DNA-RNA hybrid secondary structures

DNA-RNA and DNA-DNA duplexes are even more polymorphic than observed previously. DNA-RNA hybrids can have secondary structures like A-DNA or A-RNA, but double helices of the synthetic DNA-RNA hybrids poly(dA) X poly(rU) and poly(dI) X poly(rC), respectively, form 11-fold and 10-fold double-helical structures in which the two chains have quite different conformations. Extensive X-ray fiber diffraction analyses show that in both structures the DNA chains have C-2'-endo-puckered furanose rings, while the anti-parallel RNA chains have C-3'-endo-puckered rings. The bidirectional properties of such duplexes may be important in the transfer of biological information from nucleic acids.     

Action of ribonuclease BS-1 on a DNA-RNA hybrid

Evidence has been obtained that ribonuclease BS-1, structurally and catalytically related to ribonuclease A, is capable of degrading a DNA-RNA hybrid. This finding has been discussed with regard to the recently reported biological effects of the enzyme.     

Crystallization and preliminary crystallographic investigation of a DNA-RNA hybrid

A complex between the complementary hexanucleotides ribo-C-A-A-A-A-G and deoxyribo-C-T-T-T-T-G has been crystallized. The space group was determined to be P2₁ with unit cell dimensions $\text{a = 26.9}\text{A}\text{?}\text{, b = 50.9}\text{A}\text{?}\text{, c = 62.0}\text{A}\text{?}\text{; α = γ = 90 °, β = 108 °}$. The crystals contain both the ribo- and the deoxyribo-hexanucleotide strands in a ratio of approximately 1:1. Each asymmetric unit contains the equivalent of four hybrid molecules.     

Theoretical studies of DNA-RNA hybrid conformations.

Molecular modelling has been used to probe the conformational preferences of double stranded DNA-RNA hybrids. As might be expected, the sugars of the DNA strand have higher conformational flexibility, but, for the majority of the repetitive sequences studied, these sugars prefer a C2-endo pucker, while ribose sugars uniformly adopt a C3-endo pucker. This gives rise to a strongly heteronomous duplex conformation. One exception to this rule involves the thymidine strand of poly(dT).poly(rA), which marginally prefers a C3-endo pucker. Our study further indicates that the DNA strands of the hybrids favour backbone torsions in the canonical B domain, rather than the modified values proposed on the basis of fibre diffraction studies. Backbone conformational transitions can nevertheless be induced leading to an alpha gamma-flip (alpha:gamma, g-/g(+)-->t/t) or to the alpha beta gamma-flip form proposed from fibre studies (alpha:beta:gamma, g-/t/g(+)-->t/g+/t). The latter transition is also found to be linked to BI-->BII transitions (epsilon:zeta, t/g(-)-->g-/t).     

DNA-RNA hybridization.

Interest in nucleic acid hybridization stems mainly from its great power as a tool in biological research. It is used in several quite distinct ways. Because of the high degree of specificity that they show, hybridization techniques can be used to measure the amount of one specific sequence within a very heterogeneous mixture of sequences. Measurements of 1/10(6)-10(7) have been recorded. In extension of this, various properties of a specific sequence can often be studied. Secondly, because the kinetics of nucleic acid hybridization are quite well understood, it can be used to characterize both a pure sequence and a very complex mixture of sequences, like the genome of a vertebrate. Thirdly, again because of its specificity, it can be used to measure homologies between different populations of nucleic acids. Lastly, in conjunction with other techniques, it can be used as a basis for the fractionation of nucleic acid populations and the purification of specific sequences. Specific examples of these applications are given, with special reference to the organization of the genome in higher eukaryotes.     

Renaturation of bacteriophage phiX174 DNA-RNA hybrid: RNA length effect and nucleation rate constant

A one to one RNA-DNA hybrid was synthesized using bacteriophage φX174 DNA as a template for the Escherichia coli RNA polymerase system. The renaturation rate for the hybrid was found to vary as the square root of the molecular weight of the RNA. The RNA molecular weights were determined using a standardized dimethylsulfoxide, cesium sulfate equilibrium density gradient. The optimum nucleation rate constant for the renaturation of the hybrid in 0.4 m-sodium chloride was found to be 1.3 × 10⁵, as compared with to 1.7 × 10⁵ for DNA renaturation.     

CD of homopolymer DNA-RNA hybrid duplexes and triplexes containing A-T or A-U base pairs.

CD spectra and difference-CD spectra of (a) two DNA X RNA hybrid duplexes (poly[r(A) X d(U)] and poly[r(A) X d(T)]) and (b) three hybrid triplexes (poly-[d(T) X r(A) X d(T)], poly[r(U) X d(A) X r(U)], and poly[r(T) X d(A) X r(T)]) were obtained and compared with CD spectra of six A X U- and A X T-containing duplex and triplex RNAs and DNAs. We found that the CD spectra of the homopolymer duplexes above 260 nm were correlated with the type of base pair present (A-U or A-T) and could be interpreted as the sum of the CD contributions of the single strands plus a contribution due to base pairing. The spectra of the duplexes below 235 nm were related to the polypurine strands present (poly-[r(A)] or poly[d(A)]). We interpret the CD intensity in the intermediate 255-235 nm region of these spectra to be mainly due to stacking of the constituent polypurine strands. Three of the five hybrids (poly[r(A) X d(U)], poly[r(A) X d(T)], and poly[d(T) X r(A) X d(T)]) were found to have heteronomous conformations, while poly[r(U) X d(A) X r(U)] was found to be the most A-like and poly[r(T) X d(A) X r(T)], the least A-like.