Calmodulin dissociation constants

Fig. 7. (A) Aligned, partial sequences of a number of calmodulin-binding peptides. The boxes indicate residues that are generally occupied by apolar residues. Reported dissociation constants for interaction with calmodulin are given on the right. LK2, A mode peptide VIP, vasoactive intestinal peptide GIP, gastric inhibitory peptide. (B) The mean hydropho-bicities for the residues at a given position were plotted versus their position in the aligned sequence. The horizontal bar indicates the period of an a helix. From Cox et al. (1985).

Reversible Ligand Binding The protein calcineurin binds to the protein calmodulin with an association rate of 8.9 X 10s M-1s-1 and an overall dissociation constant, Kd, of 10 nM. Calculate the dissociation rate, kd, including appropriate units. 

Surface plastnon resonance Association constant Dissociation constant Hexaethylene glycol spacer Sambucus nigra agglutinin High-mobility-group transcriptional factor Soluble green fluorescent protein Calmodulin... 

Fluorescence and affinity measurements - Peptide in 25 mM Tris, 100 mM KCl and 1 mM CaCl2 at pH 7.5 and 30 C was titrated with a stock solution of calmodulin in UV transmitting plastic cuvettes since the peptides appear to bind to glass. Fluorescence titration spectra were recorded using a SPEX FluoroMax fluorescence spectrometer with excitation at 280 nm and emission scanned from 310 to 390 nm. The value of fluorescence intensity at 330nm was plotted as a function of calmodulin concentration and fitted using standard non-linear least squares methods (6) to obtain optimal values of the dissociation constant (Kj) and the maximum fluorescence enhancement (F/F ). The detection limit under our experimental conditions was 50 nM peptide and all quoted Kj values are the average of at least 3 independent determinations. 

Table 1 - Dissociation constants of wildtvne and four mutant calmodulins for WFF and FFW peptides and fluorescence enhancement (at X=330 nm) upon complex formation ...

Fig. 1 shows that lathyrls calmodulin is a good stimulator of bovine heart phosphodiesterase. The data were best fitted with two distinct dissociation constants, indicating an interaction by two different modes. 

Figure 3. Simulation of the kinetic scheme from Figure 2 using the constants from Table 3. The top figure represents the kinetics of Ca dissociation which has a biphasic response (fast phase 663 s and slow phase 9 s ). The middle figure represents the titration of calmodulin by Ca ". The signal rising between 0 and 2-3 Ca " ions is associated with the occupancy of the sites from the COOH terminus and the other signal is associated with the occupancy of the N-terminal sites. The bottom figure is a Scatchard representation of the direct calcium binding isotherm.

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