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W K Olson

Publications and source records attributed to W K Olson.

At least 37 records · Page 2Linked to original sources
PubMed1997

Minor groove-directed and intercalative ligand-DNA interactions in the poisoning of human DNA topoisomerase I by protoberberine analogs.

Spectroscopic, calorimetric, DNA cleavage, electrophoretic, and computer modeling techniques have been employed to characterize the DNA binding and topoisomerase poisoning properties of three protoberberine analogs, 8-desmethylcoralyne (DMC), 5,6-dihydro-8-desmethylcoralyne (DHDMC), and palmatine, which differ in the chemical structures of their B- and/or D-rings. DNA topoisomerase-mediated cleavage assays revealed that these compounds were unable to poison mammalian type II topoisomerase. By contrast, the three protoberberine analogs poisoned human topoisomerase I according to the following hierarchy: DHDMC > DMC > palmatine. DNA binding by all three protoberberine analogs induced negative flow linear dichroism signals as well as unwinding of the host duplex. These two observations are consistent with an intercalative mode of protoberberine binding to duplex DNA. However, a comparison of the DNA binding properties for DMC and DHDMC, which differ only by the state of saturation at the 5,6 positions of the B-ring, revealed that the protoberberine analogs do not "behave" like classic DNA intercalators. Specifically, saturation of the 5-6 double bond in the B-ring of DMC, thereby converting it to the DHDMC molecule, was associated with enhanced DNA unwinding as well as a reversal of DNA binding preference from a DNA duplex with an inaccessible or occluded minor groove {poly[d(G-C)]2} to DNA duplexes with accessible or unobstructed minor grooves {poly[d(A-T)]2 and poly[d(I-C)]2}. In addition, a comparison of the DNA binding properties for DHDMC and palmatine revealed that transferring the 11-methoxy moiety on the D-ring of DHDMC to the 9 position, thereby converting it to palmatine, was associated with a reduction in binding affinity for both duplexes with unobstructed minor grooves as well as for duplexes with occluded minor grooves. These DNA binding properties are consistent with a "mixed-mode" DNA binding model for protoberberines in which a portion of the ligand molecule intercalates into the double helix, while the nonintercalated portion of the ligand molecule protrudes into the minor groove of the host duplex, where it is thereby available for interactions with atoms lining the floor and/or walls of the minor groove. Furthermore, saturation at the 5,6 positions of the B-ring, which causes the A-ring to be tilted relative to the plane formed by rings C and D, appears to stabilize the interaction between the host duplex and the minor groove-directed portion of the protoberberine ligand. Computer modeling studies on the DHDMC-poly[d(A-T)]2 complex suggest that this interaction may involve van der Waals contacts between the ligand A-ring and backbone sugar atoms lining the minor groove of the host duplex. The hierarchy of topoisomerase I poisoning noted above suggests that this minor groove-directed interaction may play an important role in topoisomerase I poisoning by protoberberine analogs. In the aggregate, our results presented here, coupled with the recent demonstration of topoisomerase I poisoning by minor groove-binding terbenzimidazoles [Sun, Q., Gatto, B., Yu, C., Liu, A. , Liu, L. F., & LaVoie, E. J. (1995) J. Med. Chem. 38, 3638-3644], suggest that minor groove-directed ligand-DNA interactions may be of general importance in the poisoning of topoisomerase I.

Berberine Alkaloids↗
PubMed1997

Properties of ideal composite knots.

The shortest tube of constant diameter that can form a given knot represents the 'ideal' form of the knot. Ideal knots provide an irreducible representation of the knot, and they have some intriguing mathematical and physical features, including a direct correspondence with the time-averaged shapes of knotted DNA molecules in solution. Here we describe the properties of ideal forms of composite knots-knots obtained by the sequential tying of two or more independent knots (called factor knots) on the same string. We find that the writhe (related to the handedness of crossing points) of composite knots is the sum of that of the ideal forms of the factor knots. By comparing ideal composite knots with simulated configurations of knotted, thermally fluctuating DNA, we conclude that the additivity of writhe applies also to randomly distorted configurations of composite knots and their corresponding factor knots. We show that composite knots with several factor knots may possess distinct structural isomers that can be interconverted only by loosening the knot.

Computer Simulation↗
PubMed1997

Modeling protein-induced configurational changes in DNA minicircles.

A method is offered for obtaining minimum energy configurations of DNA minicircles constrained by one or more DNA-binding proteins. The minicircles are modeled as elastic rods, while the presence of bound protein is implied by rigidly fixing portions of these chains. The configurations of the geometrically constrained circular rods are sampled stochastically and optimized according to a simple elastic energy model of nicked DNA. The shapes of the minimum energy structures identified after a simulated annealing process are analyzed in terms of relative protein orientation and writhing number. The procedure is applied to minicircles 500 base pairs in length, bound to two evenly spaced DNA-wrapping proteins. The presence of histone octamers is suggested by rigidly fixing the two protein-bound portions of each minicircle as small superhelices similar in dimension to nucleosomal DNA. The folded minimum energy forms of sample chains with different degrees of protein wrapping are noteworthy in themselves in that they offer a new resolution to the well-known minichromosome linking number paradox and point to future minicircle simulations of possible import.

DNA, Circular↗
PubMed1997

Configurational transitions in Fourier series-represented DNA supercoils.

A new Fourier series representation of supercoiled DNA is employed in Langevin dynamics simulations to study large-scale configurational motions of intermediate-length chains. The polymer is modeled as an ideal elastic rod subject to long-range van der Waals' interactions. The van der Waals' term prevents the self-contact of distant chain segments and also mimics attractive forces thought to stabilize the association of closely spaced charged rods. The finite Fourier series-derived polymer formulation is an alternative to the piecewise B-spline curves used in past work to describe the motion of smoothly deformed supercoiled DNA in terms of a limited number of independent variables. This study focuses on two large-scale configurational events: the interconversion between circular and figure-8 forms at a relatively low level of supercoiling, and the transformation between branched and interwound structures at a higher superhelical density.

Biophysical Phenomena↗
PubMed1997

Modulation of nucleic acid structure by ligand binding: induction of a DNA.RNA.DNA hybrid triplex by DAPI intercalation.

The aromatic diamidine, DAPI (4',6-diamidino-2-phenylindole), is used as an important biological and cytological tool since it forms highly fluorescent complexes with nucleic acid duplexes via minor groove-directed/intercalative modes of interaction. In this study, we find that DAPI binding can induce the formation of an RNA-DNA hybrid triplex that would not otherwise form. More specifically, through application of a broad range of spectroscopic, viscometric, and molecular modeling techniques, we demonstrate that DAPI intercalation induces the formation of the poly(dT).poly(rA).poly(dT) hybrid triple helix, a structure which does not form in the absence of the ligand. Using UV mixing studies, we demonstrate that, in the presence of DAPI, the poly(rA).poly(dT) duplex and the poly(dT) single strand form a 1:1 complex (a triplex) that does not form in the absence of DAPI. Through temperature-dependent absorbance measurements, we show that the poly(dT).poly(rA).poly(dT) triplex melts via two distinct transitions: initial conversion of the triplex to the duplex state, with the DAPI remaining bound, followed by denaturation of the duplex-DAPI complex to its component single strands and free DAPI. Using optical melting profiles, we show that DAPI binding enhances the thermal stability of the poly(dT).poly(rA).poly(dT) triplex, an observation consistent with the preferential binding of the ligand to the triplex versus the duplex and single-stranded states. Our differential scanning calorimetric measurements reveal melting of the DAPI-saturated poly(dT).poly(rA).poly(dT) triplex to be associated with a lower enthalpy but greater cooperativity than melting of the corresponding DAPI-saturated poly(rA).poly(dT) duplex. Our flow linear dichroism and viscometric data are consistent with an intercalative mode of binding when DAPI interacts with both the poly(dT).poly(rA).poly(dT) triplex and the poly(rA).poly(dT) duplex. Finally, computer modeling studies suggest that a combination of both stacking and electrostatic interactions between the intercalated ligand and the host nucleic acid play important roles in the DAPI-induced stabilization of the poly(dT).poly(rA).poly(dT) triplex. In the aggregate, our results demonstrate that ligand binding can be used to induce the formation of triplex structures that do not form in the absence of the ligand. This triplex-inducing capacity has potentially important implications in the design of novel antisense, antigene, antiviral, and diagnostic strategies.

Calorimetry, Differential Scanning↗
PubMed1996

Simulating DNA at low resolution.

The past year has witnessed the development of several new mathematical approaches to analyzing the structure of double-helical DNA and to incorporating the sequence-dependent features of the chain in computer simulations of long polymers. Of special interest in this respect are the local and global structural changes induced by the binding of various proteins to DNA, ranging from subtle bending, untwisting and sliding motions at the base-pair level to the apparent organization of supercoiled structure in chains that are thousands residues long. The computational effort has also included both new ways to incorporate the polyelectrolyte character of DNA and other environmental forces in simulations of long chains and new methods to keep track of the multitude of configurations so generated. The collective advances are pointing to ways that will soon connect the sequences of base pairs in large genomes to folded three-dimensional structures based on natural bending, twisting and translational tendencies and in response to deformations produced by the binding of different proteins.

Base Sequence↗
PubMed1995

B-DNA twisting correlates with base-pair morphology.

The observed sequence dependence of the mean twist angles in 38 B-DNA crystal structures can be understood in terms of simple geometrical features of the constituent base-pairs. Structures with low twist appear to unwind in response to severe steric clashes of large exocyclic groups (such as NH2-NH2) in the major and minor grooves, while those with high twist are subjected to lesser contacts (H-O and H-H). We offer a simple clash function that depends on base-pair morphology (i.e. the chemical constitution of base-pairs) and satisfactorily accounts for the twist angles of the ten common Watson-Crick dimer steps both in the solid state and in solution. The twist-clash correlation that we find here still holds when extended to modified bases. In addition to Calladine's purine-purine clashes, we add other close contacts between bases in the grooves, and consider the conformational restrictions on the geometry of the sugar-phosphate backbone (namely, we emphasize the tendency of DNA to conserve virtual backbone length). The significance of this finding is threefold: (1) sequence-dependent DNA twisting is directly involved in protein-DNA interactions; (2) strong correlation between Twist and Roll helps to elucidate the bending of the double helix as a function of base sequence; (3) it is possible to anticipate the effects of chemical modifications on twisting and bending. The mutual correlations of other structural parameters with the twist make this angle a primary determinant of DNA conformational heterogeneity.

Base Sequence↗
PubMed1995

Effects of localized bending on DNA supercoiling.

The DNA double helix is straight only in the idealized case. In reality, it bends, twists and stretches in response to local base sequence and to specific interactions with proteins and other bound ligands. Naturally occurring bends appear to promote the assembly of nucleosomes, and in some cases can effectively replace regulatory DNA-binding proteins in vivo. Recently, a computational method known as 'finite element analysis', which is used routinely by engineers to analyse the stability of buildings and bridges, has been applied to the quantitative assessment of natural curvature in supercoiled DNA structures, providing new insight into the relationship between local, sequence-dependent features and the overall topology of these chains.

Computer Simulation↗
PubMed1994

Computer simulation of protein-induced structural changes in closed circular DNA.

The effect of protein-induced wrapping on overall DNA folding is studied using Monte Carlo computer simulation techniques. A new modeling scheme is devised to represent configurations of closed circular DNA containing fragments of the double helix partially wrapped around a core of proteins. The DNA consists of two regions, a fragment wrapped in a left-handed superhelical path around a 'phantom' protein core and a free connecting loop. The loop has at least one single-stranded scission so that it can assume a torsionally relaxed state. The configuration of the loop is varied during the course of the computer simulations and the three-dimensional spatial arrangements of lowest total energy are identified. The axis of the DNA loop is represented by a finite three-dimensional Fourier series perturbation of an initial Bézier curve, making it possible to fix the position and orientation of the chain ends as well as the contour length of the free loop. The energy is approximated by elastic terms for the bending and twisting of the DNA and an excluded volume contribution that prevents the self-intersection of sequentially distant chain segments. The proportions of the protein-DNA complex, the number of superhelical turns, the chain length and the imposed linking number difference of the closed DNA are varied in the calculations. The resulting minimum energy structures are consistent with physical models and suggest new ways to think about how proteins add and remove supercoils from DNA. Of special note in this regard is the sudden collapse of three-dimensional structure that accompanies small incremental wrapping of the DNA around the idealized protein core. These observations offer new structural insight into the mechanisms of action of proteins which add or remove supercoils from DNA and provide a first step in thinking about the activity of such systems at the chemical level whereby small fluctuations in local molecular structure are translated into large-scale macromolecular changes. The configurations identified in the simulations can also be examined in the context of the well known "linking number paradox" associated with nucleosome formation on closed circular plasmids. The findings bear relevance to DNA with natural curvature as well as to protein-induced bending and deformations of the double helix.

Computer Simulation↗
PubMed1994

Computer models of DNA four-way junctions.

A modeling scheme that combines a constrained backbone generating algorithm with simple hard-sphere packing calculations is offered to build the four-stranded structures of DNA found in Holliday junctions. Two standard B-DNA duplexes are oriented side by side with helix axes at different relative inclinations and then systematically rotated and translated to identify closely spaced contact-free states. Attempts are subsequently made to introduce a low-energy sugar-phosphate linkage that serves as the site of strand exchange between the two duplexes. The chemical connection is sought using an algorithm which identifies the possible arrangements of the intervening backbone torsions between arbitrarily positioned bases. The goal is to identify the multiple conformational solutions associated with a particular arrangement of neighboring DNA helices in the four-way junction rather than a single optimum structure. The methodology is general, in terms of accommodating four-way junctions with arms of variable conformation and chain length and of dimensions much greater than treated heretofore. The only deformation in the four-way structures relative to B-DNA occurs at the site of backbone exchange, with base stacking and Watson-Crick pairing completely preserved in all models. The arrangements of neighboring bases at these sites resemble the unusual conformational steps found in a number of small molecule nucleic acid crystal structures. An interesting outcome of the calculations is the formation of sterically acceptable four-arm Holliday junctions over a wide range of angles at the cross. The potential mobility of the Holliday junctions is inferred from visualization and energetic analysis of the various models. Long-range electrostatic energies based on different currently available treatments of the dielectric constant are used to estimate the conformational preferences and flexibility of the four-stranded structures. The various dielectric schemes, however, are not in complete agreement on the likely conformational variability of the four-way junctions. The structures suggest a possible mechanism for branch migration and detail a pathway linking the antiparallel uncrossed Holliday structure inferred from solution measurements and the parallel cross-packed helical arrangements observed in single-crystal X-ray studies.

Algorithms↗
PubMed1994

The effect of mathematics and coordinate system on comparability and "dependencies" of nucleic acid structure parameters.

This paper critically examines the methodologies used to analyze nucleic acid three-dimensional structure based on guidelines set at a 1988 EMBO workshop. The implications of these analyses cannot be fully understood without a thorough knowledge of how the numbers are calculated. This paper addresses one aspect of the calculations, namely the observed correlations between various parameters. These correlations are addressed in the mathematics by explicitly incorporating the concept of a pivot point, which is the point about which a base rotates as it buckles, propeller twists and opens. Pivot points enable one to model the physical motion of bases more accurately. As a result, they greatly reduce and/or eliminate the statistical correlations between rotational and translational parameters found in other approaches. The correlations that are reduced or eliminated are actually artifacts of the mathematics employed and do not reflect true structural properties of nucleic acids. The mathematics we have developed, including the mathematics of pivot points, are presented in the companion paper. Here, we explain how some of the observed correlations occur as a by-product of the method of calculation, while others are truly structural, and we show how optimum pivot points can be determined to minimize artifactual correlations. The observation that experimental bases often rotate about the long axis in a "propeller" motion as well as rotate about the Z-axis of each base, "opening" into the major groove, is evident in the location of the optimum region for the pivot point as determined in this study. We consider locating a pivot point as a calibration step to increase the agreement between physical intuition and the mathematics of our program.

Base Sequence↗
PubMed1994

Nucleic acid structure analysis. Mathematics for local Cartesian and helical structure parameters that are truly comparable between structures.

Analyzing nucleic acid structures in a comparable manner has become increasingly important as the number of solved structures has increased. This paper presents the concepts, mathematics, theorems, and proofs that form the basis of a new program to analyze three-dimensional DNA and RNA structures. The approach taken here provides numerical data in accordance with guidelines set at a 1988 EMBO workshop. Mathematical definitions are provided for all local structural parameters described in the guidelines. The definitions satisfy the guideline requirements while preserving the original physical intuition of the parameters. In particular, the rotational parameters are true rotations based on a simple physical model (net rotation at constant angular velocity), not Euler angles or angles between vectors and planes as is the case with other approaches. As a result, the mathematical definitions are symmetrical with the property that a 5 degrees tilt is the same as a 5 degrees roll and a 5 degrees twist, except that the rotations take place about different axes. In other approaches, a 5 degrees tilt can mean a different amount of net rotation than a 5 degrees roll or a 5 degrees twist. A second unique feature of the mathematics is that it explicitly incorporates the concept of a pivot point, which is the point about which a base in a base-pair rotates as it buckles, propeller twists, and opens. Pivot points enable one to model the physical motion of bases more accurately. As a result, they greatly reduce and/or eliminate the statistical correlations between rotational and translational parameters that arise as mathematically induced artifacts in other approaches. This paper, together with the statistical analysis in the companion paper for determining the locations of the pivot points, provides everything needed to understand the output of the program as it relates to individual structures.

Base Composition↗
PubMed1994

Configurational statistics of the DNA duplex: extended generator matrices to treat the rotations and translations of adjacent residues.

The base-to-base virtual bond treatment of nucleic acids used in statistical mechanical calculations of polynucleotide chain properties has been refined by incorporating the six parameters that relate the positions and orientations of sequential rigid bodies. The scheme allows for the sequence-dependent bending, twisting, and displacement of base pairs as well as for asymmetry in the angular and translational fluctuations of individual residues. Expressions are developed for the generator matrices required for the computation, as a function of chain length, of various parameters measuring the overall mean extension and shape of the DNA. Quantities of interest include the end-to-end vector r, the square of the end-to-end distance r2, the square radius of gyration s2, the center-of-gravity vector g, the second moments of inertia Sx2, and the higher moments of r and g. The matrix expressions introduced in the 1960s by Flory and co-workers for the determination of configuration-dependent polymer chain averages are decomposed into their translational and orientational contributions so that the methods can be extended to the rigid body analysis of chemical moieties. The new expressions permit, for the first time, examination of the effects of sequence-dependent translations, such as the lateral sliding of residues in A- and B-helices and the vertical opening of base pairs in drug-DNA complexes, on the average extension and shape of the long flexible double helix. The approach is in the following paper using conformational energy estimates of the base sequence-dependent flexibility of successive B-DNA base pairs.

Biometry↗
PubMed1994

Spatial translational motions of base pairs in DNA molecules: application of the extended matrix generator method.

We have used the elementary generator matrices outlined in the preceding paper to examine the conformational plasticity of the nucleic acid double helix. Here we investigate kinked DNA structures made up of alternating B- and A-type helices and intrinsically curved duplexes perturbed by the intercalation of ligands. We model the B-to-A transition by the lateral translation of adjacent base pairs, and the intercalation of ligands by the vertical displacement of neighboring residues. We report a complete set of average configuration-dependent parameters, ranging from scalars (i.e., persistence lengths) to first- and second-order tensor parameters (i.e., average second moments of inertia), as well as approximations of the associated spatial distributions of the DNA and their angular correlations. The average structures of short chains (of lengths less than 100 base pairs) with local kinks or intrinsically curved sequences are essentially rigid rods. At the smallest chain lengths (10 base pairs), the kinked and curved chains exhibit similar average properties, although they are structurally perturbed compared to the standard B-DNA duplex. In contrast, at lengths of 200 base pairs, the curved and kinked chains are more compact on average and are located in a different space from the standard B- or A-DNA helix. While A-DNA is shorter and thicker than B-DNA in x-ray models, the long flexible A-DNA helix is thinner and more extended on average than its B-DNA counterpart because of more limited fluctuations in local structure. Curved polymers of 50 base pairs or longer also show significantly greater asymmetry than other DNAs (in terms of the distribution of base pairs with respect to the center of gravity of the chain). The intercalation of drugs in the curved DNA straightens and extends the smoothly deformed template. The dimensions of the average ellipsoidal boundaries defining the configurations of the intercalated polymers are roughly double those of the intrinsically curved chain. The altered proportions and orientations of these density functions reflect the changing shape and flexibility of the double helix. The calculations shed new light on the possible structural role of short A-DNA fragments in long B-type duplexes and also offer a model for understanding how GC-specific intercalative ligands can straighten naturally curved DNA. The mechanism is not immediately obvious from current models of DNA curvature, which attribute the bending of the chain to a perturbed structure in repeating tracts of A.T base pairs.

Base Sequence↗
PubMed1994

On higher buckling transitions in supercoiled DNA.

A combination of detailed energy minimization and molecular dynamics studies of closed circular DNA offers here new information that may be relevant to the dynamics of short DNA chains and/or low superhelical densities. We find a complex dependence of supercoiled DNA energies and geometries on the linking number difference delta Lk as physiological superhelical densities (magnitude of sigma approximately 0.06) are approached. The energy minimization results confirm and extend predictions of classical elasticity theory for the equilibria of elastic rods. The molecular dynamics results suggest how these findings may affect the dynamics of supercoiled DNA. The minimization reveals sudden higher order configurational transitions in addition to the well-known catastrophic buckling from the circle to the figure-8. The competition among the bending, twisting, and self-contact forces leads to different families of supercoiled forms. Some of those families begin with configurations of near-zero twist. This offers the intriguing possibility that nicked DNA may relax to low-twist forms other than the circle, as generally assumed. Furthermore, for certain values of delta Lk, more than one interwound DNA minimum exists. The writhing number as a function of delta Lk is discontinuous in some ranges; it exhibits pronounced jumps as delta Lk is increased from zero, and it appears to level off to a characteristic slope only at higher values of delta Lk. These findings suggest that supercoiled DNA may undergo systematic rapid interconversions between different minima that are both close in energy and geometry. Our molecular dynamics simulations reveal such transitional behavior. We observe the macroscopic bending and twisting fluctuations of interwound forms about the global helix axis as well as the end-over-end tumbling of the DNA as a rigid body. The overall mobility can be related to magnitude of sigma and to the bending, twisting, and van der Waals energy fluctuations. The general character of molecular motions is thus determined by the types of energy minima found at a given delta Lk. Different time scales may be attributed to each type of motion: The overall chain folding occurs on a time scale almost an order of magnitude faster than the end-over-end tumbling. The local bending and twisting of individual chain residues occur at an even faster rate, which in turn correspond to several cycles of local variations for each large-scale bending and straightening motion of the DNA.

DNA, Superhelical↗
PubMed1994

Electrostatic effects in short superhelical DNA.

We present Monte Carlo simulations of the equilibrium configurations of short closed circular DNA that obeys a combined elastic, hard-sphere, and electrostatic energy potential. We employ a B-spline representation to model chain configuration and simulate the effects of salt on chain folding by varying the Debye screening parameter. We obtain global equilibrium configurations of closed circular DNA, with several imposed linking number differences, at two salt concentrations (specifically at the extremes of no added salt and the high salt regime), and for different chain lengths. Minimization of the composite elastic/long-range potential energy under the constraints of ring closure and fixed chain length is found to produce structures that are consistent with the configurations of short supercoiled DNA observed experimentally. The structures generated under the constraints of an electrostatic potential are less compact than those subjected only to an elastic term and a hard-sphere constraint. For a fixed linking number difference greater than a critical value, the interwound structures obtained under the condition of high salt are more compact than those obtained under the condition of no added salt. In the case of no added salt, the electrostatic energy plays a dominant role over the elastic energy in dictating the shape of the closed circular DNA. The DNA supercoil opens up with increasing chain length at low salt concentration. A branched three-leaf rose structure with a fixed linking number difference is higher in energy than the interwound form at both salt concentrations employed here.

DNA, Superhelical↗
PubMed1994

The influence of salt on the structure and energetics of supercoiled DNA.

We present a detailed computational study of the influence of salt on the configurations, energies, and dynamics of supercoiled DNA. A potential function that includes both elastic and electrostatic energy components is employed. Specifically, the electrostatic term, with salt-dependent coefficients, is modeled after Stigter's pioneering work on the effective diameter of DNA as a function of salt concentration. Because an effective charge per unit length is used, the electrostatic formulation does not require explicit modeling of phosphates and can be used to study long DNAs at any desired resolution of charge. With explicit consideration of the electrostatic energy, an elastic bending constant corresponding to the nonelectrostatic part of the bending contribution to the persistence length is used. We show, for a series of salt concentrations ranging from 0.005 to 1.0 M sodium, how configurations and energies of supercoiled DNA (1000 and 3000 base pairs) change dramatically with the simulated salt environment. At high salt, the DNA adopts highly compact and bent interwound states, with the bending energy dominating over the other components, and the electrostatic energy playing a minor role in comparison to the bending and twisting terms. At low salt, the DNA supercoils are much more open and loosely interwound, and the electrostatic components are dominant. Over the range of three decades of salt examined, the electrostatic energy changes by a factor of 10. The buckling transition between the circle and figure-8 is highly sensitive to salt concentration: this transition is delayed as salt concentration decreases, with a particularly sharp increase below 0.1 M. For example, for a bending-to-twisting force constant ratio of A/C = 1.5, the linking number difference (delta LK) corresponding to equal energies for the circle and figure-8 increases from 2.1 to 3.25 as salt decreases from 1.0 to 0.005 M. We also present in detail a family of three-lobed supercoiled DNA configurations that are predicted by elasticity theory to be stable at low delta Lk. To our knowledge, such three-dimensional structures have not been previously presented in connection with DNA supercoiling. These branched forms have a higher bending energy than the corresponding interwound configurations at the same delta Lk but, especially at low salt, this bending energy difference is relatively small in comparison with the total energy, which is dominated by the electrostatic contributions. Significantly, the electrostatic energies of the three-lobed and (straight) interwound forms are comparable at each salt environment. We show how the three-lobed configurations change slowly with ALk, resulting in branched interwound forms at higher salt. In longer chains, the branched forms are highly interwound, with bent arms. At low salt, the branched supercoils are asymmetric, with a longer interwound stem and two shorter arms. From molecular dynamics simulations we observe differences in the motions of the DNA as a function of salt. At high salt, the supercoiled chain is quite compact but fairly rigid, whereas at low salt the DNA is loosely coiled but more dynamic. Especially notable at low salt are the large-scale opening and closing of the chain as a whole and the rapid "slithering"of individual residues past one another. Toroidal forms are not detected under these conditions. However, the overall features of the open, loose supercoils found at low salt are more similar to those of toroidal than interwound configurations. Indeed,simulated x-ray scattering profiles reveal the same trends observed experimentally and are consistent with a change from closed to open forms as salt is decreased. Like the minimization studies, the dynamics reveal a critical point near 0.1 M associated with the collapse of loose to tight supercoils. Near this physiological concentration, enhanced flexibility of the DNA is noted. The collective observations suggest a potential regulatory role for salt on supercoiled DNA function, not only for closed circular DNA,but also for linear DNA with small looped regions.

Bacteriophage phi X 174↗