Density functional studies on the electron affinities of DNA and RNA bases

Influence of basis sets on electron affinities (EAs) of DNA and RNA bases has been investigated using density functional method (B3LYP functional) with different basis sets (6-31G, TZVP and 6-311+ + G**). Effect of some PBE functionals namely, PBEOP, PBELYP and PBEVWN, on EA values of the nucleobases was studied using basis set which predicted the most reliable values with B3LYP functional. Observation of the trends in the values of EA and dipole moment of the molecules enable us to identify the features of a basis set that shows the presence of dipole-bound state of some of the nucleobases. The vertical electron affinities with B3LYP and PBEOP functionals are close to the experimental values. The adiabatic electron affinities of uracil and thymine were found to be positive for basis set with diffuse functions using B3LYP functional. Adenine does not have a stable covalently bound anion at all levels of basis sets and functionals. The sign of adiabatic electron affinity value of cytosine is inconsistent with that of experimental value but in agreement with previous theoretical results. For guanine the adiabatic electron affinity value with 6-311+ + G** basis set was found to be very high as comparison with other two basis sets confirming the formation of mixed covalent-dipole character.     

An absolute method for the determination of the persistence length of native DNA from electron micrographs.

Information on spatial correlation in the tangent direction along electron microscope images of filamentous molecule is shown to be obtainable by the analysis of statistical fluctuations in curvature, yielding an absolute measure of the persistence parameter a_micro. The relationship of a_micro, a local, microscopic parameter, to the persistence length introduced by Kratky and Porod is discussed. The hypotheses underlying the assumed theoretical model concern (1) the shape of the angle distribution, assumed to be Gaussian; (2) the passage from a three- to a two-dimensional situation, which is supposed to occur by deformation of the flexible chain in a manner that preserves the memory of the spatial correlation in orientation (except for the blocking of one degree of freedom); and (3) the adsorption conditions, which should meet the equilibrium requirement as closely as possible. The analytical method has been checked on computer simulated “Gaussian” molecules: the study of the simulated sample was essential in solving the problems connected with minimum statistics requirements and the effect of the reading error. Experimental images obtained for T2 DNA fragments at different ionic strengths by Kleinschmidt's adsorption technique have been analyzed by means of an automatic flying spot digitizer, the “Precision Encoder and Pattern Recognition.” The results show that adsorbed molecules do in fact “remember” the rigidity they possessed in solution and that the Gaussian hypothesis is well verified. Consequently, the slopes of log cosθ (l) or θ² (l) may be used indifferently in the estimate of a_micro. The dependence of this parameter on ionic strength in the range explored shows the expected behavior.     

Influence of purines and pyrimidines on cold hardiness of plants. V. Uptake and translocation of purines and pyrimidines in alfalfa

Differential uptake of radioactive purines and pyrimidines by alfalfa leaves was observed. Translocation of radioactive constituents from leaves to stem-crown or root tissues (metabolic sinks) was not proportional to uptake. After 40 days of hardening at low temperatures, ranking of radioactivity levels changed for the base treatments and the change in leaves was different from that in whole plants. Ratios calculated for radioactivity from guanine and cytosine/adenine and uracil for RNA of leaves was twice as large as that for stem-crown or root tissues.More than $\text{3}\text{4}$ of the total radioactivity was found in soluble cytoplasm, although less than 10% of the RNA of hardened plants has been found in this fraction. This has special significance because soluble cytoplasm is the fraction that contains most of the polynucleotide phosphorylase activity and contains 60% of the soluble proteins that have been closely associated with cold tolerance.     

Amino acid catalyzed condensation of purines and pyrimidines with 2-deoxyribose.

Mild heating of aqueous mixtures containing 2-deoxyribose, amino compounds, and purines or pyrimidines produces derivatives of the purines and pyrimidines in high yield. Among the major products formed are 2,3-dideoxy-3-(1'-pyrimidino)pentose and 2,3-dideoxy-3-(9'-purino)pentose. The mechanism of the reaction includes amine-catalzyed dehydration of the alpha, beta positions of the sugar followed by addition of the purine or pyrimidine to the double bond. Rapid addition of purines and pyrimidines to alpha, beta-unsaturated carbonyl compounds (such as acrolin) is a general phenomenon which does not require an amine catalyst. While multiple derivatization of the purines will take place, the N-9 derivative is formed first.     

The sequential enzymatic determination of DNA and RNA

Enzymatically small quantities of native DNA and RNA in crude tissue homogenates can readily be specifically determined using ethidium bromide as a fluorescent indicator. The ethidium bromide-polynucleotide complexes of DNA and RNA serve as effective substrates for deoxyribonuclease and ribonuclease, respectively. Optimal divalent metal cation requirements were determined for a common reaction medium that is compatible for both the DNase and RNase reactions. To a single reaction mixture, that contains a biological sample, sequential addition of DNase and RNase produces a specific and rapid decrease in fluorescence that is proportional to the respective amounts of DNA and RNA present. Levels of DNA and RNA found in six different tissues of the rat were determined enzymatically by this method and compared to that obtained by alternate techniques. Enzymatically determined values were highly reproducible and correlate well with those values obtained by more time-consuming, conventional methods. Most enzymatically determined RNA levels in tissues, however, were significantly greater than those levels obtained spectrophotometrically. Advantages of the enzymatic procedure for analysis of tissue polynucleotide content are: (1) rapid determination of both DNA and RNA within a single sample aliquot allowing maximum use of available sample; (2) extensive fractionation and extraction of the tissue are not required; (3) it is especially useful when quantities of tissue are limited; and (4) sensitivity to 0.05 and 0.25 μg of DNA and RNA, respectively. Reaction conditions developed for the assay also provide for a highly sensitive means to continuously monitor DNA hydrolysis; DNase activity is directly proportional to the amount of enzyme added.