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Biomedical subjects

D Sprous

Publications and source records attributed to D Sprous.

7 recordsLinked to original sources
PubMed1999

Molecular dynamics studies of axis bending in d(G5-(GA4T4C)2-C5) and d(G5-(GT4A4C)2-C5): effects of sequence polarity on DNA curvature.

Gel retardation studies and other experiments indicate that DNA sequences containing the d(GA4T4C)n motif are curved, whereas those of identical composition but with a reverse sequence polarity, the d(GT4A4C)n motif, are straight. Hydroxyl radical cleavage experiments show that d(GA4T4C)n shows a unique signature, whereas d(GT4A4C)n behaves normally. To explain these results at a molecular level, molecular dynamics (MD) simulations were performed on the DNA duplexes d(G5-(GA4T4C)2-C5) and d(G5-(GT4A4C)2-C5) to 3.0 and 2.5 ns, respectively. The MD simulations are based on the Cornell force field implemented in the AMBER 4.1 modeling package and performed in a neutral solution of anionic DNA with K+, Cl- and Mg2+ at concentrations roughly comparable to a ligase buffer. Long range interactions were treated by the particle mesh Ewald method. Analysis of the results shows that the calculated dynamical structure of d(G5-(GA4T4C)2-C5) exhibits strong gross curvature, consistent with the observed behavior. The most significant locus of curvature in the MD structure is found at the central C15-G16 step, with an average roll angle of 12.8(+/-6.40)deg. The d(G5-(GT4A4C)2-C5) MD structure exhibited significantly less gross curvature. Analysis of results indicates that the reduction in gross curvature in the d(G5-(GT4A4C)2-C5) trajectory originates from the effect of the T10-A11 and T20-A21 steps, which showed average roll angles of 12.5(+/-5)deg. These three steps, T10-A11, C15-G16 and T20-A21, are half-helix turns away from one another, and their contributions to concerted bending cancel out. The A-tracts in the MD structure are essentially straight. The dynamical structure of d(G5-(GA4T4C)2-C5) exhibited minor groove deformation comprised of expansion at the 5' end of A-tracts and progressive narrowing towards the 3' end, consistent with and elaborating the interpretation of hydroxyl radical chemical probing results.

Base Sequence↗
PubMed1996

Molecular modeling of closed circular DNA thermodynamic ensembles.

Many modeling studies of supercoiled DNA are based on equilibrium structures from theoretical calculations or energy minimization. Since closed circular DNAs are flexible, it is possible that errors are introduced by calculating properties from a single minimum energy structure, rather than from a complete thermodynamic ensemble. We have investigated this question using molecular dynamics simulations on a low resolution molecular mechanics model in which each base pair is represented by three points (a plane). This allows the inclusion of sequence-dependent variations of tip, inclination, and twist. Three kinds of sequences were tested: (1) homogeneous DNA, in which all base pairs have the helicoidal parameters of an ideal, average B-DNA; (2) random sequence DNA; and (3) curved DNA. We examined the rate of convergence of various structural parameters. Convergence for most of these is slowest for homogeneous sequences, more rapid for random sequences, and most rapid for curved sequences. The most slowly converging parameter is the antipodes profile. In a plasmid with N base pairs (bp), the antipodes distance is the distance dij from base pair i to base pair j halfway around the plasmid, j = i + N/2. The antipodes profile at time tau is a plot of dij over the range i = 1, N/2. In a homogeneous plasmid, convergence requires that the antipodes profile averaged over time must be flat. Even in the small plasmids examined here, the average properties of the ensembles were found to differ from those of static equilibrium structures. These effects will be even more dramatic for larger plasmids. Further, average and dynamic properties are affected by both plasmid size and sequence.

Chemical Phenomena↗
PubMed1996

Molecular dynamics simulations of small DNA plasmids: effects of sequence and supercoiling on intramolecular motions.

Small (600 base pair) DNA plasmids were modeled with a simplified representation (3DNA) and the intramolecular motions were studied using molecular mechanics and molecular dynamics techniques. The model is detailed enough to incorporate sequence effects. At the same time, it is simple enough to allow long molecular dynamics simulations. The simulations revealed that large-scale slithering occurs in a homogeneous sequence. In a heterogeneous sequence, containing numerous small intrinsic curves, the centers of the curves are preferentially positioned at the tips of loops. With more curves than loop tips (two in unbranched supercoiled DNA), the heterogeneous sequence plasmid slithers short distances to reposition other curves into the loop tips. However, the DNA is immobilized most of the time, with the loop tips positioned over a few favored curve centers. Branching or looping also appears in the heterogeneous sequence as a new method of repositioning the loop tips. Instead of a smooth progression of increasing writhing with increasing linking difference, theoretical studies have predicted that there is a threshold between unwrithed and writhed DNA at a linking difference between one and two. This has previously been observed in simulations of static structures and is demonstrated here for dynamic homogeneous closed DNA. Such an abrupt transition is not found in the heterogeneous sequence in both the static and dynamic cases.

DNA, Superhelical↗
PubMed1996

Action at a distance in supercoiled DNA: effects of sequence on slither, branching, and intramolecular concentration.

We report a computer modeling study of DNA supercoiling in model plasmids over the size range of 140-1260 bp. We used a computer model with basepair resolution. Molecular dynamics was used to produce ensembles at 300 K and to investigate intramolecular motions. The plasmid models varied by their sequence. The sequence types employed for comparison included a curve-bearing plasmid, a heterogenous sequence plasmid, and a homogenous sequence. Within the three sequence types tested at the 1260-bp plasmid size, we observed several sequence-dependent phenomena. Writhe, radius of gyration, slither motion, and branching probability were seen to be sequence dependent. Branching probability was the least in the homogenous plasmid and the greatest in the curve-bearing plasmid. The curve imposed a symmetry on the plasmid that was absent in the heterogenous sequence. Significant localizations and enhancements of intramolecular concentration were seen to a persistence length. Molecular dynamics allowed us to observe the mechanism of branch formation and reabsorption. We observed a size-dependent change in the types of motion observed in plasmids. Slither motion predominated in plasmids up to 600 bp in size, whereas global rearrangements were more important in the 1260 mer.

Base Sequence↗
PubMed1995

Dehydrating agents sharply reduce curvature in DNAs containing A tracts.

The structural basis of DNA curvature remains elusive, because models for curvature based on crystallographic structures of molecules containing A tracts do not agree with any of the models for sequence-directed curvature based on solution studies. Here we demonstrate that the difference is probably due to MPD (2-methyl-2,4-pentanediol), the dehydrating agent commonly used in crystallography. One characteristic signature of curved DNA molecules is that they run anomalously slowly on polyacrylamide gels, appearing to be larger than they actually are. The gel anomalies of three curved DNAs from trypanosome kinetoplast minicircles drop monotonically with increasing MPD concentration, indicating that MPD straightens molecules that are curved in aqueous solution. This is not due to some non-specific effect of MPD on poly(dA) or polypurine tracts, because control molecules containing dA70 and dG43 run normally over the full range of MPD concentrations. Circular dichroism spectra are not affected by MPD, ruling out a conformational change to a structure outside the B-DNA family. The effect is not due to MPD-induced changes in phasing of the curved sequences, because MPD has virtually no effect on the linking numbers of relaxed plasmids containing either curved sequences or dA70. At the concentrations of MPD used in X-ray crystallography, the curvature of DNAs containing A tracts is substantially lower than in solution, which probably explains the ongoing discrepancies between the crystallographic results and models based on solution studies.

Animals↗
PubMed1995

What is the basis of sequence-directed curvature in DNAs containing A tracts?

A variety of solution and gel experiments show that DNAs containing tracts of 4-8 A's repeated in phase with the helix repeat are curved. Several independent analyses of these experiments argue that curvature resides in the A tracts themselves. In x-ray crystallographic studies of several DNAs containing A tracts, however, the A tracts are uncurved, leading to models in which curvature resides in the non-A tracts. This "curved general sequence model" has several problems, in our view. We review those, and we describe recent experiments that show that the dehydrating agents commonly used in x-ray crystallography markedly reduce curvature in gels and in solution, calling into question the ability of crystallography to determine the structural basis of DNA curvature. Finally, we discuss the critical role of hydration in curved DNAs and suggest new experiments that we hope could finally determine exactly which sequences are responsible for curvature.

Adenine↗