-form DNA. The idea of localized complexation of otherwise mobile counterions in electronegative pockets in the grooves of DNA helices introduces an additional source of sequence-dependent effects on local conformational, helicoidal, and morphological structure, and may have important implications for understanding the functional energetics and specificity of the interactions of DNA and RNA with regulatory proteins, pharmaceutical agents, and other ligands.

Analysis of local helix bending in crystal structures of DNA oligonucleotides and DNA-protein complexes

Sequence-dependent bending of the helical axes in 112 oligonucleotide duplex crystal structures resident in the Nucleic Acid Database have been analyzed and compared with the use of bending dials, a computer graphics tool. Our analysis includes structures of both A and B forms of DNA and considers both uncomplexed forms of the double helix as well as those bound to drugs and proteins. The patterns in bending preferences in the crystal structures are analyzed by base pair steps, and emerging trends are noted. Analysis of the 66 B-form structures in the Nucleic Acid Database indicates that uniform trends within all pyrimidine-purine and purine-pyrimidine steps are not necessarily observed but are found particularly at CG and GC steps of dodecamers. The results support the idea that AA steps are relatively straight and that larger roll bends occur at or near the junctions of these A-tracts with their flanking sequences. The data on 16 available crystal structures of protein-DNA complexes indicate that the majority of the DNA bends induced via protein binding are sharp localized kinks. The analysis of the 30 available A-form DNA structures indicates that these structures are also bent and show a definitive preference for bending into the deep major groove over the shallow minor groove.

Molecular dynamics studies of DNA

Abstract Progress has recently been made towards the development of an accurate theoretical model of DNA structure and motions on the basis of molecular dynamics computer simulations. Recent quantitative comparisons between calculated properties with those observed in crystal structure determinations and nuclear magnetic resonance experiments allow improvement of modelling techniques. Current issues in molecular dynamics methodology, particularly truncation of potentials, are discussed, followed by a survey of results from in vacuo and in aquo treatments of solvent. The results of recent in aquo DNA trajectories extended into the nanosecond regime, together with a comparison of calculated and observed axis bending behavior, close the gap between. the timescale of molecular dynamics simulation and NMR techniques. Examples illustrating the analysis of DNA molecular dynamics simulations are included and discussed.

Differential stability of beta-sheets and alpha-helices in beta-lactamase: a high temperature molecular dynamics study of unfolding intermediates

beta-Lactamase, which catalyzes beta-lactam antibiotics, is prototypical of large alpha/beta proteins with a scaffolding formed by strong noncovalent interactions. Experimentally, the enzyme is well characterized, and intermediates that are slightly less compact and having nearly the same content of secondary structure have been identified in the folding pathway. In the present study, high temperature molecular dynamics simulations have been carried out on the native enzyme in solution. Analysis of these results in terms of root mean square fluctuations in cartesian and [phi, psi] space, backbone dihedral angles and secondary structural hydrogen bonds forms the basis for an investigation of the topology of partially unfolded states of beta-lactamase. A differential stability has been observed for alpha-helices and beta-sheets upon thermal denaturation to putative unfolding intermediates. These observations contribute to an understanding of the folding/unfolding processes of beta-lactamases in particular, and other alpha/beta proteins in general.

Molecular Dynamics Simulations of DNA and a Protein-DNA Complex Including Solvent

The results of a recent nanosecond (ns) molecular dynamics (MD) simulation of the d(CGCGAATTCGCG) double helix in water and a 100 ps MD study of the λ repressor-operator complex are described. The DNA simulations are analyzed in terms of the structural dynamics, fluctuations in the groove width and bending of the helical axis. The results indicate that the ns dynamical trajectory progresses through a series of three substates of B form DNA, with lifetimes of the order of hundreds of picoseconds (ps). An incipient dynamical equilibrium is evident. A comparison of the calculated axis bending with that observed in corresponding crystal structure data is presented. Simulation of the DNA in complex with the protein and that of the free DNA in solution, starting from the crystal conformation, reveal the dynamical changes that occur on complex formation.

Molecular Dynamics Information Extraction

MD-trajectories currently span nanoseconds of simulated time. The parameters most commonly presented in MD papers are RMS, energy and temperature presented as functions of time. These are easily obtained from a variety of sources, most commonly the output files of the simulation itself. However, the most critical structural parameters are not available directly from program output files, but require post processing. Typically, these parameters are specific for a given molecule type. Pucker angle and other helicoidals are important for DNA. Ramachandran plots are obviously relevant for proteins. Hydrogen bond distances would be relevant for a ligand-DNA or ligand-protein simulation. The various modeling packages normally incorporate an analysis program such as the AMBER CARNAL program. The MD-ToolChest (MDTC) suite of programs was designed to analyze data from a broad range of sources and can read trajectory files from AMBER, GROMOS and CHARMm. For DNA, its flagship program dials_and_windows_dna serves as a front end for passing coordinates to theCURVES (Lavery and Sklenar, 1996) program. Complete helicoidal parameters are presented as time series. For proteins, a variety of torsional parameters will be analyzed and presented as graphs. AMBER4.1/PME simulations involving the “Hagerman” (1986) sequences d[G 5 – {GA 4 T 4 C}2 – C 5] and d[G 5 – {GT 4 A 4 C}2 – C 5] are presented here to illustrate the process involved in using MDTC2.0 and CURVES. Experimental evidence characterizes the d[GA 4 T 4 C] n as having a compressed minor groove and significant curvature. Conversely, d[GT 4 A 4 C] n lacks these features. MDTC provides utilities to determine RMS, to extract pdb files from AMBER trajectory files, and to view the complete set of DNA helicoidal parameters. CURVES analysis is capable of returning measures of curvature and groove width, which are critical in comparing experiment to simulation. A web page auxiliary (http://linus.chem.wesleyan.edu/ima/mdinfo.html) has been created which holds a complete set of MDTC2.0 example scripts, complete output and links to other programs.