Poly rU Research Articles

Melting studies on spin‐labeled polyadenylic–polyuridylic complexes

AbstractTemperature‐dependent conformational transitions of spin‐labeled poly rA, spin‐labeled poly rU and the two‐stranded helical complexes consisting either of spin‐labeled rA·poly rU or spin‐labeled poly rU·poly rA have been measured by electron spin resonance spectrocopy. The polynucleotides were spin labeled with 4‐(2‐iodoacetamido)2,2,6,6‐tetramethylpiperidinooxyl and the spin label to nucleotide base ratio was approximately 1:600. The relationship between the log of tumbling time τ and the reciprocal absolute temperature for the spin‐labeled single and double‐stranded polynucleotides is presented. An agreement between TmOD (optical density melting) and Tmsp (spin melting) is found for the complexes, which strongly supports the conclusion that the same temperature‐dependent structural changes are monitored with both techniques.

Studies on spin-labeled polyriboadenylic acid.

AbstractSpin‐labeled samples of poly rA, poly rU, and poly rG have been prepared, and physicochemical properties primarily of labeled poly rA are reported. The nitroxide radical, 4‐(2‐iodoacetamido)‐2,2,6,6‐tetramethylpiperidinooxyl, is incorporated to a greater extent in poly rA and poly rU, as compared to poly rG. No incorporation is observed in the case of poly rC. Special attention has been paid to the separation of the covalently attached labels from the free labels, and to the preservation of the integrity of the chain length of the labeled polymers. The determination of molar extinction coefficients of the three labeled polymers indicates virtually no difference from those known for the chemically unpertubed polyribonucleotides. The correlation times for the spin‐labeled single stranded poly rA and poly rU have been calculated. More mobile building blocks are found in poly rU as compared to poly rA. Conformational properties of labeled poly rA in aqueous solutions have been investigated using electron spin resonance, circular dichroism, and absorption spectroscopy. The objective of the study of labeled poly rA was to examine its conformational transitions upon the uptake of protons by the adenine bases. Based on electron spin resonance data there is strong evidence that the single strand‐double strand transition can take place in three steps. In addition to the already known two forms of double‐stranded poly rA in acidic solution, called A and B, it is suggested that a third phase, consisting possibly of large aggregates, is involved in the transition of the less protonated double strands to those of complete protonation.

Studies on the antiviral activity and cell interaction of synthetic double-stranded polyribo- and polydeoxyribonucleotides

A variety of (radiolabeled) RNA and DNA polymers, including (poly rA)·(poly rU), poly r(A-U), (poly rI)·(poly rC), poly r(I-C), (poly dA)·(poly dT), poly d(A-T), (poly dI)·(poly dC), and poly d(I-C), were compared in their antiviral activity as measured in terms of inhibition of virus plaque formation by vesicular stomatitis virus in human skin fibroblasts. These polymers were also studied with respect to the kinetics of their binding to cells and with respect to their persistence at the outer cell membrane. The kinetics of cell binding was monitored by the loss of acid-insoluble radioactivity from the supernatant fluid; persistence of the polymer at the cell surface was determined by measuring the amount of cell-bound radioactivity that was released upon nuclease treatment. Although preincubation of the polymer at 37° increased the antiviral activity and rate of cell binding, and persistence at the cell surface of each individual polymer, there was no direct correlation between antiviral activity, rate of cell binding, and persistence at the cell surface with all polymers studied. These findings suggest that the differences in antiviral activity among polyribo- and polydeoxyribonucleotides are not related to differences in kinetics of cell binding or persistence of polymer at the cell surface, but reflect an as yet undefined interaction of the polymer with the cell.

Poly 2′-deoxy-2′-aminouridylic acid

The synthesis of poly 2′-deoxy-2′-aminouridylic acid is described. This analog of poly rU does not seem to form a double stranded complex with the complementary poly rA under normal conditions.

The conformation of single-strand polynucleotides in solution: sedimentation studies of apurinic acid.

AbstractTo elucidate the role of the bases in single‐strand polynucleolide conformation, we have studied apurinic acid, a single‐strand polydeoxyribonucleotide from which almost half the bases have been removed. The conformation of apurinic acid in aqueous solution near θ condition has been investigated by sedimentation velocity and sedimentation equilibrium measurements. The unperturbed coil dimensions of apurinic acid are essentially identical to those of poly rU of the same degree of polymerization. The dimensions are also similar to those of poly rA at high temperature, where the adenine residues are not stacked upon one another. We conclude that the considerable rigidity of these polynuclotides is conferred not by residual, undetected base stacking, but by restrictions in rotation about the bonds of the backbone. Furthermore, the rigidity of the ribose‐phosphate backbone cannot be attributed to interactions involving the 2'—OH group.

Conformation of polyribouridylic acid in solution

The conformation of polyribouridylic acid in aqueous solution has been studied under ideal solvent conditions, using light scattering, viscometry and sedimentation velocity and equilibrium measurements. The unperturbed dimensions of the poly rU molecule do not vary greatly with temperature, as expected for a polynucleotide with little or no base stacking. Despite the absence of stacking, the characteristic ratio < h 2 > nl 2 is 17.6, corresponding to a quite highly extended random coil. Only slightly smaller values are found by extrapolation of the data for poly rA to the completely unstacked form, suggesting that the degree of extension of an unstacked polynucleotide does not depend upon the nature of the base. This degree of extension can only be accounted for if rotation about all of the bonds of the polynucleotide backbone is highly restricted, as has been suggested recently by Sundaralingam (1969) on the basis of his survey of the crystallographic literature. The restrictions he proposes appear to be sufficient to account for our result.

Topography of nucleic acid helices in solutions. The interaction specificities of optically active amino acid derivatives

The synthesis and interactions of the d- and l-enantiomers of the amino acid amide derivatives [Formula: see text] (I) and lysyl dipeptides [Formula: see text] (II) with poly rI.poly rC, poly rA.poly rU and calf thymus DNA is reported. The following results were found. (1) The degree of stabilization of the helices as measured by the T(m) (;melting' temperature) of the helix-coil transition was dependent on the nature of the amino acid. (2) For the poly rI.poly rC helix, the l-enantiomers of salts (I) and (II) stabilized more than the d-enantiomers. The same was true for calf thymus DNA in the presence of salts (II) and for poly rA.poly rU in the presence of salts (II) and the proline derivatives of salts (I). (3) As R increased in size and became more apolar, the amount of stabilization of the poly rI.poly rC helix in the presence of salts (I) decreased. On the other hand, the amount of stabilization increased with more polar substituents. An attempt was then made to determine whether the difference in stabilization of the double-stranded helices at the T(m) in the presence of the l- and d-enantiomers of salts (I) is due to the interaction with the helix, the random coil or both. A new method was developed for determining the binding of small ions to polyions that involves a competition between an insoluble polystyrene ion-exchange resin and the soluble polyion for the counterion. Dissociation constants are obtained for the complexes of single- and double-stranded helices with the salts (I). The results are illuminating and indicate that with certain helices, i.e. poly rA.poly rU, the interactions of salts (I) with the single strands may not be ignored. It is concluded that the high optical specificity found in Nature, i.e. d-ribose in nucleic acids and l-amino acids in proteins, cannot be attributed solely to monomer-polymer interactions described by Gabbay (1968).