Stoichiometry of interaction

Expression of a recombinant protein using an inducible vector system would permit expression at endogenous levels to simulate physiologic levels of expression of a protein of interest. Tandem affinity purification strategies have recently been employed and facilitate the analyses of highly interactive proteins when the bait protein is expressed at endogenous levels. Immunoaffinity or immunoprecipitation followed by LC-MS/MS does not readily permit determination of the stoichiometry of interacting partners. Additionally, when compared to yeast hybrid experiments, it is difficult to determine whether interactions are binary when identified in complexes by MS/MS. 

Titration. This technique provides a convenient method for following the stoichiometry of a reaction between a gas and a solution or pure liquid. Also, similar techniques can be applied to the study of gas-liquid equilibria and gas-solid equilibria. In a tensimetric titration, measured amounts of a gaseous reactant are added to a solution. After each addition a pressure measurement is made. A plot of these pressures versus the moles of added gas is then inspected for the break-point, which represents the stoichiometry of interaction (Fig. 9.4). 

Above we described only rather simple cases, for which the stoichiometry of interaction is 1 1 or n l, and cooperativity effect is absent. In strict terms, neither of the mentioned linear transformations is suited for more complex systems such as, e.g., bivalent antibodies and multivalent antigens. Only rough estimation of the total affinity is possible. Separate determinations of the affinity of all interactions occurring in a studied multivalent system (see, e.g., Ref. 7)... 

FIGURE 9.14 Stoichiometry of interaction in alginate-pectin mixed gels. (Redrawn from Toft et al., 1986). 

Because SPR responses are generally proportional to mass, it is possible to gain some insight into the stoichiometry of an interaction by comparing the amount of ligand immobihzed to the amount of analyte it can bind. If one knows the mass ratio of the two species then a stoichiometry of interaction can be calculated according to the following equation ... 

The aqueous method has been used for the determination of tautomeric structures (Miles, 1961 Miles et al., 1963) the demonstration of helix strandedness [i.e., the stoichiometry of interaction of polynucleotides to form two- or three-stranded helices (Miles and Frazier, 1964a)] the demonstration of a strand disproportionation reaction (Miles and Frazier, 1964b), of a strand displacement reaction (Sigler et al., 1962), and the quantitative analysis of different helical structures (Miles and Frazier, 1964a,b) in complex mixtures. It has been important in most of these applications that there are comparatively large differences of the spectra of the components from one another and there are marked and characteristic changes that the spectra undergo upon interaction to form helical secondary structures. 

Clipson, J.A., Roberts, G.A.R, 1994. Differential dyeing cotton. Part 2—stoichiometry of interaction with acid and direct dyes. J. Soc. Dyers Color. 110 (2), 69-73. 

Determination of stoichiometry of interaction between the components of the mixed solvent is discussed elsewhere. ... 

The stoichiometry of the prepared compounds depends not only on the composition of the initial mixture, but also on the initial oxide s fluorination activity. Unlike tantalum oxide, fluorination of niobium oxide by an ammonium hydrofluoride melt results in the formation of oxyfluoroniobates, but not of fluoroniobates. During the first step of Nb205 fluorination, (NH4)3NbOF6 is formed according to the following interaction [51, 52, 105, 111, 121, 122] ... 

The main advantages of the method can be formulated as follows. First, hydrofluoric acid is not needed for the decomposition stage the amount of fluorine required for the raw material decomposition can be calculated and adjusted as closely as possible to the stoichiometry of the interaction. Since the leaching of the fluorinated material is performed with water, a significant fraction of the impurities are precipitated in the form of insoluble compounds that can be separated from the solution, hence the filtrated solution is essentially purified. There is no doubt that solutions prepared in this way can be of consistent concentrations of tantalum and niobium, independent of the initial raw material composition. 

Kim et al. [601] investigated the influence of both temperature and the excess amount of sodium compared with the stoichiometry of the interaction. The molten system K2TaF7 - KC1 - KF was used for the experiments and the temperature varied in the range of 800-980°C. The excessive amount of sodium ranged from -10% to +10%. It was found that increasing either the temperature or the excess amount of added sodium led to an enhanced yield and increased the particle size of the tantalum powder. Optimal conditions were found to be 920°C and 5% excess reductant. 

Addition of trimethylaluminum to norcamphor (3), regardless of the stoichiometry of the reactants, leads to a mixture of the diastereomeric alcohols in a ratio of 95 5 also in favor of the erafo-alcohol6. Examination of the norcamphor model indicates that endo attack is sterical-ly more hindered than exo attack. However, steric interaction may not fully account for the exceptionally high exo selectivity. On the other hand, no severe torsional strain is involved if the nucleophile approaches the carbonyl group from the exo side, however, a nucleophile approaching from the endo side encounters torsional strain between the incipient bond and the C-l to C-6 carbon-carbon bond. Thus, in the case of norcamphor, steric and stereoelectronic factors reinforce each other, resulting in a strong directional influence for exo attack. 

These CD studies confirmed the binding stoichiometries of our aPNA-DNA complexes and provided further support for our binding model. Comparisons between the CD spectra of the individual components and the aPNA DNA complex suggest a template effect (not unhke that observed with certain DNA-binding proteins) where the components induce mutual conformational changes upon their interaction with each other. 

Based on the fact that pi-acids interact with the trinuclear gold] I) pi-bases, TR(carb) and TR(bzim), the trinuclear 3,5-diphenylpyrazolate silver(I) complex was reacted with each. Mixing [Au3(carb)3] or [Au3(bzim)3] with [Ag3(p,-3,5-Ph2pz)3] in CH2CI2 in stoichiometric ratios of 1 2 and 2 1 produced the mixed metal/mixed ligand complexes in the same gold-silver ratios. The crystalline products were not the expected acid-base adducts. It is suspected that the lability of the M-N bond (M=Au, Ag) in these complexes results in the subsequent cleavage of the cyclic complexes to produce the products statistically expected from the stoichiometry of materials used [74]. As a result of the lability of Au-N and Ag-N bonds, and the stability of... 

The value of EM for a cooperative self-assembled structure provides a measure of the monomer concentration at which trivial polymeric structures start to compete, and therefore EM represents the upper limit of the concentration range within which the cooperative structure is stable (Scheme 2). The lower limit of this range is called the critical self-assembly concentration (csac) and is determined by the stoichiometry of the assembly and the strength of the non-covalent binding interactions weaker interactions and larger numbers of components raise the csac and narrow the stability window of the assembly (8). Theoretical treatments of the thermodynamics of the self-assembly process have been reported by Hunter (8), Sanders (9), and Mandolini (10). The value of EM is lowered by enthalpic contributions associated with... 

The number of binding sites can be determined in this model by a plot of d Ink /dlnm at constant temperature, pH, and ion valency. To do that, it may be assumed that dlny /dlnm is approximately zero. The actual value is -0.04 for 0.1 to 0.5 M sodium chloride and less at lower concentrations. To a first approximation, the stoichiometry of water molecules released by binding protein could be determined from the slope of the plot of dink /dlnm vs. m. However, especially at low salt concentration and near the isoelectric point, the slope of such plots is nonlinear. The nonlinearity may be due to hydrophobic interaction between stationary phase and protein or a large change of ionic hydration on binding.34... 

By plotting i versus the ratio R = (CHX)t/(CB)t during the titration, they determined simultaneously the extent of acid-base interaction, the stoichiometry of that interaction and the degree of association of the acid-base adduct. Fig. 4.13 shows hypothetical titration curves line ABC corresponds to the interaction between B and HX as monomers without further reaction between BHX and HX, and the subsequent occurrence of the latter reaction to a small extent is indicated by the line ABC and to the full extent by line ABDE, when no more HX can react with BHX HX line AFDE arises when formation of BHX HX starts right away in the case of previous partial dimerization of B, the various lines will begin at A instead of A. 

The stoichiometry of an interaction between gas molecules and preadsorbed species may thus be deduced from the modifications of the Q-6 curves for a given reactant which are produced by the presence of preadsorbed species on the solid. The results are, of course, particularly conclusive when the differential heats of adsorption of small doses of reactant are measured in a sensitive calorimeter. But, such a qualitative analysis of the calorimetric data, though very useful, does not allow definite conclusions. In the preceding example, for instance, a fraction of carbon dioxide may remain adsorbed on the solid ... 

Numbers not in parentheses refer to the heat of adsorption or interaction, in kilocalories per mole or kilocalories per atom depending on the stoichiometry of the proposed interaction numbers in parentheses refer to the corresponding surface coverages in cubic centimeters per gram. 

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