Protein environment affects the water-tryptophan binding mode. MD, QM/MM, and NMR studies of engrailed homeodomain mutants.

Water molecules can interact with aromatic moieties using either their O-H bonds or their lone-pairs of electrons. In proteins, water-π interactions have been reported to occur with tryptophan and histidine residues, and dynamic exchange between O-Hπ hydrogen bonding and lone-pairπ interactions was suggested to take place, based on ab initio calculations. Here we used classical and QM/MM molecular dynamics simulations, complemented with an NMR study, to examine a specific water-indole interaction observed in the engrailed homeodomain and in its mutants. Our simulations indicate that the binding mode between water and indole can adapt to the potential created by the surrounding amino acids (and by the residues at the DNA surface in protein-DNA complexes), and support the model of dynamic switching between the O-Hπ hydrogen bonding and lone-pairπ binding modes.

Structural dynamics of the box C/D RNA kink-turn and its complex with proteins: the role of the A-minor 0 interaction, long-residency water bridges, and structural ion-binding sites revealed by molecular simulations.

Kink-turns (K-turns) are recurrent elbow-like RNA motifs that participate in protein-assisted RNA folding and contribute to RNA dynamics. We carried out a set of molecular dynamics (MD) simulations using parm99 and parmbsc0 force fields to investigate structural dynamics of the box C/D RNA and its complexes with two proteins: native archaeal L7ae protein and human 15.5 kDa protein, originally bound to very similar structure of U4 snRNA. The box C/D RNA forms K-turn with A-minor 0 tertiary interaction between its canonical (C) and noncanonical (NC) stems. The local K-turn architecture is thus different from the previously studied ribosomal K-turns 38 and 42 having A-minor I interaction. The simulations reveal visible structural dynamics of this tertiary interaction involving altogether six substates which substantially contribute to the elbow-like flexibility of the K-turn. The interaction can even temporarily shift to the A-minor I type pattern; however, this is associated with distortion of the G/A base pair in the NC-stem of the K-turn. The simulations show reduction of the K-turn flexibility upon protein binding. The protein interacts with the apex of the K-turn and with the NC-stem. The protein-RNA interface includes long-residency hydration sites. We have also found long-residency hydration sites and major ion-binding sites associated with the K-turn itself. The overall topology of the K-turn remains stable in all simulations. However, in simulations of free K-turn, we observed instability of the key C16(O2')-A7(N1) H-bond, which is a signature interaction of K-turns and which was visibly more stable in simulations of K-turns possessing A-minor I interaction. It may reflect either some imbalance of the force field or it may be a correct indication of early stages of unfolding since this K-turn requires protein binding for its stabilization. Interestingly, the 16(O2')-7(N1) H- bond is usually not fully lost since it is replaced by a water bridge with a tightly bound water, which is adenine-specific similarly as the original interaction. The 16(O2')-7(N1) H-bond is stabilized by protein binding. The stabilizing effect is more visible with the human 15.5 kDa protein, which is attributed to valine to arginine substitution in the binding site. The behavior of the A-minor interaction is force-field-dependent because the parmbsc0 force field attenuates the A-minor fluctuations compared to parm99 simulations. Behavior of other regions of the box C/D RNA is not sensitive to the force field choice. Simulation with net-neutralizing Na(+) and 0.2 M excess salt conditions appear in all aspects equivalent. The simulations show loss of a hairpin tetraloop, which is not part of the K-turn. This was attributed to force field limitations.

Single Stranded Loops of Quadruplex DNA As Key Benchmark for Testing Nucleic Acids Force Fields.

We have carried out a set of explicit solvent molecular dynamics (MD) simulations on two DNA quadruplex (G-DNA) molecules, namely the antiparallel d(G4T4G4)2 dimeric quadruplex with diagonal loops and the parallel-stranded human telomeric monomolecular quadruplex d[AGGG(TTAGGG)3] with three propeller loops. The main purpose of the paper was testing of the capability of the MD simulation technique to describe single-stranded topologies of G-DNA loops, which represent a very challenging task for computational methods. The total amount of conventional and locally enhanced sampling (LES) simulations analyzed in this study exceeds 1.5 µs, while we tested several versions of the AMBER force field (parm99, parmbsc0, and a version with modified glycosidic χ torsion profile) and the CHARMM27 force field. Further, we compared minimal salt and excess salt simulations. Postprocessing MM-PBSA (Molecular Mechanics, Poisson-Boltzmann, Surface Area) free energy calculations are also reported. None of the presently available force fields is accurate enough in describing the G-DNA loops. The imbalance is best seen for the propeller loops, as their experimental structure is lost within a few ns of standard simulations with all force fields. Among them, parmbsc0 provides results that are clearly closest to the experimental target values but still not in full agreement. This confirms that the improvement of the γ torsional profile penalizing the γ trans substates in the parmbsc0 parametrization was a step in the right direction, albeit not sufficient to treat all imbalances. The modified χ parametrization appears to rigidify the studied systems but does not change the ultimate outcome of the present simulations. The structures obtained in simulations with the modified χ profile are predetermined by its combination with either parm99 or parmbsc0. Experimental geometries of diagonal loops of d(G4T4G4)2 are stable in standard simulations on the ∼10 ns time scale but are becoming progressively lost in longer and LES simulations. In addition, the d(G4T4G4)2 quadruplex contains, besides the three genuine binding sites for cations in the channel of its stem, also an ion binding site at each stem-loop junction. This arrangement of five cations in the quadruplex core region is entirely unstable in all 24 simulations that we attempted. Overall, our results confirm that G-DNA loops represent one of the most difficult targets for molecular modeling approaches and should be considered as reference structures in any future studies aiming to develop or tune nucleic acids force fields.

Molecular dynamics simulations and their application to four-stranded DNA.

This review provides a critical assessment of the advantages and limitations of modeling methods available for guanine quadruplex (G-DNA) molecules. We characterize the relations of simulations to the experimental techniques and explain the actual meaning and significance of the results. The following aspects are discussed: pair-additive approximation of the empirical force fields, sampling limitations stemming from the simulation time and accuracy of description of base stacking, H-bonding, sugar-phosphate backbone and ions by force fields. Several methodological approaches complementing the classical explicit solvent molecular dynamics simulations are commented on, including enhanced sampling methods, continuum solvent methods, free energy calculations and gas phase simulations. The successes and pitfalls of recent simulation studies of G-DNA are demonstrated on selected results, including studies of cation interactions and dynamics of G-DNA stems, studies of base substitutions (inosine, thioguanine and mixed tetrads), analysis of possible kinetic intermediates in folding pathway of a G-DNA stem and analysis of loop regions of G-DNA molecules.

Impact of an extruded nucleotide on cleavage activity and dynamic catalytic core conformation of the hepatitis delta virus ribozyme.

The self-cleaving hepatitis delta virus (HDV) ribozyme is essential for the replication of HDV, a liver disease causing pathogen in humans. The catalytically critical nucleotide C75 of the ribozyme is buttressed by a trefoil turn pivoting around an extruded G76. In all available crystal structures, the conformation of G76 is restricted by stacking with G76 of a neighboring molecule. To test whether this crystal contact introduces a structural perturbation into the catalytic core, we have analyzed approximately 200 ns of molecular dynamics (MD) simulations. In the absence of crystal packing, the simulated G76 fluctuates between several conformations, including one wherein G76 establishes a perpendicular base quadruplet in the major groove of the adjacent P1 stem. Second-site mutagenesis experiments suggest that the identity of the nucleotide in position 76 (N76) indeed contributes to the catalytic activity of a trans-acting HDV ribozyme through its capacity for hydrogen bonding with P1. By contrast, in the cis-cleaving genomic ribozyme the functional relevance of N76 is less pronounced and not correlated with the P1 sequence. Terbium(III) footprinting and additional MD show that the activity differences between N76 mutants of this ribozyme are related instead to changes in average conformation and modified cross-correlations in the trefoil turn.

Molecular dynamics simulations of RNA: an in silico single molecule approach.

RNA molecules are now known to be involved in the processing of genetic information at all levels, taking on a wide variety of central roles in the cell. Understanding how RNA molecules carry out their biological functions will require an understanding of structure and dynamics at the atomistic level, which can be significantly improved by combining computational simulation with experiment. This review provides a critical survey of the state of molecular dynamics (MD) simulations of RNA, including a discussion of important current limitations of the technique and examples of its successful application. Several types of simulations are discussed in detail, including those of structured RNA molecules and their interactions with the surrounding solvent and ions, catalytic RNAs, and RNA-small molecule and RNA-protein complexes. Increased cooperation between theorists and experimentalists will allow expanded judicious use of MD simulations to complement conceptually related single molecule experiments. Such cooperation will open the door to a fundamental understanding of the structure-function relationships in diverse and complex RNA molecules. .

Calculation of structural behavior of indirect NMR spin-spin couplings in the backbone of nucleic acids.

Calculated indirect NMR spin-spin coupling constants (J-couplings) between (31)P, (13)C, and (1)H nuclei were related to the backbone torsion angles of nucleic acids (NAs), and it was shown that J-couplings can facilitate accurate and reliable structural interpretation of NMR measurements and help to discriminate between their distinct conformational classes. A proposed stepwise procedure suggests assignment of the J-couplings to torsion angles from the sugar part to the phosphodiester link. Some J-couplings show multidimensional dependence on torsion angles, the most prominent of which is the effect of the sugar pucker. J-couplings were calculated in 16 distinct nucleic acid conformations, two principal double-helical DNAs, B- and A-, the main RNA form, A-RNA, as well as in 13 other RNA conformations. High-level quantum mechanics calculations used a baseless dinucleoside phosphate as a molecular model, and the effect of solvent was included. The predicted J-couplings correlate reliably with available experimental data from the literature.

Molecular dynamics simulations of sarcin-ricin rRNA motif.

Explicit solvent molecular dynamics (MD) simulations were carried out for sarcin-ricin domain (SRD) motifs from 23S (Escherichia coli) and 28S (rat) rRNAs. The SRD motif consists of GAGA tetraloop, G-bulged cross-strand A-stack, flexible region and duplex part. Detailed analysis of the overall dynamics, base pairing, hydration, cation binding and other SRD features is presented. The SRD is surprisingly static in multiple 25 ns long simulations and lacks any non-local motions, with root mean square deviation (r.m.s.d.) values between averaged MD and high-resolution X-ray structures of 1-1.4 A. Modest dynamics is observed in the tetraloop, namely, rotation of adenine in its apex and subtle reversible shift of the tetraloop with respect to the adjacent base pair. The deformed flexible region in low-resolution rat X-ray structure is repaired by simulations. The simulations reveal few backbone flips, which do not affect positions of bases and do not indicate a force field imbalance. Non-Watson-Crick base pairs are rigid and mediated by long-residency water molecules while there are several modest cation-binding sites around SRD. In summary, SRD is an unusually stiff rRNA building block. Its intrinsic structural and dynamical signatures seen in simulations are strikingly distinct from other rRNA motifs such as Loop E and Kink-turns.

Structural dynamics of precursor and product of the RNA enzyme from the hepatitis delta virus as revealed by molecular dynamics simulations.

The hepatitis delta virus (HDV) ribozyme is a self-cleaving RNA enzyme involved in the replication of a human pathogen, the hepatitis delta virus. Recent crystal structures of the precursor and product of self-cleavage, together with detailed kinetic analyses, have led to hypotheses on the catalytic strategies employed by the HDV ribozyme. We report molecular dynamics (MD) simulations (approximately 120 ns total simulation time) to test the plausibility that specific conformational rearrangements are involved in catalysis. Site-specific self-cleavage requires cytidine in position 75 (C75). A precursor simulation with unprotonated C75 reveals a rather weak dynamic binding of C75 in the catalytic pocket with spontaneous, transient formation of a H-bond between U-1(O2') and C75(N3). This H-bond would be required for C75 to act as the general base. Upon protonation in the precursor, C75H+ has a tendency to move towards its product location and establish a firm H-bonding network within the catalytic pocket. However, a C75H+(N3)-G1(O5') H-bond, which would be expected if C75 acted as a general acid catalyst, is not observed on the present simulation timescale. The adjacent loop L3 is relatively dynamic and may serve as a flexible structural element, possibly gated by the closing U20.G25 base-pair, to facilitate a conformational switch induced by a protonated C75H+. L3 also controls the electrostatic environment of the catalytic core, which in turn may modulate C75 base strength and metal ion binding. We find that a distant RNA tertiary interaction involving a protonated cytidine (C41) becomes unstable when left unprotonated, leading to disruptive conformational rearrangements adjacent to the catalytic core. A Na ion temporarily compensates for the loss of the protonated hydrogen bond, which is strikingly consistent with the experimentally observed synergy between low pH and high Na+ concentrations in mediating residual self-cleavage of the HDV ribozyme in the absence of divalents.

Principles of RNA base pairing: structures and energies of the trans Watson-Crick/sugar edge base pairs.

Due to the presence of the 2'-OH hydroxyl group of ribose, RNA molecules utilize an astonishing variability of base pairing patterns to build up their structures and perform the biological functions. Many of the key RNA base pairing families have no counterparts in DNA. In this study, the trans Watson-Crick/sugar edge (trans WC/SE) RNA base pair family has been characterized using quantum chemical and molecular mechanics calculations. Gas-phase optimized geometries from density functional theory (DFT) calculations and RIMP2 interaction energies are reported for the 10 crystallographically identified trans WC/SE base pairing patterns. Further, stable structures are predicted for all of the remaining six possible members of this family not seen in RNAs so far. Among these novel six base pairs, the computations substantially refine two structures suggested earlier based on simple isosteric considerations. For two additional trans WC/SE base pairs predicted in this study, no arrangement was suggested before. Thus, our study brings a complete set of trans WC/SE base pairing patterns. The present results are also contrasted with calculations reported recently for the cis WC/SE base pair family. The computed base pair sizes are in sound correlation with the X-ray data for all WC/SE pairing patterns including both their cis and trans isomers. This confirms that the isostericity of RNA base pairs, which is one of the key factors determining the RNA sequence conservation patterns, originates in the properties of the isolated base pairs. In contrast to the cis structures, however, the isosteric subgroups of the trans WC/SE family differ not only in their H-bonding patterns and steric dimensions but also in the intrinsic strength of the intermolecular interactions. The distribution of the total interaction energy over the sugar-base and base-base contributions is controlled by the cis-trans isomerism.

Non-Watson-Crick base pairing in RNA. quantum chemical analysis of the cis Watson-Crick/sugar edge base pair family.

Large RNA molecules exhibit an astonishing variability of base-pairing patterns, while many of the RNA base-pairing families have no counterparts in DNA. The cis Watson-Crick/sugar edge (cis WC/SE) RNA base pairing is investigated by ab initio quantum chemical calculations. A detailed structural and energetic characterization of all 13 crystallographically detected members of this family is provided by means of B3LYP/6-31G and RIMP2/aug-cc-pVDZ calculations. Further, a prediction is made for the remaining 3 cis WC/SE base pairs which are yet to be seen in the experiments. The interaction energy calculations point at the key role of the 2'-OH group in stabilizing the sugar-base contact and predict all 16 cis WC/SE base-pairing patterns to be nearly isoenergetic. The perfect correlation of the main geometrical parameters in the gas-phase optimized and X-ray structures shows that the principle of isosteric substitutions in RNA is rooted from the intrinsic structural similarity of the isolated base pairs. The present quantum chemical calculations for the first time analyze base pairs involving the ribose 2'-OH group and unambiguously correlate the structural information known from experiments with the energetics of interactions. The calculations further show that the relative importance and absolute value of the dispersion energy in the cis WC/SE base pairs are enhanced compared to the standard base pairs. This may by an important factor contributing to the strength of such interactions when RNA folds in its polar environment. The calculations further demonstrate that the Cornell et al. force field commonly used in molecular modeling and simulations provides satisfactory performance for this type of RNA interactions.

Theoretical study of the guanine --> 6-thioguanine substitution in duplexes, triplexes, and tetraplexes.

Molecular dynamics and thermodynamic integration calculations have been carried out on a set of G-rich single-strand, duplex, triplex, and quadruplex DNAs to study the structural and stability changes connected with the guanine --> 6-thioguanine (G --> S) mutation. The presence of 6-thioguanine leads to a shift of the geometry from the B/A intermediate to the pure B-form in duplex DNA. The G --> S mutation does not largely affect the structure of the antiparallel triplex when it is located at the reverse-Hoogsteen position, but leads to a non-negligible local distortion in the structure when it is located at the Watson-Crick position. The G --> S mutation leads to destabilization of all studied structures: the lowest effect has been observed for the G --> S mutation in the reverse-Hoogsteen strand of the triplex, a medium effect has been observed in the Watson-Crick strand of the triplex and duplex, and the highest influence of the G -->S mutation has been found for the quadruplex structures.

Long-residency hydration, cation binding, and dynamics of loop E/helix IV rRNA-L25 protein complex.

Molecular dynamics simulations of RNA-protein complex between Escherichia coli loop E/helix IV (LE/HeIV) rRNA and L25 protein reveal a qualitative agreement between the experimental and simulated structures. The major groove of LE is a prominent rRNA cation-binding site. Divalent cations rigidify the LE major groove geometry whereas in the absence of divalent cations LE extensively interacts with monovalent cations via inner-shell binding. The HeIV region shows bistability of its major groove explaining the observed differences between x-ray and NMR structures. In agreement with the experiments, the simulations suggest that helix-alpha1 of L25 is the least stable part of the protein. Inclusion of Mg2+ cations into the simulations causes perturbation of basepairing at the LE/HeIV junction, which does not, however, affect the protein binding. The rRNA-protein complex is mediated by a number of highly specific hydration sites with long-residing water molecules and two of them are bound throughout the entire 24-ns simulation. Long-residing water molecules are seen also outside the RNA-protein contact areas with water-binding times substantially enhanced compared to simulations of free RNA. Long-residency hydration sites thus represent important elements of the three-dimensional structure of rRNA.

Ribosomal RNA kink-turn motif--a flexible molecular hinge.

Ribosomal RNA K-turn motifs are asymmetric internal loops characterized by a sharp bend in the phosphodiester backbone resulting in "V" shaped structures, recurrently observed in ribosomes and showing a high degree of sequence conservation. We have carried out extended explicit solvent molecular dynamics simulations of selected K-turns, in order to investigate their intrinsic structural and dynamical properties. The simulations reveal an unprecedented dynamical flexibility of the K-turns around their X-ray geometries. The K-turns sample, on the nanosecond timescale, different conformational substates. The overall behavior of the simulations suggests that the sampled geometries are essentially isoenergetic and separated by minimal energy barriers. The nanosecond dynamics of isolated K-turns can be qualitatively considered as motion of two rigid helix stems controlled by a very flexible internal loop which then leads to substantial hinge-like motions between the two stems. This internal dynamics of K-turns is strikingly different for example from the bacterial 5S rRNA Loop E motif or BWYV frameshifting pseudoknot which appear to be rigid in the same type of simulations. Bistability and flexibility of K-turns was also suggested by several recent biochemical studies. Although the results of MD simulations should be considered as a qualitative picture of the K-turn dynamics due to force field and sampling limitations, the main advantage of the MD technique is its ability to investigate the region close to K-turn ribosomal-like geometries. This part of the conformational space is not well characterized by the solution experiments due to large-scale conformational changes seen in the experiments. We suggest that K-turns are well suited to act as flexible structural elements of ribosomal RNA. They can for example be involved in mediation of large-scale motions or they can allow a smooth assembling of the other parts of the ribosome.

Molecular dynamics simulations of Guanine quadruplex loops: advances and force field limitations.

A computational analysis of d(GGGGTTTTGGGG)(2) guanine quadruplexes containing either lateral or diagonal four-thymidine loops was carried out using molecular dynamics (MD) simulations in explicit solvent, locally enhanced sampling (LES) simulations, systematic conformational search, and free energy molecular-mechanics, Poisson Boltzmann, surface area (MM-PBSA) calculations with explicit inclusion of structural monovalent cations. The study provides, within the approximations of the applied all-atom additive force field, a qualitatively complete analysis of the available loop conformational space. The results are independent of the starting structures. Major conformational transitions not seen in conventional MD simulations are observed when LES is applied. The favored LES structures consistently provide lower free energies (as estimated by molecular-mechanics, Poisson Boltzmann, surface area) than other structures. Unfortunately, the predicted optimal structure for the diagonal loop arrangement differs substantially from the atomic resolution experiments. This result is attributed to force field deficiencies, such as the potential misbalance between solute-cation and solvent-cation terms. The MD simulations are unable to maintain the stable coordination of the monovalent cations inside the diagonal loops as reported in a recent x-ray study. The optimal diagonal and lateral loop arrangements appear to be close in energy although a proper inclusion of the loop monovalent cations could stabilize the diagonal architecture.

Molecular dynamics simulations of RNA kissing-loop motifs reveal structural dynamics and formation of cation-binding pockets.

Explicit solvent molecular dynamics (MD) simulations were carried out for three RNA kissing-loop complexes. The theoretical structure of two base pairs (2 bp) complex of H3 stem-loop of Moloney murine leukemia virus agrees with the NMR structure with modest violations of few NMR restraints comparable to violations present in the NMR structure. In contrast to the NMR structure, however, MD shows relaxed intermolecular G-C base pairs. The core region of the kissing complex forms a cation-binding pocket with highly negative electrostatic potential. The pocket shows nanosecond-scale breathing motions coupled with oscillations of the whole molecule. Additional simulations were carried out for 6 bp kissing complexes of the DIS HIV-1 subtypes A and B. The simulated structures agree well with the X-ray data. The subtype B forms a novel four-base stack of bulged-out adenines. Both 6 bp kissing complexes have extended cation-binding pockets in their central parts. While the pocket of subtype A interacts with two hexacoordinated Mg2+ ions and one sodium ion, pocket of subtype B is filled with a string of three delocalized Na+ ions with residency times of individual cations 1-2 ns. The 6 bp complexes show breathing motions of the cation-binding pockets and loop major grooves.

Formation pathways of a guanine-quadruplex DNA revealed by molecular dynamics and thermodynamic analysis of the substates.

The formation of a cation-stabilized guanine quadruplex (G-DNA) stem is an exceptionally slow process involving complex kinetics that has not yet been characterized at atomic resolution. Here, we investigate the formation of a parallel stranded G-DNA stem consisting of four strands of d(GGGG) using molecular dynamics simulations with explicit inclusion of counterions and solvent. Due to the limitations imposed by the nanosecond timescale of the simulations, rather than watching for the spontaneous formation of G-DNA, our approach probes the stability of possible supramolecular intermediates (including two-, three-, and four-stranded assemblies with out-of-register base pairing between guanines) on the formation pathway. The simulations suggest that "cross-like" two-stranded assemblies may serve as nucleation centers in the initial formation of parallel stranded G-DNA quadruplexes, proceeding through a series of rearrangements involving trapping of cations, association of additional strands, and progressive slippage of strands toward the full stem. To supplement the analysis, approximate free energies of the models are obtained with explicit consideration of the integral cations. The approach applied here serves as a prototype for qualitatively investigating other G-DNA molecules using molecular dynamics simulation and free-energy analysis.

Unique tertiary and neighbor interactions determine conservation patterns of Cis Watson-Crick A/G base-pairs.

X-ray, phylogenetic and quantum chemical analysis of molecular interactions and conservation patterns of cis Watson-Crick (W.C.) A/G base-pairs in 16S rRNA, 23S rRNA and other molecules was carried out. In these base-pairs, the A and G nucleotides interact with their W.C. edges with glycosidic bonds oriented cis relative to each other. The base-pair is stabilised by two hydrogen bonds, the C1'-C1' distance is enlarged and the G(N2) amino group is left unpaired. Quantum chemical calculations show that, in the absence of other interactions, the unpaired amino group is substantially non-planar due to its partial sp(3) pyramidalization, while the whole base-pair is internally propeller twisted and very flexible. The unique molecular properties of the cis W.C. A/G base-pairs make them distinct from other base-pairs. They occur mostly at the ends of canonical helices, where they serve as interfaces between the helix and other motifs. The cis W.C. A/G base-pairs play crucial roles in natural RNA structures with salient sequence conservation patterns. The key contribution to conservation is provided by the unpaired G(N2) amino group that is involved in a wide range of tertiary and neighbor contacts in the crystal structures. Many of them are oriented out of the plane of the guanine base and utilize the partial sp(3) pyramidalization of the G(N2). There is a lack of A/G to G/A covariation, which, except for the G(N2) position, would be entirely isosteric. On the contrary, there is a rather frequent occurrence of G/A to G/U covariation, as the G/U wobble base-pair has an unpaired amino group in the same position as the cis W.C. G/A base-pair. The cis W.C. A/G base-pairs are not conserved when there is no tertiary or neighbor interaction. Obtaining the proper picture of the interactions and phylogenetic patterns of the cis W.C. A/G base-pairs requires a detailed analysis of the relation between the molecular structures and the energetics of interactions at a level of single H-bonds and contacts.

Non-Watson-Crick basepairing and hydration in RNA motifs: molecular dynamics of 5S rRNA loop E.

Explicit solvent and counterion molecular dynamics simulations have been carried out for a total of >80 ns on the bacterial and spinach chloroplast 5S rRNA Loop E motifs. The Loop E sequences form unique duplex architectures composed of seven consecutive non-Watson-Crick basepairs. The starting structure of spinach chloroplast Loop E was modeled using isostericity principles, and the simulations refined the geometries of the three non-Watson-Crick basepairs that differ from the consensus bacterial sequence. The deep groove of Loop E motifs provides unique sites for cation binding. Binding of Mg(2+) rigidifies Loop E and stabilizes its major groove at an intermediate width. In the absence of Mg(2+), the Loop E motifs show an unprecedented degree of inner-shell binding of monovalent cations that, in contrast to Mg(2+), penetrate into the most negative regions inside the deep groove. The spinach chloroplast Loop E shows a marked tendency to compress its deep groove compared with the bacterial consensus. Structures with a narrow deep groove essentially collapse around a string of Na(+) cations with long coordination times. The Loop E non-Watson-Crick basepairing is complemented by highly specific hydration sites ranging from water bridges to hydration pockets hosting 2 to 3 long-residing waters. The ordered hydration is intimately connected with RNA local conformational variations.

Molecular dynamics simulations and thermodynamics analysis of DNA-drug complexes. Minor groove binding between 4',6-diamidino-2-phenylindole and DNA duplexes in solution.

An extended set of nanosecond-scale molecular dynamics simulations of DNA duplex sequences in explicit solvent interacting with the minor groove binding drug 4',6-diamidino-2-phenylindole (DAPI) are investigated for four different and sequence specific binding modes. Force fields for DAPI have been parametrized to properly reflect its internal nonplanarity. Sequences investigated include the binding modes observed experimentally, that is, AATT in d(CGCGAATTCGCG)(2) and ATTG in d(GGCCAATTGG)(2) and alternative shifted binding modes ATTC and AATT, respectively. In each case, stable MD simulations are obtained, well reproducing specific hydration patterns seen in the experiments. In contrast to the 2.4 A d(CGCGAATTCGCG)(2) crystal structure, the DAPI is nonplanar, consistent with its gas-phase geometry and the higher resolution crystal structure. The simulations also suggest that the DAPI molecule is able to adopt different conformational substates accompanied by specific hydration patterns that include long-residing waters. The MM_PBSA technology for estimating relative free energies was utilized. The most consistent free energy results were obtained with an approach that uses a single trajectory of the DNA-DAPI complex to estimate all free energy terms. It is demonstrated that explicit inclusion of a subset of bound water molecules shifts the calculated relative binding free energies in favor of both crystallographically observed binding modes, underlining the importance of structured hydration.