A conformational analysis of daunomycin considering D-ring repucker.

We present a complete molecule mechanics optimization of daunomycin. Two different D-ring puckers are found to be of comparable energy, consistent with NMR data, although only one of these low-energy structures has been observed by X-ray crystallography. Our results are more consistent with the previous conformational analysis of daunomycin by Neidle and Taylor (Neidle, S. and Taylor, G.L. (1979) FEBS Lett. 107, 348-354) than that of Nakata and Hopfinger (Nakata, Y. and Hopfinger, A.J. (1980) FEBS Lett. 117, 259-264).

Temperature-dependent molecular dynamics and restrained X-ray refinement simulations of a Z-DNA hexamer.

Molecular dynamics simulations of the Z-DNA hexamer 5BrdC-dG-5BrdC-dG-5BrdC-dG were performed at several temperatures between 100 K and 300 K. Above 250 K, a strong sequence-dependent flexibility in the nucleic acid is observed, with the guanine sugar and the phosphate of GpC sequences much more mobile than the cytosine sugar and phosphate of CpG sequences. At 300 K, the hexamer is in dynamic equilibrium between several Z forms, including the crystallographically determined ZI and ZII forms. The local base-pair geometry, however, is not very variable, except for the roll of the base-pairs. The hexamer molecular dynamics trajectories have been used to test the restrained parameter crystallographic refinement model for nucleic acids. X-ray diffraction intensities corresponding to observed diffraction data were computed. The average structures obtained from the simulations were then refined against the calculated intensities, using a restrained least-squares program developed for nucleic acids in order to analyse the effects of the refinement model on the derived quantities. In general, the temperature dependence of the atomic fluctuations determined directly from the refined Debye-Waller factors is in reasonably good agreement with the results obtained by calculating the atomic fluctuations directly from the Z-DNA molecular dynamics trajectories. The agreement is best for refinement of temperature factors without restraints. At the highest temperature studied (300 K), the effect of the refinement on the most mobile atoms (phosphates) is to significantly reduce the mean-square atomic fluctuations estimated from the refined Debye-Waller factors below the actual values (less than (delta r)2 greater than congruent to 0.5 A2). Analysis of the temperature-dependence of the mean-square atomic fluctuations provides information concerning the conformational potential within which the atoms move. The calculated temperature-dependence and anharmonicity of the Z-DNA helix are compared with the results observed for proteins. The average structures from the simulations were refined against the experimental X-ray intensities. It is found that low-temperature molecular dynamics simulations provide a useful tool for optimizing the refinement of X-ray structures.

Theoretical studies of the structure and energies of base-paired nucleotides and the dissociation kinetics of a proflavine-dinucleotide complex.

We presented calculations of base-paired dinucleoside phosphates and hexanucleoside pentaphosphates of varying compositions. Complete energy minimizations were performed for (a) the ten base-pair combinations of dinucleoside phosphates, starting from a B-DNA conformation, (b) six hexanucleoside pentaphosphates--base-paired CGCGCG, GCGCGC, G6-C6, TATATA, ATATAT, and A6-T6--starting with a B-DNA geometry, and (c) the four hexanucleoside pentaphosphates that have alternating pyrimidine-purine sequences, starting with a Z-DNA geometry. In addition, we studied the proflavine-base-paired CpG complex, using both complete energy minimization and energetic constraints to force the drug to dissociate from the dinucleoside phosphate. In many of these calculations, we examined the dependence of the calculated energies and structures on the potential function, focusing mainly on the effect of nonbonded potentials, the effective dielectric constant, and the role of counterions. These calculations allow us to explain why pur-(3',5')-pyr sequence isomers are more stable than pyr-(3'-5')-pur isomers. Both base-base and base-backbone energies are important in this differentiation, with the former being mainly van der Waals attraction and the latter mainly electrostatic energies. The calculations also allow us to understand the differences in double helical stabilities found by Wells et al. These differences, caused by electrostatic interactions between those bases not Watson-Crick hydrogen bonded, allow us to explain the following experimental data: poly(dG-dC) melts 12 degrees C higher than poly dG-poly dC, poly(dA-dT) melts 6 degrees C lower than poly dA-poly dT, and poly(dA-dG)-poly(dC-dT) melts 6 degrees C lower than poly(dA-dC)-poly(dT-dG). These results have interesting implications for drug binding: they imply that simple intercalators, such as ethidium, will exhibit a greater affinity for hetero- than for homopolymers and that this preference will be greater in the AT polymers than it is in the GC polymers. Our calculations allow us to explain the fact that Z-DNA is more stable than B-DNA under high salt conditions and to suggest some sequence dependence for the Z to B transition. We found that the activation energy for proflavine dissociating from dCpG is almost equal to the dissociation energy.

Molecular mechanical studies of DNA flexibility: coupled backbone torsion angles and base-pair openings.

Molecular mechanics studies have been carried out on "B-DNA-like" structures of [d(C-G-C-G-A-A-T-T-C-G-C-G)](2) and [d(A)](12).[d(T)](12). Each of the backbone torsion angles (psi, phi, omega, omega', phi') has been "forced" to alternative values from the normal B-DNA values (g(+), t, g(-), g(-), t conformations). Compensating torsion angle changes preserve most of the base stacking energy in the double helix. In a second part of the study, one purine N3-pyrimidine N1 distance at a time has been forced to a value of 6 A in an attempt to simulate the base opening motions required to rationalize proton exchange data for DNA. When the 6-A constraint is removed, many of the structures revert to the normal Watson-Crick hydrogen-bonded structure, but a number are trapped in structures approximately 5 kcal/mol higher in energy than the starting B-DNA structure. The relative energy of these structures, some of which involve a non-Watson-Crick thymine C2(carbonyl)[unk]adenine 6NH(2) hydrogen bond, are qualitatively consistent with the DeltaH for a "base pair-open state" suggested by Mandal et al. of 4-6 kcal/mol [Mandal, C., Kallenbach, N. R. & Englander, S. W. (1979) J. Mol. Biol. 135, 391-411]. The picture of DNA flexibility emerging from this study depicts the backbone as undergoing rapid motion between local torsional minima on a nanosecond time scale. Backbone motion is mainly localized within a dinucleoside segment and generally not conformationally coupled along the chain or across the base pairs. Base motions are much smaller in magnitude than backbone motions. Base sliding allows imino N-H exchange, but it is localized, and only a small fraction of the N-H groups is exposed at any one time. Stacking and hydrogen bonding cause a rigid core of bases in the center of the molecule accounting for the hydrodynamic properties of DNA.

Electrostatic potential molecular surfaces.

Color-coded computer graphics representations of the electrostatic potentials of trypsin, trypsin-inhibitor, prealbumin and its thyroxine complex, fragments of double-helical DNA, and a netropsin--DNA complex illustrate the electrostatic and topographic complementarity in macromolecule-ligand interactions. This approach is powerful in revealing intermolecular specificity and shows promise of having predictive value in drug design.

Electrostatic recognition between superoxide and copper, zinc superoxide dismutase.

Electrostatic forces have been implicated in a variety of biologically important molecular interactions including drug orientation by DNA, protein folding and assembly, substrate binding and catalysis and macromolecular complementarity with inhibitors, drugs and hormones. To examine enzyme-substrate interactions in copper, zinc superoxide dismutase (SOD), we developed a method for the visualization and analysis of an enzyme's three-dimensional electrostatic vector field that allows the contributions of specific residues to be identified. We report here that the arrangement of electrostatic charges in SOD promotes productive enzyme-substrate interaction through substrate guidance and charge complementarity: sequence-conserved residues create an extensive electrostatic field that directs the negatively charged superoxide (O-2) substrate to the highly positive catalytic binding site at the bottom of the active-site channel. Dissection of the electrostatic potential gradient indicated the relative contributions of individual charged residues: Lys 134 and Glu 131 seem to have important roles in directing the long-range approach of O-2, while Arg 141 has local orienting effects. The reported methods of analysis may have general application for the elucidation of intermolecular recognition processes.

A hypothetical model of the flavodoxin-tetraheme cytochrome c3 complex of sulfate-reducing bacteria.

A hypothetical model of the flavodoxin-tetraheme cytochrome c3 electron-transfer complex from the sulfate-reducing bacterium Desulfovibrio vulgaris has been constructed by using interactive computer graphics based on electrostatic potential field calculations and previous NMR experiments. Features of the proposed complex are (1) van der Waals contact between the flavin mononucleotide prosthetic group of flavodoxin and one heme of the cytochrome, (2) unique complementarity of electrostatic fields between the region surrounding this heme and the region surrounding the exposed portion of the flavin mononucleotide group of flavodoxin, and (3) no steric interferences between the two polypeptide chains in the complex. This complex is consistent with all structural and spectroscopic data available.

Electron transport in sulfate-reducing bacteria. Molecular modeling and NMR studies of the rubredoxin--tetraheme-cytochrome-c3 complex.

A hypothetical model of the complex formed between the iron-sulfur protein rubredoxin and the tetraheme cytochrome c3 from the sulfate-reducing bacteria Desulfovibrio vulgaris (Hildenborough) has been proposed utilizing computer graphic modeling, computational methods and NMR spectroscopy. The proposed complex appears feasible on the basis of complementary electrostatic interaction and steric factors and is consistent with the data from NMR experiments. In this model, the non-heme iron atom of rubredoxin is in close proximity to heme 1 of cytochrome c3. The complex is stabilized by charge-pair interactions and hydrogen bonds. This complex is compared to the flavodoxin-cytochrome c3 complex previously proposed [Stewart, D. E., LeGall, J., Moura, I., Moura, J. J. G., Peck, H. D. Jr, Xavier, A. V., Weiner, P. K. & Wampler, J. E. (1988) Biochemistry 27, 2444-2450] and new NMR data shows that both proteins interact with the same heme group of the cytochrome as postulated.