Conformational diversity versus nucleic acid triplex stability, a combinatorial study.

The stability of a triple helix formed between a DNA duplex and an incoming oligonucleotide strand strongly depends on the solvent conditions and on intrinsic chemical and conformational factors. Attempts to increase triple helix stability in the past included chemical modification of the backbone, sugar ring, and bases in the third strand. However, the predictive power of such modifications is still rather poor. We therefore developed a method that allows for rapid screening of conformationally diverse third strand oligonucleotides for triplex stability in the parallel pairing motif to a given DNA double helix sequence. Combinatorial libraries of oligonucleotides of the requisite (fixed) base composition and length that vary in their sugar unit (ribose or deoxyribose) at each position were generated. After affinity chromatography against their corresponding immobilized DNA target duplex, utilizing a temperature gradient as the selection criterion, the oligonucleotides forming the most stable triple helices were selected and characterized by physicochemical methods. Thus, a series of oligonucleotides were identified that allowed us to define basic rules for triple helix stability in this conformationally diverse system. It was found that ribocytidines in the third strand increase triplex stability relative to deoxyribocytidines independently of the neighboring bases and position along the strand. However, remarkable sequence-dependent differences in stability were found for (deoxy)thymidines and uridines.     

Conformational constraints of amino acid side chains in alpha-helices

The conformational freedom of amino acid side chains is strongly reduced when the side chains occur on an α-helix. A quantitative evaluation of this freedom has been carried out by means of conformational energy computations for all naturally occurring amino acids and for α-aminobutyric acid when they are placed in the middle of a right-handed poly(L-alanine) α-helix. One of the three possible rotameric states for rotation around the C^α ? C^β bond (viz. g⁺) is excluded completely on the helix because of steric hindrance, and the relative populations of the other two rotamers (t and g^?) are altered because of steric interactions and the reduction of hydrogen-bonding possibilities. The computed tendencies of the changes in distributions of rotamers, on going from an ensemble of all backbone conformations to the α-helix, agree with the observed tendencies in proteins. Minimum-energy side-chain conformations in an α-helix have been tabulated for use in conformational energy computations on polypeptides.     

Conformational preferences of amino acid side chains in collagen

Conformational energy computations were carried out on collagenlike triple-stranded conformations of several poly(tripeptide)s with the general structure CH₃CO? (Gly? X? Y)₃? NHCH₃. The sequences considered had various amino acid residues in position X or Y of the central tripeptide, with either Pro or Ala as a neighbor, i.e., Gly-X-Pro, Gly-X-Ala, Gly-Pro-Y, and Gly-Ala-Y. Minimum-energy conformations were computed for the side chains, and their distributions were compared for the four sequences. The residues used were Abu (= α-aminobutyric acid), Leu, Phe, Ser, Asp, Asn, Val, Ile, and Thr. The conformational energy of a ? Ch₂? CH₃ side chain in Abu was mapped as a function of the dihedral angle χ¹. Intrastrand interactions with neighboring residues do not affect the conformations of a side chain in position Y, and they have a minor effect on it in the X-Ala sequence, but they strongly restrict the conformational freedom of the side chain in the X-Pro sequence. Conversely, interstrand interactions do not affect side chains in position X, but they strongly restrict the conformational freedom of a side chain in position Y if there is a nearby Pro residue in a neighboring strand. Hydrogen bonds with the backbone can be formed in some conformations of long polar side chains, such as Asp, Asn, or Gln. All amino acid residues can be accommodated in collagen. Because of the interactions mentioned above, steric and energetic constraints can be correlated with observed preferences of certain amino acids for positions X or Y in collagen. Hence, these preferences may be explained, in part, in terms of differences in the conformational freedom of the side chains in the triple-stranded structure.     

Raman spectral studies of nucleic acids. XVII. Conformational structures of polyinosinic acid.

Laser-Raman spectra of poly(rI) show the formation of an ordered complex in aqueous solutions of high ionic strength. This structure exhibits the A-helix geometry, contains stacked bases and is apparently stabilized by specific hydrogen bonding involving hypoxanthine C6=0 groups. Thermal dissociation of the poly(rI) complex (Tm=45 degrees C) yields single-stranded and disordered poly (RI) chains. A disordered structure also occurs for poly (rI) in aqueous solutions of low ionic strength. In oriented films, poly (rI) forms an ordered structure probably the same as that which occurs in solutions of high ionic strength. Raman intensities measured at 815 and 1100 cm-1 in spectra of poly (rI) and poly (rU)-poly (rA)-poly(rU) indicate that the correlation previously established for single- and double-stranded ribopolymer structures is valid also for these multi-stranded structures. X-ray diffraction and model-building studies confirm the A-helix structure.     

Conformational studies of nucleic acids: III. Empirical multiple correlation functions for nucleic acid torsion angles

There are seven significantly variable torsion angles in each monomer unit of a polynucleotide. Because of this, it is computationally infeasible to consider the energetics of all conformations available to a nucleic acid without the use of simplifications. In this paper, we develop functions suggested by and regression fit to crystallographic data which allow three of these torsion angles, alpha (O3'-P-O5'-C5'), delta (C5'-C4'-C3'-O3') and epsilon (C4'-C3'-O3'-P), to be calculated as dependent variables of those remaining. Using these functions, the seven independent torsions are reduced to four, a reduction in complexity sufficient to allow an examination of the global conformational energetics of a nucleic acid for the remaining independent torsion angles. These functions are the first to quantitatively relate a dependent nucleic acid torsion angle to several different independent angles. In all three cases the data are fit reasonably well, and in one case, alpha, the fit is exceptionally good, lending support for the suitability of the functions in conformational searches. In addition, an examination of the most significant terms in each of the correlation functions allows insight into the physical basis for the correlations.     

Conformational transitions in human translin enable nucleic acid binding

Translin is a highly conserved RNA- and DNA-binding protein that plays essential roles in eukaryotic cells. Human translin functions as an octamer, but in the octameric crystallographic structure, the residues responsible for nucleic acid binding are not accessible. Moreover, electron microscopy data reveal very different octameric configurations. Consequently, the functional assembly and the mechanism of nucleic acid binding by the protein remain unclear. Here, we present an integrative study combining small-angle X-ray scattering (SAXS), site-directed mutagenesis, biochemical analysis and computational techniques to address these questions. Our data indicate a significant conformational heterogeneity for translin in solution, formed by a lesser-populated compact octameric state resembling the previously solved X-ray structure, and a highly populated open octameric state that had not been previously identified. On the other hand, our SAXS data and computational analyses of translin in complex with the RNA oligonucleotide (GU)₁₂ show that the internal cavity found in the octameric assemblies can accommodate different nucleic acid conformations. According to this model, the nucleic acid binding residues become accessible for binding, which facilitates the entrance of the nucleic acids into the cavity. Our data thus provide a structural basis for the functions that translin performs in RNA metabolism and transport.     

Conformational classification of functional nucleic acids.

The kink parameters would provide the tolerant aspect for irregular helical structure of nucleic acid. Using these kink parameters, the classification of conformation space was carried for the functional nucleic acid molecules. The kink parameters could afford us the simple structural aspects about the constructive parts of functional molecules. Local elastic kink phenomena can be classified by rod like models with the combination of kink parameters. The constructive parts, such as the stable tetra nucleotides loop, U-turn conformation and adenosine platform, were selected and the statistical analyses were carried on the parameters calculated by program BIOCON.     

Metal ion-mediated specific interactions between nucleic acid bases of polynucleotides and amino acid side chains of polypeptides.

Interactions between copolypeptides containing Glu and Tyr residues and polynucleotides can be mediated through divalent metal ions such as Zn-2+ and Ci-2+. Circular dichroism studies show that the binding of metal ion - polypeptide complexes to poly(A) induces an unstacking of adenine bases. Fluorescence investigations demonstrate that Tyrosine - Adenine interactions result from the formation of ternary complexes polypeptide-Zn-2 plus-polynucleotide.     

Torsion angle approach to nucleic acid distance geometry: TANDY

Summary An efficient algorithm for generating DNA structures from a given set of distance constraints has been developed. The present implementation is suited for single-stranded DNA. The performance of the program has been tested with constraint sets representative of most stringent theoretical cases and also with usually available experimental ones. The results indicate that use of NOE-derived constraints alone generates an extremely large family of conformers and suggest that the quality of structure determination may be enhanced by incorporating additional constraints obtained by other means. The speed of the program makes it ideal for interactive use in conjunction with other complementary algorithms such as those for spectral simulation, energy minimization and molecular dynamics calculations.Dedicated to the memory of Professor V.F. Bystrov     

Theoretical studies of nucleic acid interactions. I. Estimates of conformational mobility in intercalated chains

A combined geometric and potential-energy analysis has been carried out to identify the torsional arrangements of the nucleic acid chain that can accommodate the intercalation of small planar moieties. In contrast to previous theoretical efforts, which detail local conformations after adjacent bases are positioned in space, the likely geometries are found here on the basis of the base orientations that result from all feasible combinations of the nine torsional variables of the basic dinucleotide intercalation unit. The relatively mobile nature of the sugar-phosphate backbone, together with the fairly long stretches of chemical bonds between adjacent units, is apparently responsible for the large number of feasible binding geometries. Some previously overlooked conformations with unusual sugar-puckering combinations and various phosphodiester arrangements are found in the survey. A large proportion of the energetically favored intercalation states are closely related to the backbone conformations of familiar double-helical models such as A-, B-, and Z-DNA, as well as the Watson-Crick model. Moreover, the intercalated forms are found to interconvert smoothly along a continuous conformational pathway. The intercalation structures derived from x-ray crystallographic analyses of drug-oligonucleotide complexes, in contrast, are stiff three-dimensional forms essentially frozen in a single domain of conformation space. Specific ligand-nucleic acid interactions that may be responsible for the experimental observations are not included in this study. The classical intramolecular potential energies reported here are highly approximate, providing only rough gauges of the relative importance of the many competing conformations.     

Conformational changes induced in tobacco mosaic virus nucleic acid by ultraviolet radiation

Summary In order to study any Conformational changes associated with ultraviolet irradiation of TMV-RNA, methods of circular dichroism and absorbance-temperature profile were applied. RNA irradiated in water at 254 nm showed a distinct change in CD spectrum, but there was only a slight change accompanying irradiation in phosphate buffer.A small change in absorbance-temperature profile at 258 nm is associated with irradiation of RNA in water; RNA irradiated in phosphate buffer and Mg⁺⁺ solutions showed essentially no changes.It is concluded that conformational changes accompanying irradiation of RNA in water are greater than those taking place in phosphate or magnesium solutions, and the enhanced change in water is related to the larger quantum yield for inactivation found in water than in buffer solutions.Photochemistry of Macromolecules XXXIII, supported in part by the U.S. Atomic Energy Commission, contract AT(11-1)-34, Project 116.     

Conformational characteristics and correlations in crystal structures of nucleic acid oligonucleotides: evidence for sub-states

Sugar phosphate backbone conformations are a structural element inextricably involved in a complete understanding of specific recognition nucleic acid ligand interactions, from early stage discrimination of the correct target to complexation per se, including any structural adaptation on binding. The collective results of high resolution DNA, RNA and protein/DNA crystal structures provide an opportunity for an improved and enhanced statistical analysis of standard and unusual sugar-phosphate backbone conformations together with corresponding dinucleotide sequence effects as a basis for further exploration of conformational effects on binding. In this study, we have analyzed the conformations of all relevant crystal structures in the nucleic acids data base, determined the frequency distribution of all possible epsilon, zeta, alpha, beta and gamma backbone angle arrangements within four nucleic acid categories (A-RNA and A-DNA, free and bound B-DNA) and explored the relationships between backbone angles, sugar puckers and selected helical parameters. The trends in the correlations are found to be similar regardless of the nucleic acid category. It is interesting that specific structural effects exhibited by the different unusual backbone sub-states are in some cases contravariant. Certain alpha/gamma changes are accompanied by C3' endo (north) sugars, small twist angles and positive values of base pair roll, and favor a displacement of nucleotide bases towards the minor groove compared to that of canonical B form structures. Unusual epsilon/zeta combinations occur with C2' (south) sugars, high twist angles, negative values of base pair roll, and base displacements towards the major groove. Furthermore, any unusual backbone correlates with a reduced dispersion of equilibrium structural parameters of the whole double helix, as evidenced by the reduced standard deviations of almost all conformational parameters. Finally, a strong sequence effect is displayed in the free oligomers, but reduced somewhat in the ligand bound forms. The most variable steps are GpA and CpA, and, to a lesser extent, their partners TpC and TpG. The results provide a basis for considering if the variable and non-variable steps within a biological active sequence precisely determine morphological structural features as the curvature direction, the groove depth, and the accessibility of base pair for non covalent associations.