Water and Ions in a High Resolution Structure of B-DNA

Soler-López, Montserrat; Malinina, Lucy; Liu, Jie; Huynh‐Dinh, Tam; Subirana, Juan A.

doi:10.1074/jbc.274.34.23683

Cited by 75 publications

(66 citation statements)

References 21 publications

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“…Minor Groove Structure-The central region of the duplex has a spine of hydration (5,6,9) identical to that found in the P2 1 2 1 2 1 structure, as it has been discussed in detail elsewhere (21). At both ends of the spine of hydration, there are two heavily hydrated Ca 2ϩ ions.…”

Section: Alcium Form Of D(cgcgaattcgcg)mentioning

confidence: 67%

Structure of d(CGCGAATTCGCG) in the Presence of Ca2+Ions

Liu¹,

Subirana²

1999

Journal of Biological Chemistry

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The dodecamer d(CGCGAATTCGCG) was the first oligonucleotide to be crystallized as a B-DNA duplex. Its structure was analyzed in detail in the early 1980s. Here we show that, in the presence of Ca 2؉ , it crystallizes in a different way (R3 space group). The dodecamers form parallel columns of straight duplexes with ten base pairs in the B form. The terminal cytosines in each molecule are disordered, whereas the terminal guanines are placed in the minor groove of neighbor duplexes. The central GAATTC region is practically identical to that found in the classic structure of the same dodecamer crystallized in the P2 1 2 1 2 1 space group in the presence of Mg 2؉ and spermine. Its structure is thus independent of the crystallization conditions which have been used.The first detailed structure of a DNA oligonucleotide in the B-form determined by single crystal x-ray diffraction methods was published in this journal in 1982 (1). The structure of a bromine derivative was compared with that previously determined (2, 3) for the native dodecamer at different temperatures. Its features were analyzed in detail in several other publications (4 -7). Many other derivatives from the same oligonucleotide and related sequences, alone and in association with drugs, have also been studied. Coordinates and references may be found in the Nucleic Acid Database (8). Most of them are in practically identical unit cells in the P2 1 2 1 2 1 space group. The original work (1-3) was a landmark in the study of DNA because it confirmed unequivocally the double helical structure of B-form DNA and, at the same time, showed many features of conformation as a function of sequence. More recently, the same dodecamer has been crystallized under various ionic conditions (9 -13). A high resolution (1.4 Å) was obtained (9) in the presence of Na ϩ /Mg 2ϩ /spermine, giving a structure essentially similar to those first reported by Dickerson and co-workers (1-3). Given the higher resolution of that structure (9), we used it for comparison with our results.Most of the work described in the previous paragraph was carried out with crystals obtained in the presence of Mg 2ϩ and spermine. Recently we discovered (14) that, in the presence of a high concentration of Ca 2ϩ ion, this oligonucleotide could crystallize in the R3 space group. Here we report the results we have obtained under the latter crystallization conditions at a higher resolution than that previously reported (14). Our results show that the central sequence GAATTC is practically identical in both cases, whereas the conformation of the terminal CGC/GCG sequences is much more variable because of the interaction with neighbor molecules in the crystal. EXPERIMENTAL PROCEDURESCrystallization-The crystal was grown by the vapor diffusion hanging drop method. The crystallization solution contained 0.4 mM dodecamer (NH 4 ϩ salt), 300 mM CaCl 2 , 10% MPD (2-methyl-2,4-pentanediol), and 20 mM cacodylic acid (pH 7.0). The concentration of monovalent cations (Na ϩ from the buffer and NH 4 ϩ from the dodecame...

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Section: Alcium Form Of D(cgcgaattcgcg)mentioning

confidence: 67%

Structure of d(CGCGAATTCGCG) in the Presence of Ca2+Ions

Liu¹,

Subirana²

1999

Journal of Biological Chemistry

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“…R-free was calculated with 5% of the data excluded from the refinement. of different cations (Na ϩ , K ϩ , Mg 2ϩ , Ca 2ϩ ) were determined at high resolution (29)(30)(31)(32)(33)(34)(35), and several of them were isomorphous to the Dickerson-Drew dodecamer (36). The superpositioning of such structures with the present one is shown in Fig.…”

Section: Resultsmentioning

confidence: 90%

DNA bending by an adenine–thymine tract and its role in gene regulation

Hizver

Rozenberg

Frolow

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

138

153

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To gain insight into the structural basis of DNA bending by adenine-thymine tracts (A-tracts) and their role in DNA recognition by gene-regulatory proteins, we have determined the crystal structure of the high-affinity DNA target of the cancer-associated human papillomavirus E2 protein. The three independent B-DNA molecules of the crystal structure determined at 2.2-Å resolution are examples of A-tract-containing helices where the global direction and magnitude of curvature are in accord with solution data, thereby providing insights, at the base pair level, into the mechanism of DNA bending by such sequence motifs. A comparative analysis of E2-DNA conformations with respect to other structural and biochemical studies demonstrates that (i) the A-tract structure of the core region, which is not contacted by the protein, is critical for the formation of the high-affinity sequence-specific protein-DNA complex, and (ii) differential binding affinity is regulated by the intrinsic structure and deformability encoded in the base sequence of the DNA target.A-tract ͉ papillomavirus E2-DNA target ͉ transcriptional control T he relationship between DNA sequence, structure, and function has been studied and discussed extensively for the last 20 years. A particular effort has been directed toward the structural elucidation of short runs of four to six adeninethymine residues, known as A-tracts, in an attempt to reveal the structural basis of DNA curvature induced by such sequence motifs when they are inserted in phase with the helical periodicity (1, 2). Despite numerous efforts, including x-ray crystallographic and solution studies, the structural basis of A-tractinduced bending has remained enigmatic (3). Because no single structure could explain the whole phenomenon, it was necessary to rely on models, several of which had been proposed (4). They generally conform to the gel migration data, which suggest that the center of curvature is toward the minor groove of the A-tracts and toward the major groove of the intervening general sequences (5). However, they differ substantially in the details of the stereochemical origin of curvature. This issue is of particular biological significance, as sequence-dependent DNA curvature or bending is an important determinant of DNA recognition by proteins (6).The E2 regulatory system of human papillomaviruses provides an example where sequence-specific binding of proteins to A-tracts is crucial to the organism's life cycle. The E2 proteins from all viral strains activate or repress transcription in a context-dependent manner and are required for the initiation of replication in vivo. Their function depends on sequence-specific binding to a highly conserved 12-bp sequence of the general form ACCGNNNNCGGT, where N4 is variable (7,8). However, the E2 binding sites in the human papillomavirus (HPV) genomes, including the cancer-related strains HPV-16 and HPV-18 (9), exhibit a further level of specificity in the interaction with their cognate E2 proteins. This specificity is manifested by th...

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“…The spine of hydration, which we analyzed in a previous paper (2), is also shown in yellow in Fig. 13.…”

Section: Discussionmentioning

confidence: 90%

“…Therefore, we released this restraint from the dictionary, lowering the R factor from 0.148 to 0.135. This restraint release was not introduced in our preliminary model (2).…”

Section: Methodsmentioning

confidence: 99%

Solvent Organization in an Oligonucleotide Crystal

Soler-López¹,

Malinina²,

Subirana³

2000

Journal of Biological Chemistry

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We describe the crystal structure of d(GCGAATTCG) determined by x-ray diffraction at atomic resolution level (0.89 Å). The duplex structure is practically identical to that described at 2.05 Å resolution (Van Meervelt, L., Vlieghe, D., Dautant, A., Gallois, B., Pré cigoux, G., and Kennard, O. (1995) Nature 374, 742-744), however about half of the phosphate groups show multiple conformations. The crystal has three regions with different solvent structure. One of them contains several ordered Mg ؉2 ions and can be considered as an ionic crystal. A second region is formed by a network of ordered water molecules with a polygonal organization that binds three duplexes. The third region is formed by channels of solvent in which very few ordered solvent molecules are visible. The less ordered phosphates are found facing this channel. The latter region provides a view of DNA with highly movable charges, both negative phosphates and counterions, without a precise location.Two periods could be distinguished in the x-ray diffraction studies of DNA: the era of fiber diffraction of polymeric DNA/ polynucleotides and the time of single crystal x-ray structures of short fragments. The data available from fiber diffraction were limited and allowed no detailed analysis. The goal of DNA crystallography is, therefore, the study of detailed DNA structures. However, crystallographic data are also limited in some important aspects of DNA structure. For example, it is well known (1) that the solvent environment plays an important role in the conformational behavior and interactions of DNA. To study this feature in detail, it is necessary to work with x-ray data at atomic resolution level. Until recently, such data were available only for Z-DNA fragments. This situation is now changing. Synchrotron radiation of increasing power, together with better detectors and cryotechniques, make possible a third period of DNA crystallography: the epoch of high resolution DNA structures.We recently presented (2) a preliminary account of the structure of the B-DNA fragment (GCGAATTCG) solved at atomic resolution (0.89 Å). This structure had already been solved at 2.05-Å resolution (3, 4). In our previous analysis we described the water spine in the minor groove and compared it with related structures. Here we present a detailed analysis of the same oligonucleotide, which has multiple conformations in several parts of the phosphodiester backbone. We also describe the organization of the solvent, which in some regions shows well resolved water molecules, whereas in other regions it remains disordered, despite the fact that our crystals have been studied at 120 K. Thus, a frozen sample at atomic resolution, instead of showing a highly ordered duplex, demonstrates multiple conformations and regions of disordered solvent. MATERIALS AND METHODSCrystallization-The d(GCGAATTCG) nonamer was crystallized, using a batch method, in sitting drops containing 0.5 mM DNA duplex, 1 mM acridine-(Arg 4 ) drug-peptide aduct, 20 mM sodium cacodylate buffer, pH 7, 100 mM...

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Water and Ions in a High Resolution Structure of B-DNA

Cited by 75 publications

References 21 publications

Structure of d(CGCGAATTCGCG) in the Presence of Ca2+Ions

Structure of d(CGCGAATTCGCG) in the Presence of Ca2+Ions

DNA bending by an adenine–thymine tract and its role in gene regulation

Solvent Organization in an Oligonucleotide Crystal

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