Spectra polynucleotides

Infrared and Raman spectroscopy are in current use fo r elucidating the molecular structures of nucleic acids. The application of infrared spectroscopy to studies of the structure of nucleic acids has been reviewed,135 as well as of Raman spectroscopy.136 It was noted that the assignments are generally based on isotopic substitution, or on comparison of the spectrum of simple molecules that are considered to form a part of the polynucleotide chain to that of the nucleic acid. The vibrational spectra are generally believed to be a good complementary technique in the study of chemical reactions, as in the study76 of carbohydrate complexation with boric acid. In this study, the i.r. data demonstrated that only ribose forms a solid complex with undissociated H3B03, and that the complexes are polymeric. 

Thermodynamic and kinetic studies have demonstrated that the alternating purine-pyrimidine polynucleotide poly(dA-dT) folds into smaller duplexes which melt independently of each other in solution (16,17). The NMR spectrum should be considerably simplified since all base pairs are structurally equivalent due to the symmetry of the alternating purine-pyrimidine duplex. 

The deprotected oligonucleotide synthetic product is precipitated twice in ethanol, and a 0.5 fig/fd solution in water is prepared (concentration is measured from a UV absorption spectrum). One microliter of the oligo-deoxynucleotide solution is mixed with 2 fd of 10X PL, 5 fd of [y-32P]ATP (or [y-35S]ATP), 1 fd of T4 polynucleotide kinase, and 11 fd water. After incubation at 37 ° (for 45 min with [y-32P]ATP or for 2 hr with [y-35S]ATP), the reaction is stopped by the addition of 150 [A of 5 M ammonium acetate, pH 5.5, and 130 fd water and 10 fd of the yeast tRNA solution are added to the mixture before precipitation with 1 ml ethanol. After chilling at —70° for at least 15 min, the precipitate is collected by centrifugation (12,000 g, 15 min), redissolved, and submitted to two additional cycles of precipitation-redissolution. Finally, the precipitate is redissolved in 20 fd of gel loading mix and the mixture analyzed on a 8% acrylamide-7 Af urea slab gel in IX electrophoresis buffer, until the bromphenol blue has reached the middle of the gel. 

While the importance of IR and Raman spectroscopy for the structural elucidation of purine bases has diminished over the last few decades, its advantages for the study of oligo- and polynucleotides in particular with respect to base pairing is apparent. The IR absorption spectrum of a single-stranded polynucleotide is very similar to that of its component nucleotides, but drastic changes occur on formation of hydrogen-bonded helical structures. 

To illustrate the versatility of the metal complexes of polynucleotides, it has recently been shown that zinc (II) can bind to the phosphate groups of the macromolecules to which bases silver (I) has already been attached. Evidence for this simultaneous binding is provided by the fact that the characteristic effect of silver (I) on the spectrum of the polynucleo-... 

Figures 15 and 16 show typical Py-El mass spectra of herring-sperm DNA and yeast RNA recorded on a Riber 1010 quadrupole mass spectrometer. The spectrum of the synthetic polynucleotide polydeoxyinosine-deoxycytosine (Fig. 17), recorded in a negative ion mode e capture), shows the essential characteristics of this polymer (jn/z 13 5 B for I and 110 B for C) as expected.

An increase in the contribution of transients involving the bases to the DNA transient spectrum as the pH is increased is consistent with chemical evidence that bases in the double helix of polynucleotides are protected from radiation to a degree which is related to helix stability. Radiation sensitivity of the pentose on the other hand, is unrelated to macromolecular structure (16). 

The natural geometry of biomolecules can be exploited to clarify the SERS sensitivity dependence on distance from the surface. This aspect of SERS spectroscopy is important for the interpretation of the SERS spectrum of a biopolymer. The electromagnetic model predicts very rapid decay of SERS with increasing distance Thus, in small molecules with a dimension of approximately 0.6 nm (benzene) all vibrations of the molecule can be enhanced. In large biomolecules with diameters of about 6 nm (hemoglobin protein) only groups which are attached directly to the surface will yield SERS. This important aspect is illustrated by SERS studies of three examples mono-, di-, and polynucleotides. 

Hilleman and his co-workers have demonstrated interferon production following the administration of reovirus RNA or the replicative form of RNA isolated from E. coli infected with MS2 coliphage. These BNAs are effective in inducing resistance to virus infection m vitro and vivo. The induction of interferon and the broad-spectrum protection against viral infection conferred by double-stranded RNA suggested the use of synthetic polynucleotides, including polyriboinosinic-polyribocytidilic acid, which possess a double-stranded conformation. Chemically modified RNA may also induce the production of interferon. ... 

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