Crystallographic structure of an RNA helix: [U(UA)6A]2 (original) (raw)
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Crystal Structure of a Bulged RNA Tetraplex at 1.1 Å Resolution
Structure, 2003
sequence d(GGGCGG) (Gralla et al., 1987; Oppenheim et al., 1992), and Saccharomyces cerevisiae telomere and Biochemistry Yale University repeat d(GGGT) (Zakian, 1989). These fragments may play an important role in biological processes. For in-New Haven, Connecticut stance, the auxiliary downstream element in SV40 liter pre-mRNA r(GGGGGAGGUGUGGG) (Bagga et al., 1995) is bound by hnRNA H/HЈ protein, and their interaction Summary may stimulate the polyadenylation of SV40 liter pre-mRNA (Bagga et al., 1998). Four consecutive guanines Bulges are an important structural motif in RNA and can form guanine tetraplex in both solution (Kim et al., can be used as recognition and interaction sites in 1991; Cheong and Moore, 1992) and crystalline state RNA-protein interaction and RNA-RNA interaction. (Laughlan et al., 1994; Phillips, et al., 1997; Deng et al., Here we report the first crystal structure of a bulged 2001a). Because pyrimidines are smaller than purines RNA tetraplex at 1.1 Å resolution. The hexamer r(U)( Br dG)
Crystallographic studies of DNA and RNA
2009
Our knowledge of nucleic acid structure grew rapidly over the past decade with the determination to high resolution of larger structures of great biological significance. Advances in sample preparation, crystallization techniques, cryocrystallography, access to synchrotron radiation, and crystallographic software continue to accelerate the structure determination of nucleic acids. Crystallographic studies of DNA and RNA molecules share many considerations that we outline here.
Structures and Energetics of Four Adjacent G·U Pairs That Stabilize an RNA Helix
The Journal of Physical Chemistry B, 2015
Consecutive G•U base pairs inside RNA helices can be destabilizing while those at the ends of helices are thermodynamically stabilizing. To determine if this paradox could be explained by differences in base stacking, we determined the high-resolution (1.32 Å) crystal structure of (5'-GGUGGCUGUU-3') 2 and studied three sequences with four consecutive terminal G•U pairs by NMR spectroscopy. In the crystal structure of (5'-GGUGGCUGUU-3') 2 , the helix is overwound but retains the overall features of A-form RNA. The penultimate base steps at each end of the helix have high base overlap and contribute to the unexpectedly favorable energetic contribution for the 5'-GU-3'/3'-UG-5' motif in this helix position. The balance of base stacking and helical twist contributes to the positional dependence of G•U pair stabilities. The energetic stabilities and similarity to A-form RNA helices suggest that consecutive G•U pairs would be recognized by RNA helix binding proteins, such as Dicer and Ago. Thus, these results will aid future searches for target sites of small RNAs in gene regulation.
An Eight-Stranded Helical Fragment in RNA Crystal Structure
Structure, 2003
Multistranded helical structures in nucleic acids play various functions in biological processes. Here we report the crystal structure of a hexamer, rU(BrdG)r(AGGU),at 1.5 Å resolution containing a structural complex of an alternating antiparallel eight-stranded helical fragment that is sandwiched in two tetraplexes. The octaplex is formed by groove binding interaction and base tetrad intercalation between two tetraplexes. Two different forms of octaplexes have been proposed, which display different properties in interaction with proteins and nucleic acids. Adenines form a base tetrad in the novel N6-H…N3 conformation and further interact with uridines to form an adenine-uridine octad in the reverse Hoogsteen pairing scheme. The conformational flexibility of adenine tetrad indicates that it can optimize its conformation in different interactions.
Journal of Molecular Biology, 2000
Crystal structure of a DNA ÁRNA hybrid, d(CTCCTCTTC) Ár(gaagagagag), with an adenine bulge in the polypurine RNA strand was determined at 2.3 A Ê resolution. The structure was solved by the molecular replacement method and re®ned to a ®nal R-factor of 19.9 % (R free 22.2 %). The hybrid duplex crystallized in the space group I222 with unit cell dimensions, a 46.66 A Ê , b 47.61 A Ê and c 54.05 A Ê , and adopts the A-form conformation. All RNA and DNA sugars are in the C3 H-endo conformation, the glycosyl angles in anti conformation and the majority of the C4 H-C5 H torsion angles in g except two trans angles, in conformity with the C3 H-endo rigid nucleotide hypothesis. The adenine bulge is looped out and it is also in the anti C3 H-endo conformation. The bulge is involved in a base-triple (C Á g) * a interaction with the end base-pair (C9 Á g10) in the minor groove of a symmetry-related molecule. The 2 H hydroxyl group of g15 is hydrogen bonded to O2P and O5 H of g17, skipping the bulged adenine a16 and stabilizing the sugar-phosphate backbone of the hybrid. The hydrogen bonding and the backbone conformation at the bulged adenine site is very similar to that found in the crystal structure of a protein-RNA complex.
Solution Structure of an RNA Duplex Including a C−U Base Pair † , ‡
Biochemistry, 2000
The formation of the C-U base pair in a duplex was observed in solution by means of the temperature profile of 15 N chemical shifts, and the precise geometry of the C-U base pair was also determined by NOE-based structure calculation. From the solution structure of the RNA oligomer, r[CGACUCAGG]‚r[CCUGCGUCG], it was found that a single C-U mismatch preferred being stacked in the duplex rather than being flipped-out even in solution. Moreover, it adopts an irregular geometry, where the amino nitrogen (N4) of the cytidine and keto-oxygen (O4) of the uridine are within hydrogenbonding distance, as seen in crystals. To further prove the presence of a hydrogen bond in the C-U pair, we employed a point-labeled cytidine at the exocyclic amino nitrogen of the cytidine in the C-U pair. The temperature profile of its 15 N chemical shift showed a sigmoidal transition curve, indicating the presence of a hydrogen bond in the C-U pair in the duplex.
Structure of an RNA Internal Loop Consisting of Tandem C-A+ Base Pairs
Biochemistry, 1998
The crystal structure of the RNA octamer 5′-CGC(CA)GCG-3′ has been determined from X-ray diffraction data to 2.3 Å resolution. In the crystal, this oligomer forms a self-complementary double helix in the asymmetric unit. Tandem non-Watson-Crick C-A and A-C base pairs comprise an internal loop in the middle of the duplex, which is incorporated with little distortion of the A-form double helix. From the geometry of the C-A base pairs, it is inferred that the adenosine imino group is protonated and donates a hydrogen bond to the carbonyl group of the cytosine. The wobble geometry of the C-A + base pairs is very similar to that of the common U-G non-Watson-Crick pair. The picture of RNA molecules being structurally and functionally limited by the use of only four building blocks and by the rules of Watson-Crick base pairing is undergoing a radical change. Noncanonical base pairing within doublehelical regions of RNA has been structurally characterized (1-5) and shown to be functionally important as binding sites for protein (6, 7) and RNA (8), in active sites of catalytic RNA (9), and in bending RNA into a folded form (10, 11). Several of the noncanonical base pairs are polymorphic (12, 13), assuming a different pairing geometry depending on their environments or the solution conditions. We must also consider that at least some of the bases involved in noncanonical interactions may exist in different charged or protonated forms in different environments. For example, protonation of adenine and cytosine has been observed in the formation of C-A + (14) and CC + (15, 16) base pairs. We report here the 2.3 Å resolution (Table 1) X-ray structure of an RNA octamer which in the crystal forms a double helix incorporating tandem C-A-A-C base pairs bounded by standard Watson-Crick pairs. From the geometry of these noncanonical base pairs, it is inferred that they are C-A + pairs with a protonated N1 of the adenine donating a hydrogen bond to the cytosine carbonyl group. This "wobble" C-A + base pairing is structurally similar to, and in some cases may be a substitute for, the more common U-G wobble pair. MATERIALS AND METHODS Synthesis, Crystallization, and Data Collection. The octaribonucleotide rCGCCAGCG was chemically synthesized (Oligos, Etc.) and purified by anion exchange (DEAE-5PW TSK-Gel, TosoHaas) FPLC 1 with a linear salt gradient from 0.4 to 2.0 M sodium acetate and 50 mM Tris-HCl (pH 6.8-7.3). Crystals were grown at room temperature by the hanging drop vapor diffusion method from a 2.4 µL drop of a solution of 2 mM RNA mixed with an equal volume of the reservoir solution consisting of 20 mM NaCl, 5 mM MgCl 2 , 2 mM spermine tetrahydrochloride, and 35% MPD (2-methyl-2,4-pentanediol). The crystals were originally grown from a buffer of 50 mM sodium cacodylate (pH 6.5), but superior crystals were later grown from a buffer of 100 mM sodium cacodylate (pH 5.5). The crystals grown at pH 6.5 were thin hexagonal plates with a maximum dimension 100 µm, while those grown at the lower pH were rods that grew to dimensions of 350 µm × 40 µm × 40 µm. X-ray data were collected on a Rigaku R-axis IIC imaging plate system using CuKR radiation (λ) 1.5418 Å) and φ
Nucleic Acids Research, 1995
A crystal structure has been solved for an analog of the r(ApU) ribodinucleotide, r(Aso2U), where a bridging non-ionic dimethylene sulfone linker replaces the phosphodiester linking group found in natural RNA. Crystals of the single-stranded state of r(Aso2U) were obtained from water at 500C. In these crystals, one hydrogen bond is formed between bases from different strands and base stacking occurs in intermolecular 'homo-A' and 'homo-U' stacks. Similar to typical oligoribonucleotides, the ribose rings adopt N-type conformations and dihedral angles X are in the anti range. The all-trans rotamer of the CH2-SO2-CH2-CH2 bridge was found, which leads to a large adenine-uracil distance. Qualitative analysis of a NOESY spectrum of the Aso2U part in r(Uso2Cso2Aso2U) dissolved in a dimethylsulfoxide-D20 mixture indicates that the conformation observed in the crystal is also populated in solution. Comparison with the structure of r(Gso2C), which has been crystallized in the Watson-Crick paired state, shows that a rotation around 4 by +1120 leads from the observed, single-stranded state to a conformation that is compatible with formation of a duplex. A concerted trans/gauche flip of a and y then yields the standard conformer of A-type RNA helices. From the observed structure of r(Gso2C) and other oligonucleotides it is anticipated that this flip will also revert the ribose pucker from C2'-exo to C3'-endo.
RNA, 2000
Chemically modified nucleotide analogs have gained widespread popularity for probing structure-function relationships. Among the modifications that were incorporated into RNAs for assessing the role of individual functional groups, the phenyl nucleotide has displayed surprising effects both in the contexts of the hammerhead ribozyme and pre-mRNA splicing. To examine the conformational properties of this hydrophobic base analog, we determined the crystal structure of an RNA double helix with incorporated phenyl ribonucleotides at 1.97 Å resolution. In the structure, phenyl residues are engaged in self-pairing and their arrangements suggest energetically favorable stacking interactions with 39-adjacent guanines. The presence of the phenyl rings in the center of the duplex results in only moderate changes of the helical geometry. This finding is in line with those of earlier experiments that showed the phenyl analog to be a remarkably good mimetic of natural base function. Because the stacking interactions displayed by phenyl residues appear to be similar to those for natural bases, reduced conformational restriction due to the lack of hydrogen bonds with phenyl as well as alterations in its solvent structure may be the main causes of the activity changes with phenyl-modified RNAs.
An eight-stranded helical fragment in RNA crystal structure: implications for tetraplex interaction
Structure (London, England : 1993), 2003
tetrads (Patel et al., 1999b; Pan et al., 2003) have been observed in both NMR and crystal structures. These 200 Johnston Lab 176 West 19th Avenue base tetrads show that hydrogen bond donors and acceptors are also exposed in the groove, and tetraplexes Columbus, Ohio 43210 2 Department of Molecular Biophysics are expected to have strong interactions in their groove. Recent research has been conducted on the interactions and Biochemistry Yale University of tetraplexes and the possible pharmaceutical applications to the tetraplexes (Kerwin, 2000; Neidle and Read, New Haven, Connecticut 06511 2001). However, we still have little structural information about the interaction.