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A proflavine derivative with bulky substituents, 2,7-di-t-butyl-proflavine, binds to DNA but apparently not by intercalation. The methods used to study this problem include equilibrium dialysis, relaxation kinetics and hydroxdynamic studies on low-molecular-weight and closed-circular DNA. These show that binding produces no detectable lengthening or unwinding of the double helix, and that the binding reaction is much faster than is the case with unsubstituted proflavine. We conclude that an outside-bound complex is formed. We also measured equilibrium binding isotherms for DNA samples of varying G · C content, and for several synthetic polynucleotides. These results are consistent with a model in which a bound 2,7-di-t-butyl-proflavine molecule occupies three base pairs, and strong binding requires that two of these three be A · T pairs. Comparison of poly(dA) · poly(dT) with poly(A) · poly(U) shows a preference of at least 100-fold for binding to the former over the latter. Also, we found that, the thymine methyl groups are probably not responsible for the A · T specificity. The optical properties of the outside complex include a red shift of the absorbance maximum, with an increase in maximum absorbance instead of the more common decrease. The fluorescence of the dye is enhanced on binding, especially to an A · T-rich-DNA. The kinetic results show that two complex forms are present, differing in intrinsic affinity by about an order of magnitude. Both are formed with second-order rate constants in excess of 108 M-1 s-I and it is therefore very unlikely that either is an intercalated form. The absorbance spectrum of the bound dye changes as the degree of binding increases, due, at least in part, to the changing contribution of the two binding mechanisms. The conclusions concerning binding specificity refer to the dominant complex form; we have no firm information about the specificity of the weaker complex. [ABSTRACT FROM AUTHOR]

The interaction of hydroxymercurybenzene sulfonate (HOHgBzS) with 2-thiouridine (s2Urd) and 2-thiocytidine (s2Cyd) by means of absorption-difference spectroscopy and spectrophotometric titration was investigated. The absorption spectra of the reaction mixtures displayed isosbestic points indicating uniform reactions. Scatchard plots of the titration data revealed the formation of 1:1 complexes with a dissociation constant for s2Urd · HOHgBzS and s2Cyd · HOHgBzS of Kd = 0.55 μM and Kd = 0.54 ???M respectively. The interactions of poly(s2U) and poly(s2C) with hydroxymereurybenzene sulfonate were established by their corresponding absorption difference spectra. The binding of hydroxymercurybenzoato (HOHgBz) to poly(s2C) and poly(s2U) was investigated by means of equilibrium dialysis. The association of HOHgBz to 2-thiocytosine bases in poly(s2C) follows a simple hyperbolic binding isotherm. The Scatchard plot of the binding data disclosed that only a fraction (n = 0.32) of the s2CMP residues in poly(s2C) was accessible to the interaction with HOHgBz. The dissociation constant of this interaction was found to be Kd = 4.3 ???M. The binding of HOHgBz to the helical poly(s2U) is a cooperative process as indicated by the sigmoidal binding cuve. Under saturating conditions all s2UMP residues in poly(s2U) interact with the ligand. [ABSTRACT FROM AUTHOR]

The soluble molecular complexes of histone-DNA in solutions of 0.3–0.5 M NaClO4 at pH 5.6 have been studied by means of sedimentation and spectrophotometric analysis. The binding isotherms of DNA with two histones, the lysine-rich fraction F1 (from calf-thymus) and the fraction V, specific of erythrocytes, appear as straight lines in the classical inverse plot. A common intercept of 250 is observed and the slopes vary strongly with ionic-strength. From a double logarithmic plot of these sloes versus ionic-strength, the number of binding sites α is estimated to be 7. Such a number of adjacent basic residues could be found in the primary structure of F1 and FV or could be caused by the folding of the molecule. One can propose two types of interpretation for the value of the intercept. If non specific binding is assumed, the number of covered phosphate per histone molecule β would be close to 170, giving an average distance of 285 Å between two successive bound proteins. This value is not compatible with the known dimensions of the protein and negates a simple steric hindrance model. Assuming specific binding, the intercept of this isotherms then corresponds to 4 specific areas per 1000 nucleotides and a part of the binding energy is non-coulombian. However, if heterologous instead of homologous components are used, no specificity can be detected. The effect of temperature upon binding constant was studied between 2 and 25 °C. The linear plot of log K versus 1⁄T enables us to determine αΔH of 42 kJ (10 kcal/mol)/mol and yields the high ΔS value of about 50 e.u./mole. This last value could be related to modification of histone structure or more likely to changes of hydratation accompanying the binding process. [ABSTRACT FROM AUTHOR]

To study the binding of proteins to DNA, it was found useful to develop a general method of interpretation of binding isotherms, which appears as an extension of previous calculations made by Latt and Sober. It is assumed that electrostatic site-binding only occurs between adjacent phosphate groups and basic residues of the protein. For each molecule of protein a number a of electrostatic bonds are made and m sequences of a sites are available on the DNA. When all the sites are available (ma equal to the total number of nucleotides) isotherms have to be calculated by means of a computer If K is the intrinsic binding constant, A0 the concentration of the free protein and &vmacr; the average number of race bound to DNA, a plot of 1⁄&vmacr; versus 1⁄KA0 is found to be practically linear in a large range of KA0 values. The easiest way to interprete the linear part of the isotherms quantitatively is to recalculate it by using the Lifson's method of sequence generating function. One gets the simple equation 1⁄r = α + γ + (1⁄KA0) where r is the amount of bound protein per nucleotide and γ a parameter determined graphically which depends only on α, and on range of KA0. Values of α and K are thus readily obtained. The number of rites involved in the binding may be smaller than the total number of phosphate groups in the DNA, either by steric hindrance of the protein and hence masking of a certain number of phosphate groups or by the existence of specific sequences of nucleotides. In the first case, if β is the number of rites covered by the protein, equation, of the linear part of the isotherms is now 1⁄r = β + γ + (1⁄KA0) and only β and K can readily be measured. It is however possible to determine a by means of competition experiment between proteins and counter-ions for which all the rites are assumed to be available. If K′ is now the apparent association constant, the change of K′ with ionic strength is represented by log K′ = a log K0 + α log [kl⁄(1 + k1c1)] where k1 and c1 are respectively the association constant with DNA and the concentration of counter-ions and K0 a parameter taking into account the non coulombian binding energy between basic groups of the proteins and phosphate groups. The plot of log K′ versus log [kl⁄(1 + k1c1)] enables the determination of K0. In the second case m and K are readily obtained from isotherms by using the classical theory of multiple equilibria and α is determined according to the method previously described. Such a method was tested on oligolysine—polyribonucleotide interactions where calculated values of α are in agreement with those expected. The method was also applied to the binding of repressor to DNA for which only three phosphate groups seem to be involved in the process. Electrostatic energy, which was predominant in the former case, is only a small part of the total energy of binding in the latter. [ABSTRACT FROM AUTHOR]

Studies the effects of light, temperature and daylength on the form, growth and net assimilation rate of the S.170 variety of tall fescue from the Great Britain and Syn I and Syn II varieties from North Africa as of 1968. Reason for small relative growth rate of S.170 in winter; Factors influencing rate of leaf growth of S.170; Implications of shading for relative growth rate and net assimilation rate of the varieties.

Presents ecological studies on the large copper butterfly Lycaena dispar (L.d.) batavus at Woodwalton Fen National Nature Reserve in Huntingdonshire, England as of 1968. Race of L.d. Haw. most widespread in Europe; Native habitat of L.d. batavus and L.d. dispar; Relationship between egg-laying by copper butterfly and air temperature; Factors influencing seasonal mortality rates of the butterflies.

Studies the expansion of leaf area per plant in field bean as related to daily maximum air temperature as of 1968. Effect of leaf area index on crop yields; Factors influencing leaf area; Impact of temperature on the rate of canopy development of plants; Decrease in time taken for leaf area index to expand from 0.1 to 2.0 per degree celsius increase of mean monthly maximum temperature.

Partition isotherms of two kinds of transfer ribonucleic (tRNA) acids from baker's yeast and calf liver and six yeast tRNAs have been determined at four temperatures (0, 10, 20, and 30°), at three pH (6.6, 7.0, and 8.0) in the following solvent system: potassium phosphate buffer 1.5 M—2-methoxyethanol—2-butoxyethanol (300:100:30–130, by vol.). The effect of various parameters on the partition coefficient of tRNA, such as molarity and pH of the buffer, temperature, concentration of the organic components, have been investigated. The semi-logarithmic variation law of the partition coefficient is expressed with respect to the temperature and the 2-butoxyethanol content of the solvent mixture. Numerical values of some solvent system constants have been calculated. Advantages of this system for separation and enrichment of specific tRNAs have been discussed for a further application to a counter-current distribution or a column partition chromatography using a decreasing 2-butoxyethanol or temperature gradient. [ABSTRACT FROM AUTHOR]