Structural Waters in the Minor and Major Grooves of DNAâA Major

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Structural Waters in the Minor and Major Grooves of DNA – A Major Factor Governing Structural Adjustments of A-T Mini-helix Tetiana A Zubatiuk, Oleg V Shishkin, Leonid Gorb, Dmytro Mykolayovich Hovorun, and Jerzy Leszczynski J. Phys. Chem. B, Just Accepted Manuscript • DOI: 10.1021/jp5075225 • Publication Date (Web): 14 Dec 2014 Downloaded from http://pubs.acs.org on December 25, 2014

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The Journal of Physical Chemistry

Structural Waters in the Minor and Major Grooves of DNA – a Major Factor Governing Structural Adjustments of A-T Mini-helix Tetiana Zubatiuk1, Oleg Shishkin1,2, Leonid Gorb3, Dmytro Hovorun4, and Jerzy Leszczynski5,* 1

Division of Functional Materials Chemistry, SSI “Institute for Single Crystals” National

Academy of Science of Ukraine, Kharkiv, 61001, Ukraine 2

Department of Inorganic Chemistry, V. N. Karazin National University, Kharkiv, 61122,

Ukraine 3

Laboratory of Computational Structural Biology, Department of Molecular Biophysics,

Institute of Molecular Biology and Genetics, National Academy of Sciences of Ukraine, Kyiv, 03143, Ukraine 4

Department of Molecular Biophysics, Institute of Molecular Biology and Genetics, National

Academy of Sciences of Ukraine, Kyiv 03143, Ukraine 5

Interdisciplinary Center for Nanotoxicity, Department of Chemistry and Biochemistry, Jackson

State University, Jackson, MS 39217, USA

ACS Paragon Plus Environment


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ABSTRACT

The role of micro-hydration in structural adjustments of AT-tract in B-DNA was studied at B97D/def2-SV(P) level. The (dA:dT)5 complexes with 10 water molecules in minor and 15 water molecules in major grooves were studied. The obtained network of hydrogen bonds revealed dependence between groove width and the types of water patterns. In minor groove the following patterns were observed: interstrand one-water bridges similar to that of Dickerson “water spine” and interstrand two-water bridges. Network of structural waters in major groove is more diverse than in minor, which agrees with crystallographic data. As the major groove is wider, it is enriched by water molecules formed two- and three-water bridges. Results suggest the nucleobase-water interactions in both grooves prevent AT-tract twisting and its “collapse” along the minor groove. Whereby, helix structure with narrow minor and wide major grooves is formed. The structural waters affect the polynucleotide conformation so that it becomes similar to poly(dA)·poly(dT) in fibers and acquires features of the A-tracts in DNA in solution. We suggest that formation of specific water patterns in both grooves is the factor responsible for stabilization of A-tracts with narrowed minor groove, leading in turn to their strong intrinsic bending in DNA.

KEYWORDS. Nucleic acids, AT-tract, micro-hydration, DFT, water spine.

INTRODUCTION It has been shown many times (for reviews see

1–6

) that water molecules which associate

strongly with DNA have much reduced mobility and occupy well defined hydration sites. They are localized enough to be observed by X-ray or neutron diffraction 7. The most ordered water


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molecules are also visible by NMR techniques 8–10. The mobility of water molecules depends on DNA form (A, B, Z, etc.), ionic strength and locations in major or minor grooves

11

. As a

consequence, DNA conformations are aptly responsive to external effects and the local structure of DNA can vary over a wide range of conformations. This results in the structural variability of DNA, such as the sequence-specific bending and sequence-specific curvature, which play a role in a number of basic processes, including protein binding and gene regulation. Therefore, the DNA interaction with structural waters is critical factor in stabilization of different nucleic acid conformation. This raises the important question of the interconnectivity of the sequencedependent structures and energetics of DNA duplex and the details of its hydration. There are overwhelming experimental evidences

12–22

that the A-tracts of B-DNA (contiguous

runs of several adenines in one strand) of lengths of four or more base pairs adopt a novel cooperative state whose structures, dynamics, and thermodynamic properties differ fundamentally from those of mixed B-DNA sequence. Namely, compared to the "mixed" GCrich sequences, A-tracts in solution are characterized by a compressed minor groove specific internal local geometrical parameters

12,13

and specific bending

18–22

. To the best of our

knowledge there is no consensus in the literature regarding explanation of this phenomenon at the molecular level. There is a suggestion that the compression of the minor groove and other changes of the structure in the A-tract (particularly, in central part of Dickerson dodecamer) are caused to some extent by the "spine of hydration" "A-tract" model 2, the "non-A-tract" model

28–30

23–27

. However, the existing models, viz. the

and various "wedge" models, both static and

flexible 31–34 have one feature in common, which appears to be consistent with the whole array of experimental data. For instance, the "spine of hydration" hypothesis, considers a regular positioning of the structural waters in the minor groove as the main factor stabilizing the B-like



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conformation of A-tracts in solution

23–27

DNA dodecamer d(CGCGAATTCGCG)2

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. This model, based on the crystal structure of the B35

, successfully predicted the larger number of bound

waters in A-tracts, compared to the alternating A-T sequences 36. In addition, the driving forces leading to such a regular water spine in the narrow minor groove compared to water strings in the relatively wide major groove of the "standard" B-form still remains unclear

37

. Also, the

primary determinant of the minor groove narrowing and changing of A-tracts geometry which we have already mentioned above is not yet understood. Furthermore, "hydration spine" concept based exclusively on the formation of the hydration spine in the minor groove, fails to explain why the DNA curvature depends upon the nature of the exocyclic groups in the purine 6- and 7-positions in the major groove

38,39

. The hydration of

the minor groove alone cannot account for the noticeable difference in the electrophoretic mobility of the DNAs containing (dA:dT)4 and (AATT)2 tracts

20,21

. In addition, the amount of

water released upon binding of netropsin to A-tract is substantially larger than would be expected if the frozen-in water melted only from the minor groove

40

. The optical, spectroscopic and

calorimetric studies of the B' → B "premelting" transition in the A-tract fragments also point to melting and release of "bound" water into the bulk solvent 41. Therefore, a thorough examination of DNA hydration, depending on DNA conformation, including the major groove consideration, is necessary. One more reason to pay attention to the major groove is that the third strand of the DNA-triplex is located there. Finally, many proteins bind in the major groove, and the "structural" waters appear to be an important component governing the protein-DNA interface. An extensive H-bonding of water molecules with the special DNA regions and each other (in minor and major grooves) is expected to contribute to a length-dependent cooperative formation of the helix`s special structure. As an example, we would like to mention one of biological


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implications showing that A-tracts might function in eukaryotes to influence the organization of nucleosomes at many genomic regions

42

. Due to their properties A-tracts may resist sharp

bending and consequently, disfavor nucleosome incorporation. In other words, there are many evidences that A-tracts within DNA intrinsically resist the structural deformations required for nucleosome formation. Consistent with this view, a model for the sequence-dependent free energy of DNA wrapping in nucleosomes suggests that the curvature-dependent DNA hydration changes are coupled to sharp DNA bending and this plays a significant role in the energetics of nucleosome formation

42

. We believe that this is the true evidence of existing of a certain

connection between the sequence-dependent structures of DNA and the features of its hydration layers inside the grooves. Moreover, the very recent, high-resolution DNA crystal structures research

43

proved that water molecules really form a stable arrangement inside the minor

groove. Interestingly, even ligands adopt an appropriate orientation around these water molecules. During the binding the existed spine of hydration is displaced by bound ligands, but it does not disappear. The new arrangement of water molecules is distinct from, but linked by hydrogen bonding to the spine of hydration.43 Recently we applied modern versions of DFT approximation to study structural and energetic features of mini-helices of B-DNA.44 It was shown that quantum-chemical modeling provides a valid and efficient way for examining duplex and strands specificity including the influence of sequence dependence for both structural and energetic characteristics. An accurate prediction of binding energies of (dG:dC)3 and (dA:dT)3 duplexes validates the applied models and evidences reliability of the applied computational techniques. Also, the published data provide new, very clear evidences supporting the statement that DNA is able to adopt the classical conformations as A, B, etc., due to the interactions of a DNA backbone with water bulk. This work extends this



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point of view in the following directions: We will shed a light on the issue how the specific characteristics of A-tracts depend on waters location in the grooves. To consider the issue, we specifically hydrated (such type of hydration is called a micro-hydration

45–48

) certain sites of

(dA:dT)5 mini-helix possessing B-DNA conformation. The final geometry was obtained after full optimization of the considered models, using an appropriate density functional theory and continuum representation of a water bulk. The paper is organized as follows: After describing computational details we very specifically explain the way how we construct our models. Since such characteristics of A-tracts as width of grooves, parameters of helix, intrinsic angle of bending of local DNA fragment

49

have specific

values; next sections reveal how the values of those parameters depend on the type of microhydration. COMPUTATIONAL METHODS The structural results discussed in this paper are based on fully optimized complexes of double stranded B-DNA-like helixes consist of 5 A-T base-pairs referred to as (dA:dT)5 with structural waters in grooves. The initial structure of the (dA:dT)5 was constructed from the canonical BDNA conformation using the 3DNA

50

software package. Micro-hydrated (dA:dT)5 complexes

were built with 10 structural water molecules in minor groove and 15 water molecules in major groove. By following the conclusions from our previous paper

44

all present calculations were

carried out using COSMO continuum solvation model with dielectric constant equal to 80 51 and in presence of sodium counterions to neutralize the negative charges of phosphate groups. The cations were placed equidistantly (approximately 2.4 Å) from the two phosphate oxygen atoms along the backbone and were allowed to move during geometry optimization. The starting locations of structural waters along the grooves are discussed below. For the sake of comparison,


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also, the negatively charged (uncompensated) micro-hydrated (dA:dT)5 mini-helices were immersed into continuum type of dielectric medium. However, this is not simply a hydrated anionic form of considered mini-helixes. In the reality, these models effectively include average influence of sodium counter-ions (see the explanation in

44

). We also use similar geometry

optimization strategy for the uncompensated mini-helices with different sequences, as follows: [(dА:dT)2(dТ:dA)(dА:dT)2] denoted as ААТАА, [(dА:dT)2(dC:dG)(dА:dT)2] denoted as ААСАА,

[(dА:dT)(dТ:dA)

(dА:dT)(dТ:dА)(dA:dT)]

denoted

as

АТАТА,

and

(dА:dT)(dG:dC)(dA:dT)(dG:dC)(dA:dT) denoted as АGAGA. All optimized geometries are published as the ESI (Tables 4S-17S). Because the size of considered systems is significantly larger than recently studied (AT)3 and (GC)3 mini-helices, the previously used

44

computational strategy has been changed as follows.

In the current study we used pure GGA DFT functional B97-D3,

52,53

which is less time

consuming comparing to hybrid meta-functional M06-2X. Also, we used the def2-SV(P) 54 basis set which on the one hand is of 6-31G* quality but on another hand provides efficient integral evaluation with multipole accelerated RI-J technique. The B97-D density functional includes empirical correction to accurately account for dispersion contribution and was benchmarked to be one of the most powerful density functionals for calculations of intra- and intermolecular interactions.

53,55,56

We believe that the application of def2-SV(P) basis set is enough to secure

the quality of obtained structural parameters. The analysis of those structural parameters is the central subject of the study. Also, due to the large size of the system we did not calculated Hessian for the obtained structures to ascertain location of true local minima on the potential surface. The calculations were performed using Turbomole 6.4 57 software.



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Figure 1. Definition of the helix bending angle β between two mean planes of adjacent base pairs. These planes are defined by four atoms: C1′, N9 of adenine and C1′, N1 of thymine (showing as balls). Numbers corresponded to the site numbering for A-T nucleobase.

The geometrical analysis of duplex structure was performed in terms of the local base-pair and helical parameters (propeller, tilt and twist), which were calculated with 3DNA 50 software. The description of the minor groove width was made in terms of the distance between the nearest phosphorus atoms of the antiparallel chains (denoted as P(Ai+3)...P(Ti)), the distance between the closest sugar ring oxygens (denoted as O4'(Ai+1)...O4'(Ti)) and the distance between N3 of adenine and O2 of adjusted thymine of the opposite strands (denoted as N3(Ai+1)...O2(Ti)). The major groove width was calculated as the distance between the closest sugar ring carbons C2'


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(denoted as C2'(Ai)...C2'(Ti+1)). For description of internal deformation of the (dA:dT)5 minihelix we introduced intrinsic angle of helix bending β (Fig. 1) between mean planes of two adjacent base pairs. The planes are defined by equivalent C1′ and RN9/YN1 atom pairs. Pyrimidine is labelled Y and purine is represented by R. RESULTS AND DISCUSSION Construction of water patterns for minor and major grooves Before discussing the results of our study we would like to classify our models and describe their geometrical details. B-DNA conformation of (dA:dT)5 built by 3-DNA program is referred as ideal model (IDL). Optimized IDL structure with explicit solvent modeling by COSMO method is referred to as no-water model (noW). Let us highlight again that the noW model does not have structural waters neither in minor nor in major grooves of (dA:dT)5 mini-helix, however it is immersed into continuum type of dielectric. Micro-hydrated models of (dA:dT)5 are constructed based on the IDL model and the positions of explicit water molecules in minor groove obtained from the statistical analysis of the crystals of poly(dA)poly(dT) 12,13 and Dickerson's dodecamer 35. As follows from these experimental data the prominent hydration sites in case of the minor groove are the purine N3 and pyrimidine O2 atoms. The distances between those atoms on the edges of the bases are suitable for the formation of numerous intra- and inter-strand water bridges, containing one, two or three water molecules within the same pair or within two adjacent pairs in grooves. Two patterns of water organization in the crystal structures of AT-containing DNA were found (Figure 2). In both patterns (labeled as p1 and p2) water molecules are directed "across" the groove:



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p1: the inter-strand one-water bridge between the N3 atom of adenine and O2 atom of thymine of the adjacent base-pairs. Such type of hydration forms the first layer of the well-known “water spine” in Dickerson`s dodecamer 35,58,59; p2: the inter-strand two-water bridge between the N3 atom of adenine and O2 atom of thymine of the adjacent base-pairs. Such type of hydration is described in 29 and has also formed the first layer of water molecules.

Figure 2. (a) The minor groove scheme of the AT-containing DNA fragment. The sequence chain is numbered in the 5'-3' direction; (b) two adjacent A-T pairs; (c) the known structures of water patterns p1, p2 are shown here for the adjacent base pairs which are connected on the scheme (a) and (b) with solid line through the atoms N3 and O2. Three hydrophilic centers (N3 of adenine, O2 of thymine and O4 sugar oxygens) are shown as open circles with numbers corresponded to the site numbering for A-T nucleobase. The groove profiles with water molecules on (c) represent cross section through the atoms O4'(i+3)-N3(i+2)-O2(i+1')-O4'(i') taken from scheme (a).

We use these p1 and p2 patterns to build the micro-hydrated models W1 and W2 of (dA:dT)5 with structural waters in the minor groove (Fig. 3). Repeating the p1 pattern results in five water


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molecules within the minor groove. In order to make this structure stoichiometrically consistent with the p2 model we have added five additional water molecules connecting two adjacent water molecules of the first layer. Obtained in this way W1 model with 10 structural waters within the minor groove displayed at Fig. 3a. Analogously, the W2 model has continuously repeating twowater bridge units of type p2 along the minor groove. This results in formation of the water spine consisting of single layer of 10 water molecules connecting thymine O2 and adenine N3 atoms of adjacent base pairs through the two-waters bridges. The coordinates of structural water molecules are presented as the ESI (Tables 1S and 2S).

Figure 3. The initial locations of structural waters along the minor groove in W1 and W2 models of complexes of (dA:dT)5 with 10 water molecules. The DNA bases are shown as adenine N3 and thymine O2 hydration sites. Water molecules are colour coded according to their location in different layers: from first to second as grey and white. (a) Model W1 based on the p1 water pattern, two water layers; (b) model W2 based on the p2 water pattern, one water layer.

The following patterns of water structure have been observed in the major groove according to experimental and computational studies 58–61 (Fig. 4): P1: inter-strand two water bridge across the major groove connecting thymine O4(Ti+2) and adenine H(N6)(Ai+1) from adjacent adenines. P2: the intra-adenine two water bridge between N7 and H(N6) atoms of adenine.



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P3: the intra-strand two water bridge between two N7 and H(N6) atoms of adjacent adenines. P4: inter-strand one-water bridge across the major groove connecting thymine O4(Ti+2) and adenine H(N6)(Ai+1) from adjacent adenines. The P4 hydration pattern was reported for the d(CGCAAATTTGCG)2 dodecamer 60. P5: The one-water bridge P5 is a feature intrinsic to an individual adenine; it was predicted in the course of calculations of hydration shells of the isolated adenines and AT-pairs

61

. In DNA

this one water molecule could forms three one-water bridges then it is denoted as a "water trident", i.e. a water molecule hydrogen bonded to three hydrophilic atoms of DNA simultaneously (N7 and H(N6) of the 3'-end adenine, and N7 of the 5'-end adenine). The "water trident" was observed in at least two B-DNA dodecamers: in d(CGCGAATTCGCG)2

58

and in

d(CGCAAATTTGCG)2 60.

Figure 4. (a) The major groove scheme of the AT-containing DNA fragment. The sequence chain is numbered in the 5'-3' direction; (b) the structures of water patterns P1-P5 are shown here for three adjacent base pairs which are connected in the scheme (a) with solid line through the atoms O4, N6 and N7. Three hydrophilic centers (O4 of thymine, N6 and N7 of adenine) are shown as open circles with numbers corresponded to the site numbering for A-T nucleobase. The groove profiles with water molecules in (b) represent cross section through the atoms O4(i+2)N6(i+1)-N7(i+1)-N7'(i) taken from scheme (a).


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However, the structure of water patterns in major groove is less defined than the one located in minor groove. The major groove has both donor and acceptor of protons, localized asymmetrically with respect to the dyad axis of the AT-pair (Fig. 4). Consequently, the pattern of the hydrophilic centers is essentially sequence-dependent and e.g. for the homopolymeric sequence as A-tract, is quite regular. Therefore, the interaction of a water molecules with the base`s hydrophilic centers appears to be much more variable than interactions with the backbone. The observed water bridges are directed both along and across the major groove

11

making the

structure of water spine less ordered than in the minor groove. That is why the statistical data on the geometry of water patterns in major groove cannot be normalize. To design initial structure for micro-hydration in major groove, we use the experimentally defined 11 positions of oxygen atoms in the first hydration layer of B-DNA major groove for A-T base pair. It appears that there are 15 water molecules located around three hydration sites in major groove, forming first hydration layer in (dA:dT)5. The obtained in this way structure of major groove micro-hydration is drown in Fig. 5. As suggested

11

, the water molecules are

directed both along and across the major groove. They do not form a spine of hydration, they do have the resemblance to P1 – P5 patterns but cannot be identified as those patterns confidently. Nevertheless, certainly such a structure might be used as initial guess for the model of the microhydration in major groove. The major groove of the models W1 and W2 has been hydrated in the described way. Thus, the (dA:dT)5 mini-helices having structural waters in both minor and major grooves are referred to as WW1 and WW2 models, correspondingly. The coordinates of structural waters in major groove are presented as the ESI (Table 3S).



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Figure 5. The initial locations of structural waters along and across the major groove in WW1 and WW2 models of complexes of (dA:dT)5 with water molecules. The DNA bases are shown as adenine N6, N7 and thymine O4 hydration sites. Locations of waters are based on the P1-P5 water patterns (see, Fig.4b) and form one water layer in major groove. As we mentioned in the Introduction there exists some internal characteristics of A-tracts within B-DNA which depend on the locations of structural water molecules inside the grooves, i.e. types of water patterns. Below we present the analysis of those features based on the results of calculations at the DFT B97-D level. Considering structural waters in minor and major groove The optimized geometries of un-hydrated and micro-hydrated in different ways (dA:dT)5 mini-helices are presented as the ESI (Tables 4S-8S). Since the full analysis of base-pair geometry, base-pair step, helical and backbone structural features of (dA:dT)5 will be published elsewhere, similarly as we organized presentation for (dA:dT)3 and (dG:dC)3 44, below we will discuss only features related to the subject of this study. We also would like to mention that since the structure of the hydration sites depends on the way of counter-ions compensation just in minor way, we analyze the data obtained by the application of the model which compensate the charge of backbone by Na+ counter-ions. The similar data related to negatively charged mini-


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helices (uncompensated) are immersed into continuum type of dielectric are placed in Tables 9S13S). It should be noted, that in all micro-hydrated structures discussed here all proton donating sites of water molecules form H-bonds to the DNA residues, or to each other. This results in highly ordered water patterns within the major, and especially within the minor groove. We recognize the fact, that these structures are not the only possible configuration of the water molecules within the grooves. In order to ascertain whether some disorder in starting geometry will affect the final water patterns in minor and major grooves we optimized several more micro-solvated helices with different initial positions of water molecule. In these structures the water molecules were rotated around their geometric centers using a random rotation matrix, thus introducing substantial perturbation to the patterns of micro-hydration. All obtained alternative structures contain partially reorganized and sometimes disrupted hydrogen bonding network. Six of eight optimized structure have higher (up to 25.6 kcal/mol) total energy, comparing to unperturbed one. However, two of them are placed 3.6-6.4 kcal/mol below the first set of structures. We found that the structural parameters of mini-helices that will be discussed below remain almost the same. Therefore, we decided to consider described above models W1, W2, WW1 and WW2 as being the most representative micro-hydration models. The coordinates of the alternative (perturbed) structures and their structural parameters are included in the ESI (Tables 22S-32S).



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Figure 6. The equilibrium complexes of (dA:dT)5 with structural waters along the minor groove in W1 and W2 models: (a) Model W1 with domination of p1 water pattern; (b) Model W2 with domination of p2 water pattern. The DNA bases are shown as adenine N3 and thymine O2 hydration sites. Water molecules are shown here as spheres color coded according to their positions in shells - from first to second as grey and white. Solid lines represent the network of hydrogen bonds. The position of Na+ ions is not shown.

Data in Table 1 show that equilibrium structures of water patterns in the minor groove of micro-hydrated (dA:dT)5 mini-helices differ insignificantly, comparing to the initial structures (Fig. 3, 5). The minor groove water spine structure that was predicted for W1 and WW1 complexes (Fig. 6) is very similar to that in Dickerson's dodecamer. The only major difference between initial and equilibrium geometry observed is the transition of p1 to p2 pattern for the terminal A-T base pair in the minor groove of W1 model.


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Table 1. The minor and major groove local water patterns in micro-hydrated optimized and initial (dA:dT)5 complexes. Minor groove Base pair step

initial

equilibrium

Minor groove initial

equilibrium

Model W1

Major groove initial

equilibrium

Model WW1

1AA/TT

p1

p2

p1

p2

2AA/TT

p1

p1

p1

p1

P4, P2, P3 P1, P2, P1-P5

3AA/TT

p1

p1

p1

p1

P1, P5

4AA/TT

p1

p1

p1

p1

P2

Model W2

Model WW2

1AA/TT

p2

p2

p2

p2

2AA/TT

p2

p2

p2

p2

P1, P2, P3 P1, P2, P3 P1-P5

3AA/TT

p2

p2

p2

p2

P1, P2, P3

4AA/TT

p2

p2

p2

p2

P2, P3

a

p1: the inter-strand one-water bridge between the N3 atom of adenine and O2 atom of thymine of the adjacent base-pairs, p2: the inter-strand two-water bridge between the N3 atom of adenine and O2 atom of thymine of the adjacent base-pairs (Fig. 2); P1: inter-strand two water bridge across the major groove connecting thymine O4(Ti+2) and adenine H(N6)(Ai+1) from adjacent adenines, P2: the intra-adenine two water bridge between N7 and H(N6) atoms of adenine, P3: the intra-strand two water bridge between two N7 and H(N6) atoms of adjacent adenines, P4: inter-strand one water bridge across the major groove connecting thymine O4(Ti+2) and adenine H(N6)(Ai+1) from adjacent adenines, P5: The one water bridge P5 is a feature intrinsic to an individual adenine (Fig. 4).



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Figure 7. The equilibrium complexes of (dA:dT)5 with structural waters along and across the major groove in WW1 and WW2 models: (a) P4, P5 hydration patterns dominate in model WW1; (b) P1-P3 hydration patterns dominate in model WW2. The DNA bases are shown as adenine N6, N7 and thymine O4 hydration sites. Water molecules are shown here as spheres (the water spine in minor groove is omitted). White spheres indicate water molecules which do not belong to any of discussed water patterns. Solid lines represent the network of hydrogen bonds. The position of Na+ ions is not shown.

In contrast to the minor groove, the optimized structure of the major groove micro-hydrated shell reveals significant changes. The optimization of the geometry resulted in appearance of well-defined P1 – P5 patterns of water structure described above (Fig. 7). Following data of Table 1 one notices that the ordered structure of water molecules is different for WW1 and WW2 models. However, WW2 model does not display the P4 pattern. We also would like to mention that appearance of the "water trident" in major groove (P5 water pattern) in the model WW1 locally destroys the symmetry of repeating pattern of waters’ positions along and across of major


Page 19 of 41

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groove. Most likely, appearance of P5 pattern is coupled with the local migration of one water molecule from P2 to P4 water pattern (see Fig. 4b). The similar results obtained for the uncompensated (without Na+ counter-ions) (dA:dT)5 complexes with structural waters immersed into continuum type of dielectric are shown in Table 18S. The calculations suggest some trend towards the predominance of p1 pattern over the p2 (model W has energy 10 kcal/mol lower than W2), but in case of presence of water molecules in major groove the energy differences became negligible (WW1 has energy only 0.2 kcal/mol lower than WW2). However, one needs to interpret these results cautiously, since a conformation sampling is needed to obtain this value accurately. Effect of a presence of structural waters in grooves on local helix parameters The data collected in Table 2 clearly evidence that the type of micro-hydration affects directly on local width of the minor groove. As expected, width of the minor groove (interphosphate distance) determined in framework of the IDL model is virtually the same as presented in text books

49

. However, absence of structural waters (model noW) compress it approximately twice

(according to P(Ai+3)...P(Ti) distance), what actually indicates that double helix is about to be collapsed. Presence of p1 (model W1) or p2 (model W2) patterns of water molecules effectively prevents (especially in case of p1 pattern) (dA:dT)5 mini-helix compression along the minor groove, keeping still the width significantly more narrow than the one in experimentally observed structure. Micro-hydration of both major and minor grooves results in the predicted dimensions that are very close to those recorded experimentally (models WW1 and WW2). Interesting peculiarity is revealed during the analysis of O4'(Ai+1)...O4'(Ti) and N3(Ai+1)...O2(Ti) distances. As one may see from the data presented in Table 2, the change of the type of the model (namely W1 to WW1 and W2 to WW2) does not result in significant change of their



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Page 20 of 41

values. This means that current study suggests that only external size of minor groove (P(Ai+3)...P(Ti)) is sensitive to the location of structural waters, either in minor groove, or in both grooves. The internal O4'(Ai+1)...O4'(Ti) distance is sensitive to the amount of the water molecules in the first micro-hydration layer (the variation is nearly 1Å). The (dA:dT)5 mini-helix with inter-strand one-water bridges between the N3 atom of adenine and O2 atom of thymine of the adjacent base-pairs (models W1 and WW1) has more narrow minor groove than the (dA:dT)5 mini-helix with inter-strand two-water bridges (models W2 and WW2). We also established that the tendency to a double helix collapse in absence of structural waters inside the grooves is not a specific property of A-tracts. This conclusions is based on the finding that the value of the interphosphate distance obtained in the framework of noW model for the mini-helices such as ААТАА, ААСАА, ТАТА, АGAGA is virtually the same as for (dA:dT)5 (see the data collected in Table 19S; and the appropriate equilibrium geometries of these un-hydrated mini-helices in Tables 14S-17S). More interesting situation is in the major groove. The P1 two water bridges occur locally in parts of A-tract with narrower minor groove (~8.6 Å in model WW1 and ~9.3 Å in WW2); when the minor groove is widening locally (first step 1AA/TT), the P1 pattern changes to the one water bridge pattern P4 in model WW1, see O4'(Ai+1)...O4'(Ti) distances in Table 2 and major groove patterns in Table 1. Wherein, the major groove width for the first step is narrowing locally, see C2'(Ai)...C2'(Ti+1) distances in Table 2. In general, changes of a major groove are much smaller and do not vary from model to model. In our opinion this is because the hydrophobic major groove "walls" are located farther away from the hydrophilic base atoms, compared to the minor groove, and thus these "walls" do not form a narrow cleft restraining the positions of the hydration waters. As we already mentioned, there are both hydrogen donor and


Page 21 of 41

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acceptor sites in the major groove. As a result, mobility of water molecules with the base hydrophilic centers appears to be more extensive. In accordance with water patterns presented in Table 1 the effect of changing of structural water configuration is observed locally. It may indicate that water molecules arranged mainly due to influence of local properties of minor groove, rather than of major groove. Table 2. Width of the minor and major groovesa measured from the 5′-end of the helix. Minor groove width, Å Base step

pair

Major groove width, Å

O4'(Ai+1)...O4'(Ti),

N3(Ai+1)...O2(Ti)

P(Ai+3)...P(Ti)

C2'(Ai)...C2'(Ti+1)

W1

WW1

W1

WW1

W1

W1

WW1

1AA/TT

9.9

9.5

3.9

3.8

13.1

13.5

2AA/TT

8.7

8.5

3.8

3.7

13.9

14.2

8.3

WW1

9.0

3AA/TT

8.7

8.6

3.5

3.4

14.1

14.4

4AA/TT

8.6

8.7

3.7

3.6

13.8

13.7

W2

WW2

W2

WW2

W2

WW2

1AA/TT

9.9

9.8

3.9

3.8

13.6

13.9

2AA/TT

9.3

9.4

3.8

3.7

14.2

14.3

W2

6.9

WW2

8.1

3AA/TT

9.7

9.3

3.6

3.7

13.7

14.1

4AA/TT

9.5

9.2

3.7

3.4

14.1

14.0

noW (ave.)

9.8

4.3

6.1

12.9

IDL

9.8

4.7

11.7 (11.8 49)

13.8

Exp. 12,13

-

3.3

9.2

-

a

O4'(Ai+1)...O4'(Ti) - the distance between the closest sugar ring oxygens O4' of the opposite strands; N3(Ai+1)...O2(Ti) – the distance between the adjacent base pairs of the opposite strands; P(Ai+3)...P(Ti) – the distance between the nearest phosphorus atoms of the opposite strands; C2'(Ai)...C2'(Ti+1) the distance between the closest carbons C2' of sugar ring of the opposite strands.



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Since the values of tilt, propeller and twist angles are usually discussed as specific characteristics of A-tracts.12–22 Table 3 collects these data to analyze their dependence on the type of hydration. Obtained data suggest quite satisfactory correspondence between calculated and experimental values. However, in the case of tilt and propeller the presence of structural water inside of the grooves does not seem to play a significant role. This conclusion is based on the comparison of averaged tilt and propeller values of models micro-hydrated in different way, with similar parameters that characterize noW model. However, the micro-hydration somewhat increases the propeller and changes locally the sign of base pair tilt angles. Table 3. The conformational parameters of micro-hydrated (dA:dT)5 in comparison with unhydrated (noW), and ideal (IDL) mini-helixes, and experimental data (Exp). Tilt, deg.

Propeller, deg.

Base pair

W1

1A-T

-

2A-T

3.83

2.73

4.8

3A-T

2.25

0.87

4A-T

-0.84

5A-T

W1

WW1

W2

WW2 W1

-35.4

-25.6

-28.6

-18.9

-

-0.0

-24.1

-17.9

-26.9

-17.6

42.3

38.6

41.4 39.9

3.1

0.75

-15.1

-13.9

-21.5

-23.2

39.3

38.5

37.1 37.8

-1.11

-4.3

0.92

-18.1

-19.8

-20.1

-21.5

41.0

37.6

38.3 37.5

-0.71

-1.19

-2.9

-4.86

-26.4

-27.2

-17.1

-19.2

36.3

37.7

38.1 40.5

ave.a

1.1

0.3

0.2

-0.8

-23.8

-20.9

-22.9

-20.1

39.7

38.1

38.7 38.9

noW

0.7

-18.1

45.2

IDL

0.0

-15.3

36.0

Exp.b

-6.0b

-20.0b

36.0 49

a

WW1

W2

WW2

Twist, deg.

-

WW1

W2 -

The average value of the parameter concerning five base pairs.

b

Negative sign has been assigned for this value by us. It is based on treating the data published in 12,13 by 3DNA program. Mathematically, it follows the CEHS definition 62.


WW2

Page 23 of 41

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According to data in the Table 3 the twist angle values seem to be more sensitive to the presence of structural water. The water spine perhaps stabilizes twist near 38°, in contrast to 45° in unhydrated helix (noW). Herewith, twist angle does not depend on the configuration of water spine. Moreover, tilt, propeller, and twist angle values of mini-helices that are presented in Table 3 remain almost the same as the ones obtained due to perturbation of spines of water molecules (Table 31S). Also, these values are within the distribution of angles that characterizes geometry of AT base pair in DNA surrounding

63

. This confirms that even without conformational

sampling the model of micro-hydration suggested in this paper provides reliable structural results. Effect of structural water on intrinsic bending of (dA:dT)5 duplex The structural data collected for the poly(dA:dT) tracts suggest that this part of B-DNA has the intrinsic resistance to adopting the substantially distorted structure

42

. However, some studies

suggest that poly(dA:dT) tracts are not more, but rather less resistant to bending and twisting than are the other simple sequences

64

. There are also other investigations that suggest their

stiffness to be within the normal range 65,66. However, these experiments are somewhat indirect; moreover, they monitor DNA flexibility in situations in which the DNA is rather less distorted than is DNA in biological systems. The data presented in the Table 4 do suggest that intrinsic binding depends on the type of micro-hydration. Following data in Table 4 and Table 20S (results for the uncompensated microhydrated (dA:dT)5 mini-helix), one concludes that the degree of helix bending is coupled mainly with the presence of structural waters in major groove. Interestingly, the presence of sodium cations influences to lesser degree the preservation of not bended conformation of mini-helix, than the presence of structural waters in grooves. The total absence of water molecules in



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grooves results in strong intrinsic curvature of helix. The presence of water spines in just the minor groove leads to a slight unbending. Furthermore, the presence of structural waters in both grooves makes the mini-helix significantly less bended, especially in case WW1 model (Fig. 8). Based on these results, we hypothesize that formation of specific additional water bridges in major groove is the main factor responsible for retaining B-like conformations of A-tracts not bended. The presented above results could be considered as a support of the view that AT tracts intrinsically resist the distortion due to a presence of ordered waters in the grooves. Certainly, this point requires an additional thermodynamic analysis but the structural evidence is quite convincing (Fig. 8). Thus, in the structural interpretation of experimental data on a DNA curvature the presence of structural water in major groove should be taken into account, in addition to the well-known Drew-Dickerson water spines located in the minor groove.

Figure 8. Schematic pictures of mini-helices discussed with the local helical axes (dotted line) superimposed. All structures are set with reference to the middle helical frame defined by 1, 2, 3 4, and 5-th base pairs and minor groove facing the viewer (black site).


Page 25 of 41

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Table 4. The intrinsic angles of helix bending β in micro-hydrated (dA:dT)5 mini-helices and the average value of this angle recalculated per one base pair. Base pair step

Helix bending β, deg. W1

WW1

W2

WW2

noW

1AA/TT

20.4

7.4

12.7

8.8

17.8

2AA/TT

6.2

5.6

6.8

7.5

12.5

3AA/TT

7.1

1.9

5.1

3.7

12.9

4AA/TT

6.7

2.9

9.7

3.3

14.9

Average bending per base pair

10.1

4.4

8.6

5.8

14.6

IDL

0.3

Similarly to our findings described in section 3.3, we established that the values of helix bending obtained in the framework of noW model for the following mini-helixes: ААТАА, ААСАА, ТАТА, and АGAGA have the same magnitudes as for (dA:dT)5 (see the data collected in Table 21S). Therefore, one may conclude that the tendency to a double helix strong bending in absence of structural waters inside the grooves is not a specific property of A-tracts. CONCLUSIONS Applying B97-D level of density functional theory to the calculation of geometrical characteristics of (dA:dT)5 B-DNA mini-helix, we found that such characteristics as major and minor grooves width, local helical parameters and internal bending might be reproduced with the same accuracy as corresponding values measured experimentally. Such accuracy is reached when ordered in water bridges (water spine in case of minor groove) structural water is placed in major and minor grooves. As a result, the structural waters affect the polynucleotide



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Page 26 of 41

conformation so that it becomes strikingly similar to the conformation of poly(dA)·poly(dT) in fibers. The obtained results suggest that minor groove width and internal bending are the most sensitive parameters to the presence of structural water. The specific water patterns in the major and minor grooves is the factor responsible for stabilization of conformations of A-tracts with narrowed minor groove and low intrinsic bending. In contrast such geometrical parameters as tilt, propeller and twist are relatively less sensitive. We concluded that obtained result have to be taken into account during the interpretation of experimental data on conformational dynamics and bending of A-tracts. However, seems that the tending to the collapse of minor groove and strong internal bending in the absence of structural water are more general DNA property that the just property of AT-tracts. ASSOCIATED CONTENT Supporting Information Available: B97-D/def2-SV(P) optimized reference geometries of un-hydrated and micro-hydrated (dA:dT)5 mini-helices (compensated and uncompensated forms) immersed into continuum type of dielectric medium; B97-D/def2-SV(P) optimized and initial reference geometries of micro-hydrated (dA:dT)5 mini-helices with perturbed spine of water molecules; B97-D/def2-SV(P) optimized reference geometries of un-hydrated and uncompensated B-DNA mini-helices (AACAA, AATAA, ATATA, AGAGA) immersed into continuum type of dielectric medium; the initial coordinates of structural waters in minor and major grooves; the minor and major groove local water patterns for uncompensated microhydrated (dA:dT)5 mini-helixes; the minor and major groove widths and the intrinsic angles of helix bending for uncompensated micro-hydrated (dA:dT)5 mini-helices and mini-helices with different sequences (AACAA, AATAA, ATATA, AGAGA); the structural parameters of micro-


Page 27 of 41

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hydrated (dA:dT)5 mini-helices with perturbed spine of water molecules. This material is available free of charge via the Internet at http://pubs.acs.org. AUTHOR INFORMATION Corresponding Author * To whom correspondence should be addressed. Tel: 60 1979 3482; Fax: 60 1979 7823; E-mail: [email protected].

Author Contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Funding Sources This work was supported by National Science Foundation [grant number NSF/CREST HRD0833178]. This research was supported in part by the Extreme Science and Engineering Discovery Environment (XSEDE) [National Science Foundation grant number OCI-1053575 and XSEDE award allocation number DMR110088]. ACKNOWLEDGEMENT Authors thank the Mississippi Center for Supercomputer Research (Oxford, MS) for the generous allotment of computer time. An access to computational facilities of joint computational cluster of SSI “Institute for Single Crystals” and Institute for Scintillation Materials of National Academy of Science of Ukraine incorporated into Ukrainian National Grid is gratefully acknowledged.



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DEDICATION This quantum-chemical study of DNA is dedicated to the memory of Dr. Oleg Shishkin, SSI “Institute for Single Crystals” National Academy of Science of Ukraine, for his outstanding contributions to computational chemistry science. He lead the way of the series of studies devoted to assisted elucidation of the key aspects of structure and specific features of nucleic acids and their fragments. He also inspired the major part of the great ideas of these studies. Oleg passed away suddenly and unexpectedly on 17th July, 2014. You will always be missed and we cherish the memories we had. REFERENCES (1)

Trifonov, E. N. Curved DNA. CRC Crit. Rev. Biochem. 1985, 19, 89–106.

(2)

Crothers, D. M.; Haran, T. E.; Nadeau, J. G.; Dna, B.; Crothers, D. M.; Nadeau, J. G. Intrinsically Bent DNA. J. Biol. Chem. 1990, 265, 7093–7096.

(3)

Hagerman, P. J. Sequence-Directed Curvature of DNA. Annu. Rev. Biochem. 1990, 59, 755–81.

(4)

Harrington, R. E.; Winicov, I. New Concepts in Protein-DNA Recognition: Sequence Directed DNA Bending and Flexibility. Prog. Nucleic Acids Res.Mol. Biol. 1994, 47, 195–270.

(5)

Travers, A. A. In DNA-Protein: Structural Interactions; Lilley, D. M. J., Ed.; IRL Press: Oxford, N.Y., Tokyo, 1995; pp. 49–75.

(6)

Olson, W. K.; Zhurkin, V. B. In Biological Structure and Dynamics; Sarma, R. H.; Sarma, M. H., Eds.; Adenine Press: N.Y., 1996; pp. 341–370.

(7)

Savage, H.; Wlodawer, A. Determination of Water Structure around Biomolecules Using X-Ray and Neutron Diffraction Methods. Methods Enzymol. 1986, 127, 162–183.

(8)

Kubinec, M. G.; Wemmer, D. E. NMR Evidence for DNA Bound Water in Solution. J. Am. Chem. Soc. 1992, 114, 8739–8740.

(9)

Liepinsh, E.; Otting, G.; Wüthrich, K. NMR Observation of Individual Molecules of Hydration Water Bound to DNA Duplexes: Direct Evidence for a Spine of Hydration Water Present in Aqueous Solution. Nucleic Acids Res. 1992, 20, 6549–6553.

(10)

Liepinsh, E.; Leupin, W.; Otting, G. Hydration of DNA in Aqueous Solution: NMR Evidence for a Kinetic Destabilization of the Minor Groove Hydration of D-(TTAA) 2 versus D-(AATT) 2 Segments. Nucleic Acids Res. 1994, 22, 2249–2254.


Page 29 of 41

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(11)

Schneider, B.; Cohen, D.; Berman, H. M. Hydration of DNA Bases: Analysis of Crystallographic Data. Biopolymers. 1992, 32, 725–50.

(12)

Arnott, S.; Chandrasekaran, R.; Hall, I. H.; Puigjaner, L. C. Heteronomous DNA. Nucleic Acids Res. 1983, 11, 4141–4155.

(13)

Alexeev, D. G.; Lipanov, A. A.; Skuratovskii IYa. Poly(dA).poly(dT) Is a B-Type Double Helix with a Distinctively Narrow Minor Groove. Nature. 1987, 325, 821–3.

(14)

Haran, T. E.; Crothers, D. M. Cooperativity in A-Tract Structure and the Bending Properties of Composite TnAn Blocks. Biochemistry. 1989, 28, 2763–2767.

(15)

Chan, S. S.; Breslauer, K. J.; Austin, R. H.; Hogan, M. E. Thermodynamics and Premelting Conformational Changes of Phased (dA)5 Tracts. Biochemistry. 1993, 32, 11776–11784.

(16)

Barbic, A.; Zimmer, D. P.; Crothers, D. M. Structural Origins of Adenine-Tract Bending. Proc. Natl. Acad. Sci. U. S. A. 2003, 100, 2369–73.

(17)

Stefl, R.; Wu, H.; Ravindranathan, S.; Sklenár, V.; Feigon, J. DNA A-Tract Bending in Three Dimensions: Solving the dA4T4 vs. dT4A4 Conundrum. Proc. Natl. Acad. Sci. U. S. A. 2004, 101, 1177–82.

(18)

Marini, J. C.; Levene, S. D.; Crothers, D. M.; Englund, P. T. Bent Helical Structure in Kinetoplast DNA. Proc. Natl. Acad. Sci. USA. 1982, 79, 7664–7668.

(19)

Diekmann, S. Sequence Specificity of Curved DNA. FEBS Lett. 1986, 195, 53–56.

(20)

Koo, H.-S.; Wu, H.-M.; Crothers, D. M. DNA Bending at Adenine-Thymine Tract. Nature. 1986, 320, 501–506.

(21)

Hagerman, P. J. Flexibility of DNA. Annu. Rev. Biophys. Biophys. Chem. 1988, 17, 265–286.

(22)

Beutel, B. A.; Gold., L. In Vitro Evolution of Intrinsically Bent DNA. J. Mol. Biol. 1992, 228, 803– 812.

(23)

Nelson, H. C.; Finch, J. T.; Luisi, B. F.; Klug, A. The Structure of an oligo(dA).oligo(dT) Tract and Its Biological Implications. Nature. 1987, 330, 221–6.

(24)

DiGabriele, A. D.; Steitz, T. A. A DNA Dodecamer Containing an Adenine Tract Crystallizes in a Unique Lattice and Exhibits a New Bend. J. Mol. Biol. 1993, 231, 1024–39.

(25)

Diekmann, S.; Mazzarelli, J. M.; McLaughlin, L. W.; Kitzing, E. von; Travers., A. A. DNA Curvature Does Not Require Bifurcated Hydrogen Bonds or Pyrimidine Methyl Groups. J. Mol. Biol. 1992, 225, 729–738.

(26)

Chuprina, V. P. Regularities in Formation of the Spine of Hydration in the DNA Minor Groove and Its Influence on the DNA Structure. FEBS Lett. 1985, 186, 98–102.

(27)

Chuprina, V. P. Anomalous Structure and Properties of poly(dA).poly(dT). Computer Simulation of the Polynucleotide Structure with the Spine of Hydration in the Minor Groove. Nucleic Acids Res. 1987, 15, 293–311.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 30 of 41

(28)

Calladine, C. R.; Drew, H. R.; McCall., M. J. The Intrinsic Curvature of DNA in Solution. J. Mol. Biol. 1988, 201, 127–137.

(29)

Goodsell, D. S.; Kaczor-Grzeskowiak, M.; Dickerson, R. E. The Crystal Structure of C-C- A-T-T-AA-T-G-G. Implications for Bending of B-DNA at T-A Steps. J. Mol. Biol. 1994, 239, 79–96.

(30)

Brukner, I., S.; Susic, M. D.; A. Savic; Pongor., S. Physiological Concentration of Mg++ Induces a Strong Macroscopic Curvature in GGGCCC-Containing DNA. J. Mol. Biol. 1994, 236, 26–32.

(31)

Bolshoy, A., P.; McNamara, R. E.; Harrington; Trifonov, E. N. Curved DNA without A-A: Experimental Estimation of All 16 DNA Wedge Angles. Proc. Natl. Acad. Sci. USA. 1991, 88, 2312–2316.

(32)

De Santis, P., A.; Palleschi, M. S.; Scipioni., A. Validity of the Nearest-Neighbor Approximation in the Evaluation of the Electrophoretic Manifestations of DNA Curvature. Biochemistry. 1990, 29, 9269–9273.

(33)

Zhurkin, V. B.; Ulyanov, N. B.; Gorin, A. A.; Jernigan, R. L. Static and Statistical Bending of DNA Evaluated by Monte Carlo Simulations. Proc. Natl. Acad. Sci. USA. 1991, 88, 7046–7050.

(34)

Olson, W. K.; Marky, N. L.; Jernigan, R. L.; Zhurkin, V. B. Influence of Fluctuations on DNA Curvature. A Comparison of Flexible and Static Wedge Models of Intrinsically Bent DNA. J. Mol. Biol. 1993, 232, 530–551.

(35)

Drew, H. R.; Dickerson, R. E. Structure of a B-DNA Dodecamer. III. Geometry of Hydration. J. Mol. Biol. 1981, 151, 535–556.

(36)

Buckin, V. A.; Kankiya, B. I.; Bulichov, N. V.; Lebedev, A. V.; Gukovsky, I. Y.; Chuprina, V. P.; Sarvazyan, A. P.; Williams., R. A. Measurement of Anomalously High Hydration of (dA)n .(dT)n Double Helices in Dilute Solution. Nature.1989, 340, 321–322.

(37)

Prive, G. G.; Heinemann, U.; Chandrasegaran, S.; Kan, L.-S.; Kopka, M. L.; Dickerson, R. E. Helix Geometry, Hydration, and G.A Mismatch in a B-DNA Decamer. Science. 1987, 238, 498–504.

(38)

Diekmann, S., E.; von Kitzing, L. W.; McLaughlin, J. O.; Eckstein, F. The Influence of Exocyclic Substituents of Purine Bases on DNA Curvature. Proc. Natl. Acad. Sci. USA. 1987, 84, 8257– 8261.

(39)

Seela, F.; Berg, H.; Rosemeyer., H. Bending of Oligonucleotides Containing an Isosteric nucleobase:7-Deaza-2’-Deoxyadenosine Replacing dA within d(A)6 Tracts. Biochemistry. 1989, 28, 6193–6198.

(40)

Marky, L. A.; Kupke, D. W. Probing the Hydration of the Minor Groove of A.T Synthetic DNA Polymers by Volume and Heat Changes. Biochemistry. 1989, 28, 9982–9988.

(41)

Chan, S. S.; Breslauer, K. J.; Hogan, M. E.; Kessler, D. J.; Austin, R. H.; Ojemann, J.; Passner, J. M.; Wiles, N. C. Physical Studies of Structural Equilibria within Periodic Poly (dA).poly (dT) Sequences. Biochemistry. 1990, 29, 6161–6171.

(42)

Segal, E.; Widom, J. Poly(dA:dT) Tracts: Major Determinants of Nucleosome Organization. Curr. Opin. Struct. Biol. 2009, 19, 65–71.


Page 31 of 41

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(43)

Wei, D.; Wilson, W. D.; Neidle, S. Small-Molecule Binding to the DNA Minor Groove Is Mediated by a Conserved Water Cluster. J. Am. Chem. Soc. 2013, 135, 1369–77.

(44)

Zubatiuk, T. A.; Shishkin, O. V; Gorb, L.; Hovorun, D. M.; Leszczynski, J. B-DNA Characteristics Are Preserved in Double Stranded d(A)3d(T)3 and d(G)3d(C)3 Mini-Helixes: Conclusions from DFT/M06-2X Study. Phys. Chem. Chem. Phys. 2013, 15, 18155–66.

(45)

Zelený, T.; Hobza, P.; Kabelác, M. Microhydration of Guanine...cytosine Base Pairs, a Theoretical Study on the Role of Water in Stability, Structure and Tautomeric Equilibrium. Phys. Chem. Chem. Phys. 2009, 11, 3430–5.

(46)

Close, D. M.; Crespo-Hernández, C. E.; Gorb, L.; Leszczynski, J. The Influence of Microhydration on the Ionization Energy Thresholds of Uracil and Thymine. J. Phys. Chem. A. 2005, 109, 9279– 83.

(47)

Close, D. M.; Crespo-Hernandez, C. E.; Gorb, L.; Leszczynski, J. Influence of Microhydration on the Ionization Energy Thresholds of Thymine: Comparisons of Theoretical Calculations with Experimental Values. J. Phys. Chem. A. 2006, 110, 7485–90.

(48)

Close, D. M. Ionization Energy Thresholds of Microhydrated Adenine and Its Tautomers. J. Phys. Chem. A. 2008, 112, 12702–6.

(49)

Neidle, S. Nucleic Acid Structure and Recognition; Oxford University Press, 2002; p. 272.

(50)

Lu, X.-J. 3DNA: A Software Package for the Analysis, Rebuilding and Visualization of ThreeDimensional Nucleic Acid Structures. Nucleic Acids Res. 2003, 31, 5108–5121.

(51)

Klamt, A.; Schüürmann, G. COSMO: A New Approach to Dielectric Screening in Solvents with Explicit Expressions for the Screening Energy and Its Gradient. J. Chem. Soc. Perkin Trans. 2. 1993, 799.

(52)

Grimme, S. Semiempirical GGA-Type Density Functional Constructed with a Long-Range Dispersion Correction. J. Comput. Chem. 2006, 27, 1787–1799.

(53)

Grimme, S.; Antony, J.; Ehrlich, S.; Krieg, H. A Consistent and Accurate Ab Initio Parametrization of Density Functional Dispersion Correction (DFT-D) for the 94 Elements H-Pu. J. Chem. Phys. 2010, 132, 154104–154122.

(54)

Schäfer, A.; Horn, H.; Ahlrichs, R. Fully Optimized Contracted Gaussian Basis Sets for Atoms Li to Kr. J. Chem. Phys. 1992, 97, 2571–2577.

(55)

Goerigk, L.; Grimme, S. A Thorough Benchmark of Density Functional Methods for General Main Group Thermochemistry, Kinetics, and Noncovalent Interactions. Phys. Chem. Chem. Phys. 2011, 13, 6670–6688.

(56)

Grimme, S.; Ehrlich, S.; Goerigk, L. Effect of the Damping Function in Dispersion Corrected Density Functional Theory. J. Comp. Chem. 2011, 32, 1456–65.

(57)

Ahlrichs, R.; Bär, M.; Häser, M.; Horn, H.; Kölmel, C. Electronic Structure Calculations on Workstation Computers: The Program System Turbomole. Chem. Phys. Lett. 1989, 162, 165–169.

(58)

Drew, H. R.; Dickerson, R. E. Structure of a B-DNA Dodecamer. III. Geometry of Hydration. J. Mol. Biol. 1981, 151, 535–556.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 32 of 41

(59)

Kopka, M. L.; Fratini, A. V.; Drew, H. R.; Dickerson, R. E. Ordered Water Structure around a BDNA Dodecamer. J. Mol. Biol. 1983, 163, 129–146.

(60)

Edwards, K. J.; Brown, D. G.; Spink, N.; Skelly, J. V.; Neidle, S. Molecular Structure of the B-DNA Dodecamer d(CGCAAATTTGCG)2. An Examination of Propeller Twist and Minor Groove Water Structure at 2.2Å Resolution. J. Mol. Biol. 1992, 226, 1161–1173.

(61)

Poltev, V. I.; Teplukhin, A. V.; Malenkov, G. G. Computational Investigation of the Role of Hydration in Nucleic Acid Structure and Function. Int. J. Quant. Chem. 1992, 42, 1499–1514.

(62)

Olson, W. K.; Bansal, M.; Burley, S. K.; Dickerson, R. E.; Gerstein, M.; Harvey, S. C.; Heinemann, U.; Lu, X. J.; Neidle, S.; Shakked, Z.; Sklenar, H.; Suzuki, M.; Tung, C. S.; Westhof, E.; Wolberger, C.; Berman, H. M. A Standard Reference Frame for the Description of Nucleic Acid Base-Pair Geometry. J. Mol. Biol. 2001, 313, 229–37.

(63)

Lavery, R.; Zakrzewska, K.; Beveridge, D.; Bishop, T. C.; Case, D. a; Cheatham, T.; Dixit, S.; Jayaram, B.; Lankas, F.; Laughton, C.; Maddocks, J. H.; Michon, A.; Osman, R.; Orozco, M.; Perez, A.; Singh, T.; Spackova, N.; Sponer, J. A Systematic Molecular Dynamics Study of Nearest-Neighbor Effects on Base Pair and Base Pair Step Conformations and Fluctuations in BDNA. Nucleic Acids Res. 2010, 38, 299–313.

(64)

Hogan, M.; LeGrange, J.; Austin, B. Dependence of DNA Helix Flexibility on Base Composition. Nature. 1983, 304, 752–754.

(65)

Podtelezhnikov, A. A.; Mao, C.; Seeman, N. C.; Vologodskii, A. Multimerization-Cyclization of DNA Fragments as a Method of Conformational Analysis. Biophys. J. 2000, 79, 2692–704.

(66)

Zhang, Y.; Xi, Z.; Hegde, R. S.; Shakked, Z.; Crothers, D. M. Predicting Indirect Readout Effects in Protein-DNA Interactions. Proc. Natl. Acad. Sci. U. S. A. 2004, 101, 8337–41.

Table of Contents artwork


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Figure 1. Definition of the helix`s bending angle, β. The angle β represents the angle between two average planes. These planes are defined by four atoms: C1′, N9 of adenine and C1′, N1 of thymine (showing as balls). Numbers corresponded to the site numbering for A-T nucleobase. 74x108mm (300 x 300 DPI)



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Figure 2. (a) The minor groove scheme of the AT-containing DNA fragment. The sequence chain is numbered in the 5'-3' direction; (b) two adjacent A-T pairs; (c) the known structures of water patterns p1, p2 are shown here for the adjacent base pairs which are connected on the scheme (a) and (b) with solid line through the atoms N3 and O2. Three hydrophilic centers (N3 of adenine, O2 of thymine and O4 sugar oxygens) are shown as open circles with numbers corresponded to the site numbering for A-T nucleobase. The groove profiles with water molecules on (c) represent cross section through the atoms O4'(i+3)N3(i+2)-O2(i+1')-O4'(i') taken from scheme (a). 169x97mm (300 x 300 DPI)


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Figure 3. The initial locations of structural waters along the minor groove in W1 and W2 models of complexes of (dA:dT)5 with 10 water molecules. The DNA bases are shown as adenine N3 and thymine O2 hydration sites. Water molecules are colour coded according to their location in different layers: from first to second as grey and white. (a) Model W1 based on the p1 water pattern, two water layers; (b) model W2 based on the p2 water pattern, one water layer. 169x55mm (300 x 300 DPI)



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Figure 4. (a) The major groove scheme of the AT-containing DNA fragment. The sequence chain is numbered in the 5'-3' direction; (b) the structures of water patterns P1-P5 are shown here for three adjacent base pairs which are connected in the scheme (a) with solid line through the atoms O4, N6 and N7. Three hydrophilic centers (O4 of thymine, N6 and N7 of adenine) are shown as open circles with numbers corresponded to the site numbering for A-T nucleobase. The groove profiles with water molecules in (b) represent cross section through the atoms O4(i+2)-N6(i+1)-N7(i+1)-N7'(i) taken from scheme (a). 169x60mm (300 x 300 DPI)


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Figure 5. The initial locations of structural waters along and across the major groove in WW1 and WW2 models of complexes of (dA:dT)5 with water molecules. The DNA bases are shown as adenine N6, N7 and thymine O4 hydration sites. Locations of waters are based on the P1-P5 water patterns (see, Fig.4b) and form one water layer in major groove. 74x42mm (300 x 300 DPI)



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Figure 6. The equilibrium complexes of (dA:dT)5 with structural waters along the minor groove in W1 and W2 models: (a) Model W1 with domination of p1 water pattern; (b) Model W2 with domination of p2 water pattern. The DNA bases are shown as adenine N3 and thymine O2 hydration sites. Water molecules are shown here as spheres colour coded according to their positions in shells - from first to second as grey and white. Solid lines represent the network of hydrogen bonds. The position of Na+ ions is not shown. 74x90mm (300 x 300 DPI)


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Figure 7. The equilibrium complexes of (dA:dT)5 with structural waters along and across the major groove in WW1 and WW2 models: (a) P4, P5 hydration patterns dominate in model WW1; (b) P1-P3 hydration patterns dominate in model WW2. The DNA bases are shown as adenine N6, N7 and thymine O4 hydration sites. Water molecules are shown here as spheres (the water spine in minor groove is omitted). White spheres indicate water molecules which do not belong to any of discussed water patterns. Solid lines represent the network of hydrogen bonds. The position of Na+ ions is not shown. 74x92mm (300 x 300 DPI)



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Figure 8. Schematic pictures of mini-helixes discussed with the local helical axes (dotted line) superimposed. All structures are set with reference to the middle helical frame defined by 1, 2, 3 4, and 5-th base pairs and minor groove facing the viewer (black site). 74x64mm (300 x 300 DPI)


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The nucleobase-water interactions in both grooves prevent AT-tract twisting and its “collapse” along the minor groove. Whereby, helix structure with narrow minor and wide major grooves is formed. Formation of specific water patterns in both grooves is the factor responsible for stabilization of A-tracts with narrowed minor groove, leading in turn to their strong intrinsic bending in DNA. 50x24mm (300 x 300 DPI)


Structural Waters in the Minor and Major Grooves of DNAâA Major

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