Kinetic binding association rate constants

The data in the upper and lower panels were fit simultaneously with a single association rate constant (k = 3.23 x lO s ) and separate dissociation rate constants (k = 0.0108/s, upper panel 0.083/s, lower panel). The kinetic aspects of the fit were verified by the agreement with the equilibrium binding (see Figure 4 caption). 

The second and third relaxation processes were coupled, where the observed rate constants differed by a factor of 3 to 7 and the rate constant for each relaxation process varied linearly with the DNA concentration.112 This dependence is consistent with the mechanism shown in Scheme 2, where 1 binds to 2 different sites in DNA and an interconversion between the sites is mediated in a bimolecular reaction with a second DNA molecule. For such coupled kinetics, the sum and the product of the two relaxation rate constants are related to the individual rate constants shown in Scheme 2. Such an analysis led to the values for the dissociation rate constants from each binding site, one of the interconversion rate constants and the association rate constant for the site with slowest binding dynamics (Table 2).112 The dissociation rate constant from one of the sites was similar to the values that were determined assuming a 1 1 binding stoichiometry (Table 1). 

Kinetic assays give access to the binding reaction s forward and reverse rate constants, i.e. the association rate constant fe+i and the dissociation rate constant fe i that characterize the association and the dissociation of the target-marker complex and the Kj [see Eq. (4)]. 

The binding kinetics were characterized in terms of the apparent time constant (K pp = kf C + k ) where C = analyte concentration kf = association rate constant and k = dissociation rate constant. In closed loop experiments, a plateau value for K pp of 0.0024/s was reached at a linear flow rate of 2.67 mL/min. K ppWas foxmd to decrease with decreasing antigen concentration (C), with equilibrium achieved only at the highest level (1 pg/mL). The association rate constant Kf was calculated at 3.6 x 10 M/s for IgG binding. 

A more detailed analysis of ligand binding is possible by determination of the association rate constant ( on-kinetics ) of the dissociation rate constant ( off-kinetics ). The methodology of these estimations is illustrated for the cardiac DHP receptor. 

Binding kinetics should be proportional to the rate of the in vivo response and should yield an equilibrium constant equal to the dissociation rate constant divided by the association rate constant. 

An antenna remains in a plume 1 s and an antenna is not an isolated system, as is required to reach equilibrium. The kinetic properties of the PBP-ligand complexes may be more important to the function of PBPs as potential filters than the equilibrium dissociation constants. Thus, ligands with very fast association rate constants and very slow dissociation rate constants are more likely to be bound at the pore surfaces and to traverse the sensillar lymph unharmed by the powerful pheromone-degrading enzymes in the lymph (see below). Thus, in order to understand the function of PBPs, it is essential to obtain more data on binding kinetics. 

In the kinetic studies of the adsorption process, the mass transport of the analyte to the binding sites is an important parameter to account for. Several theoretical descriptions of the chromatographic process are proposed to overcome this difficulty. Many complementary experiments are now needed to ascertain the kinetic measurements. Similar problems are found in the applications of the surface plasmon resonance technology (SPR) for association rate constant measurements. In both techniques the adsorption studies are carried out in a flow system, on surfaces with immobilized ligands. The role of the external diffusion limitations in the analysis of SPR assays has often been mentioned, and the technique is yet considered as giving an estimate of the adsorption rate constant. It is thus important to correlate the SPR data with results obtained from independent experiments, such as those from chromatographic measurements. 

Temperature-jump relaxation and the stopped-flow methods are suitable to follow the concentration changes over extremely short time intervals. Such studies have indicated that immune reaction kinetics resemble other biological systems in which ligands are bound to proteins (Weber, 1975) in that the binding strength of small molecules is largely dictated by the constant. The association rate constants ka, are very similar for various antibody-antigen systems, i.e., for... 

The binding of several small molecules to cytochrome c peroxidase has been studied kinetically. In the case of fluoride, the data are consistent with the interaction of F with the protonated form of the enzyme (pJta 5.5) or with the interaction of HF with the ionized form of the protein. The latter is the preferred interpretation, giving a pH-independent rate constant for binding of 5.1 x 10 1 mol s. With cyanide the situation is rather more complex the maximum value of the association rate constant, which occurs between pH 6 and 8, is 1.1 x 10 1 mol S . ... 

The kinetics of the binding of oxygen to the haemocyanin from the meirine gastropod Buccinum undatum under various conditions have been reported. In the presence of calcium ions the association rate constant is 7.8 x 10 1 mol s (25 "Q, and it is almost independent of pH. On the other hand, the dissociation rate constant increases significantly over the pH range 7.0 (A d=13 s ) to 8.0 (56 s ). The rate constant for the reaction of O2 with (undissociated) haemocyanin from Panulirus... 

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