Mg2+ dependence of the structure and thermodynamics of wheat germ and lupin seeds 5S rRNA.

The formation and stability of structural elements in two 5S rRNA molecules from wheat germ (WG) and lupin seeds (LS) as a function of Mg2+ concentration in solution was determined using the adiabatic differential scanning microcalorimetry (DSC). The experimentally determined thermodynamic parameters are compared with calculations using thermodynamic databases used for prediction of RNA structure. The 5S rRNA molecules which show minor differences in the nucleotide sequence display very different thermal unfolding profiles (DSC profiles). Numerical deconvolution of DSC profiles provided information about structural transformations that take place in both 5S rRNA molecules. A comparative analysis of DSC data and the theoretical thermodynamic models of the structure was used to establish a relationship between the constituting transitions found in the melting profiles and the unfolding of structural domains of the 5S rRNA and stability of its particular helical elements. Increased concentrations of Mg2+ ions induces additional internal interactions stabilising 5S rRNA structures found at low Na+ concentrations. Observed conformational transitions suggest a structural model in which the extension of helical region E dominates over the postulated tertiary interaction between hairpin loops. We propose that helix E is stabilised by a sequence of non-standard pairings extending this helix by the formation of tetra loop e and an almost total reduction of loop d between helices E and D. Two hairpin structures in both 5S rRNA molecules: the extended C-C' and the extended E-E'-E" hairpins appear as the most stable elements of the structure. The cooperativity of the unfolding of helixes in these 5S rRNA molecules changes already at 2 mM Mg2+.

Crystal structure of 2'-deoxycytidine hemidihydrogenphosphate reveals C+.C base pairs and tight, hydrogen-bonded (H2PO4-)infinity columns (1).

2'-Deoxycytidine hemidihydrogenphosphate has been crystallized in the hexagonal space group P6(2) with a = 25.839(3), c = 12.529(1) A. The structure has been solved using the Patterson search method. The asymmetric unit contains two protonated, base-paired 2'-deoxycytidine dimers and two H2PO4- anions. The C+.C base pairs are composed of a protonated and a neutral species each and are triple H-bonded, the central N(3) ... N(3) bonds being 2.850(7) and 2.884(5) A. The conformations of the four nucleosides fall in the same category (sugar puckers 2'-endo, glycosidic links anti) but in one of them the glycosidic torsion angle is quite low with consequences in other geometrical parameters. The H2PO4- anions are located on twofold axes and form two types of tight columns with P ... P separations about 4.18 A. The neighboring units along a column are linked via two very short O ... H ... O hydrogen bonds (O ... O about 2.49 A) leading to effective equalization of the P-O bonds. The base pairs of the two dC+.dC cations are coplanar and form layers perpendicular to the phosphate columns repeating every c/3. Within the layers, the dimers form a network through O(5') ... O(2) hydrogen bonds but their primary intermolecular interactions have the form of H-bond anchors [N(4)-H ... O-P and O(3')-H ... O-P] to the phosphate groups.

Comparative calorimetric studies on the dynamic conformation of plant 5S rRNA: II. Structural interpretation of the thermal unfolding patterns for lupin seeds and wheat germ.

Thermal unfolding of 5S rRNA from wheat germ (WG) and lupin seeds (LS) was studied in solution. Experimental curves of differential scanning calorimetry (DSC) were resolved into particular components according to the thermodynamic model of two-state transitions. The DSC temperature profiles for WG and LS differ significantly in spite of very high similarities in the sequence of both molecules. Those results are interpreted according to a model of the secondary and tertiary molecular structure of 5S rRNA. A comparison of the 'nearest neighbour' model of interaction with the experimental thermodynamic results enables a complete interpretation of the process of the melting of its structures. In light of our observations, the crucial differences between both DSC melting profiles are mainly an outcome of different thermodynamic properties of the first helical fragment 'A' made up of 9 complementary base pairs. It contains 6 differences in the nucleotide sequence of both types of molecules, which still retain 9-meric double helixes. The temperature stability of his helix in WG is much lower than of the LS one. Moreover, the results supply evidence for a strong specific tertiary interaction between the two hairpin loops 'c' and 'e' in both 5S rRNA molecules, modulated by small differences in the thermodynamic properties of both 5S rRNA.

Comparative calorimetric studies on the dynamic conformation of plant 5S rRNA. I. Thermal unfolding pattern of lupin seeds and wheat germ 5S rRNAs, also in the presence of magnesium and sperminium cations.

An attempt has been made to correlate differential scanning calorimetry melting profiles of 5S rRNAs from lupin seeds (L.s.) and wheat germ (W.g.) with their structure. It is suggested that the observed differences in thermal unfolding are due to differences in RNA nucleotide sequence and as a consequence in higher order structures. Interesting effects induced by magnesium cation, perprotonated and permethylated sperminium tetracations are discussed. It is suggested that the difference in the stabilizing effect of the three cations results from different mode of their interactions with RNA. "Pure" electrostatic interactions expected for permethylated tetracations are rather weak due to the steric hindrance around each positively charged nitrogen atom. Electrostatic interactions of the other two cations are significantly enhanced by coordination bonding for magnesium and by hydrogen bonding for protonated sperminium cation.

Comparative structural analysis of cytidine, ethenocytidine and their protonated salts III. 1H, 13C and 15N NMR studies at natural isotope abundance.

The 1H, 13C, 15N NMR spectra of cytidine /Cyd/, ethenocytidine /epsilon Cyd/ and their hydrochlorides /Cyd X HC1/ and /epsilon Cyd X HC1/ have been analysed to compare structural differences observed in solution with those existing in the crystalline state. The effects of ethenobridging and protonation of the hertero-aromatic base on the intramolecular stereochemistry, intermolecular interactions and electronic structure of the whole molecule are discussed on the basis of the NMR studies in DMSO solutions. Particular interest is devoted to the discussion of the conformation of the ribose ring, the presence of the intramolecular C-5'-0...H-6-C hydrogen bond, unambiguous assignment of the site of protonation, the mechanism of the 5C-H deuterium exchange in Cyd X HC1, and the intermolecular interactions in solution.