RNA Triple Helix Research Articles

Surface-Enhanced Fourier Transform Raman Scattering Study on the Adsorption Structure of an RNA Triple Helix at a Silver Electrode

In this research, we studied the adsorption structure of an RNA triplex, poly[rU]poly[rA]poly[rU], at a silver electrode by surface-enhanced Fourier transform Raman scattering (FT-SERS) spectroscopy and compared it to those of the corresponding single-stranded poly[rA] and duplex poly[rA] · poly[rU]. Some interesting phenomena have been observed. At the ex situ electrochemically roughened silver electrode, the SERS behavior of the duplex RNA is close to that of the single-stranded poly[rA], thereby indicating that the duplex RNA adsorbed at the electrode might be partly destabilized. However, on the highly positively charged surfaces, the SERS spectra revealed that the triplex was predominantly adsorbed at the electrode via the phosphate-moiety-directed mechanism, and thus the helical structure of the triplex molecules was well preserved; furthermore, since the electrode potential was set to approach the potential of zero charge (pzc) of the silver metal, in the spectral region between 1800 and 600 cm−1 the triplex gives rise to SERS spectra similar to those of the corresponding duplex, and also yields only two enhanced signals at 734 and 1382 cm−1, due to the ring-breathing and ring-vibration modes of the adsorbed adenine residues, respectively, along with the disappearance of some bands originating from the corresponding rU residues and phosphate groups. In fact, a dramatic transition of the SERS spectra of the triplex at the electrode was found to occur between −0.2 and −0.4 V. On the basis of the structural characteristics of triple-helical nucleic acids, we concluded that in this case there are two types of competitive adsorbed species—the triplex itself and the unpaired adenine residues in the incomplete region of the triplex RNA—which might be responsible for the unique potential-dependent transition.

Structural characterization of an intramolecular RNA triple helix by NMR spectroscopy.

A chemically synthesized 29-base RNA oligomer, designed to fold to form an intramolecular triple helix at acid pH, has been studied by NMR spectroscopy. The molecule consisted of seven U.A.U or C+.G.C base triples joined by two pyrimidine tetra-loops. The fold was such that the third strand was Hoogsteen base-paired in the major groove of a Watson-Crick paired double helix. The nature and size of the molecule required the use of an assignment strategy using two- and three-dimensional homonuclear methods, complemented by a natural abundance 13C correlation experiment. The assignment of the majority of the exchangeable and non-exchangeable resonances is presented. The data suggest a C3'-endo sugar puckering for all the nucleotides involved in base triples. A preliminary structural model consistent with the NMR data is presented.

Symmetry and structure of RNA and DNA triple helices

Despite wide interest in nucleic acid triple helices, there has been no stereochemically satisfactory structure of an RNA triple helix in atomic detail. AN RNA triplex structure has previously been proposed based on fiber diffraction and molecular modeling [S. Arnott and P. J. Bond (1973) Nature New Biology, Vol. 244, pp. 99-101; S. Arnott, P. J. Bond, E. Selsing, and P. J. C. Smith (1976) Nucleic Acids Research, Vol. 3, pp. 2459-2470], but it has nonallowed close contacts at every triplet and is therefore not stereochemically acceptable. We propose here a new model for an RNA triple helix in which the three chains have identical backbone conformations and are symmetry related. There are no short contacts. The modeling employs a novel geometrical approach using the linked atom least squares [P. J. C. Smith and S. Arnott (1978) Acta Crystallographica, Vol. A34, pp. 3-11] program and is not based on energy minimization. In general, the method leads to a range of possible structures rather than a unique structure. In the present case, however, the constraints resulting from the introduction of a third strand limit the possible structures to a very small range of conformation space. This method was used previously to obtain a model for DNA triple helices [G. Raghunathan, H. T. Miles, and V. Sasisekharan (1993) Biochemistry, Vol. 32, pp. 455-462], subsequently confirmed by fiber-type x-ray diffraction of oligomeric crystals [K. Liu, H. T. Miles, K. D. Parris, and V. Sasisekharan (1994) Nature Structural Biology, Vol. 1, pp. 11-12]. The above triple helices have Watson-Crick-Hoogsteen [K. Hoogsteen (1963) Acta Crystallographica, Vol. 16, pp. 907-916] pairing of the three bases. The same modeling method was used to investigate the feasibility of three-dimensional structures based on the three possible alternative hydrogen-bonding schemes: Watson-Crick-reverse Hoogsteen, Donohue [J. Donohue (1953) Proceedings of the National Academy of Science USA, Vol. 39, pp. 470-475] (reverse Watson-Crick)-Hoogsteen, and Donohue-reverse Hoogsteen. We found that none of these can occur in either RNA or DNA helices because they give rise only to structures with prohibitively short contacts between backbone and base atoms in the same chain.