The Tautomeric Form of Helical Polyribocytidylic Acid

BiochemistryStable loop in the crystal structure of the intercalated four-stranded cytosine-rich metazoan telomere (C-C+ ABSTRACTIn most metazoans, the telomeric cytosinerich strand repeating sequence is d(TAACCC). The crystal structure of this sequence was solved to 1.9-A resolution. Four strands associate via the cytosine-containing parts to form a four-stranded intercalated structure held together by C C+ hydrogen bonds. The base-paired strands are parallel to each other, and the two duplexes are intercalated into each other in opposite orientations. One TAA end forms a highly stabilized loop with the 5' thymine Hoogsteen-base-paired to the third adenine. The 5' end of this loop is in close proximity to the 3' end of one of the other intercalated cytosine strands. Instead of being entirely in a DNA duplex, this structure suggests the possibility ofan alternative conformation for the cytosine-rich telomere strands.

Section: Discussionmentioning

confidence: 78%

Stable loop in the crystal structure of the intercalated four-stranded cytosine-rich metazoan telomere.

Kang

Berger

Lockshin

et al. 1995

“…This was first observed in the single-crystal analysis of cytosine-5-acetic acid (13). Hemiprotonation was also used in interpreting the structure of polyribocytidylic acid (1), and solution studies of that polymer (2,4) as well as polydeoxycytidylic acid (3) clearly illustrated the importance of hemiprotonation. It is interesting that crystals of d(C3T) were grown by using cacodylate buffers that were fixed at pH 6.0 as well as pH 7.0.…”

Section: Discussionmentioning

confidence: 89%

“…For some time it has been known that nucleic acids containing stretches of cytidine residues can form parallel strands held together by cytosine-protonated cytosine base pairs (C-C+) (1)(2)(3)(4). In an NMR analysis of d(TC5), Gudron and his associates (5) proposed an unusual structure in which two such parallel-stranded duplexes, held together by C'C+ base pairs, intercalate with each other in opposite polarity to form a four-stranded molecule.…”

mentioning

confidence: 99%

Crystal structure of intercalated four-stranded d(C3T) at 1.4 A resolution.

Kang

Berger

Lockshin

et al. 1994

151

141

The crystal structure of d(C3T), solved at 1.4 A resolution, reveals that the molecule forms a four-stranded intercalated complex. It consists of two parallel-stranded duplexes, each of which Is held together by cytosine-protonated cytosine base pairs. The two duplexes are intercalated with each other and have opposite strand orientation. The molecule has a flat, lath-like appearance, and the covalently bonded cytosines have a slow right-handed twist of 17.1°. However, there is considerable asymmetry. On one of the flat sides, the phosphate groups are rotated away from the center of the molecule. They are held In this orientation by bridging water molecules that bind the NH of cytosine and a phosphate group of an opposite chain. There is also considerable microheterogeneity in the structure. The cytosine hemiprotonation occurs even at pH 7 where stable crystals form.For some time it has been known that nucleic acids containing stretches of cytidine residues can form parallel strands held together by cytosine-protonated cytosine base pairs (C-C+) (1-4). In an NMR analysis of d(TC5), Gudron and his associates (5) proposed an unusual structure in which two such parallel-stranded duplexes, held together by C'C+ base pairs, intercalate with each other in opposite polarity to form a four-stranded molecule. The evidence for the structure was based on strong interactions between the Hi' protons on different strands, suggesting that the backbones were close together. Recent crystal structures of d(TAACCC) (C.K., I.B., C.L., R.R., R.M., and A.R.), (unpublished data) and of d(C4) (6) were in agreement with the general conclusions deduced by the NMR studies. In telomeres at the end of chromosomes, stretches of cytosine residues are found, usually linked to thymine residues. Here, we present a crystallographic analysis of the structure of d (C3T) Rotation searches of all the models indicated a molecule with the helical axis approximately parallel to the crystallographic b axis, as we expected. Subsequent translation searches of some of the models yielded positions for the molecule that were free of close van der Waals contacts. Rigid body refinement followed by intensive simulated annealing using the program XPLOR (7) improved the density around the phosphate groups ofthe backbone and allowed us to build another model with more accurate helical twist based on the phosphate positions. We used this improved molecule as a starting model and molecular replacement was performed again. From the very beginning, we could identify two thymine residues stacked on top of six C-C+ layers and added those to the model. At this stage, simulated annealed omit maps (6) clearly showed the location of the remaining two thymine residues stacked perpendicular to the helix axis. Positional refinement followed by refinement of the temperature factors led to a final R factor of 17.7% for 5013 reflections above the 2cr level (based on F.) between 10 and 1.4 A. The free R factor (8) value based on a random 10%o subset of reflections is 22.5%...

“…However, this apparent anomaly may be explained by the fact that the protonation of the C-C pair leads to a very stable structure, which shifts upward the pK of the cytosine base. Indeed, alkaline titration studies of poly(dC) (26) and poly(rC) (27)(28)(29) have shown that the pH of the melting transition (pHm) is shifted upwards from the pK of cytosine. It has further been shown that the magnitude of the shift (pHm minus pK) is proportional to the stability of the base-paired helix-i.e., the melting temperature, Tm (26,27).…”

Section: Resultsmentioning

confidence: 99%

X-ray-structure of a cytidylyl-3',5'-adenosine-proflavine complex: a self-paired parallel-chain double helical dimer with an intercalated acridine dye.

Westhof¹,

Sundaralingam²

1980

The non-self-complementary dinucleoside monophosphate cytidylyl-3',5'-adenosine (CpA) forms a basepaired parallel-chain dimer with an intercalated proflavine.The dimer complex possesses a right-handed helical twist. The dimer helix has an irregular girth with a neutral adenine-adenine (A-A) pair, hydrogen-bonded through the N6 and N7 sites (C1'... C1' separation of 10.97 A), and a triply hydrogen-bonded protonated cytosine-cytosine (C-C) pair with a proton shared between the base N3 sites (C1'... C1' separation of 9.59 A). The torsion angles of the sugar-phosphate backbone are within their most preferred ranges and the sugar puckering sequence (5' -3') is C3'-endo, C2'-endo. There is also a second proflavine molecule sandwiched between CpA dimers on the 21-axis. Both proflavines are necessarily disordered, being on dyad axis, and this suggests possible insights into the dynamics of intercalation of planar drugs. This structure shows that intercalation of planar drugs in nucleic acids may not be restricted to antiparallel complementary Watson-Crick pairing regions and provides additional mechanisms for acridine mutagenesis.The genetic molecules DNA and RNA are targets for various chemical agents that possess carcinogenic and mutagenic properties. The molecular modes of interactions of these agents with the nucleic acid and the effects they produce on the molecular geometry and conformations of DNA and RNA have been a subject of considerable interest (1-4). X-ray diffraction studies of DNA fibers (2, 5) and of co-crystals of self-complementary dinucleoside monophosphate (6, 7) with the dye proflavine have provided important insights on the stereochemistry of intercalation of proflavine into nucleic acids. Because the known single-crystal complexes, in which the dye is intercalated between the base pairs, involve self-complementary chains, it was of interest to investigate whether similar complexes can be formed between dyes and non-self-complementary nucleic acid fragments. Also, it has been demonstrated that proflavine causes a direct inhibition of protein synthesis (8,9) and that it impairs the enzymic aminoacylation of tRNA (10). Binding studies led to the conclusion that proflavine inhibition of protein synthesis results from strong binding of proflavine to tRNA (11,12). The evidence for intercalation of proflavine in tRNA was, however, indirect (11,12). The binding of proflavine to yeast tRNAPhe was studied by x-ray crystallography after diffusing proflavine into pregrown crystals of yeast tRNAPhe (13). Intercalation of proflavine in helical stems was not found; instead, the binding sites involve electrostatic and hydrogen bonding interactions with the polynucleotide sugar-phosphate backbone (14, t).Here, we report the crystal and molecular structure of an intercalated proflavine in a novel self-paired right-handed parallel-chain double helical dimer of cytidylyl-3',5'-adenosine (CpA). The dinucleoside monophosphate CpA was chosen not only because it constitutes the end portion of the invariant CCA t...