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Figure 4


Fig. 4. Two binding sites for Crac-GFP in living cells. (A) Three states of cells; cells before stimulation, 0 to 20 seconds after stimulation, and more than 20 seconds after stimulation were referred as unstimulated, stimulated and adapted, respectively. (B) Dissociation curves of Crac-GFP and PHCrac-GFP bound to the membranes. Lifetime of Crac-GFP was measured in the cells before stimulation ({circ}), at 0-20 seconds ({triangleup}) and more than 20 seconds ({square}) after stimulation with cAMP. The curves were fitted to a sum of exponentials with the time constant {tau} and the relative ratios (see Results). The lines represent the fitting curves. Dissociation curve of PHCrac-GFP (bullet) bound to the membrane of resting cells demonstrates that the fast-dissociation site is PtdIns(3,4,5)P3.





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