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Updated: Jul 6, 2026

Measurement of Force-Sensitive Protein Dynamics in Living Cells Using a Combination of Fluorescent Techniques
Published on: November 2, 2018
Investigating complexity of protein-protein interactions in focal adhesions
Tanmay P Lele1, Charles K Thodeti, Jay Pendse
1Department of Chemical Engineering, University of Florida, Gainesville, FL 32611, USA.
Abstract:
The formation of focal adhesions governs cell shape and function; however, there are few measurements of the binding kinetics of focal adhesion proteins in living cells. Here, we used the fluorescence recovery after photobleaching (FRAP) technique, combined with mathematical modeling and scaling analysis to quantify dissociation kinetics of focal adhesion proteins in capillary endothelial cells. Novel experimental protocols based on mathematical analysis were developed to discern the rate-limiting step during FRAP. Values for the dissociation rate constant k(OFF) ranged over an order of magnitude from 0.009+/-0.001/s for talin to 0.102+/-0.010/s for FAK, indicating that talin is bound more strongly than other proteins in focal adhesions. Comparisons with in vitro measurements reveal that multiple focal adhesion proteins form a network of bonds, rather than binding in a pair-wise manner in these anchoring structures in living cells.
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