Binding Thermodynamics Of Actin-Binding Proteins: Myosin S1 And The N-Terminus Of Dystrophin

Jeffrey G. Forbes, Frederick P. Schwarz, Kurt J. Amann, and James M. Ervasti

Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742-2021; CARB/NIST, 9600 Gudelsky Drive, Rockville, MD 20850; Department of Physiology, University of Wisconsin, Madison, WI 53706

Protein-protein interactions, that occur largely through hydrophobic effects where water is excluded from the interface, are often entropy-driven processes. An entropy-driven process occurs when T S > H and in many cases H > 0. Actin polymerization, myosin assembly and F-actin-myosin binding are all entropy-driven processes. Our measurements on a recombinant fragment of the first 246 amino acids of human dystrophin yielded entirely different results. Titration microcalorimetry measurements on the dystrophin fragment gave a dissociation constant of 16 µM and an enthalpy of binding of -67 kJ/mol. The thermodynamic values for the interaction of the N-terminus of human dystrophin with rabbit muscle F-actin at 25 °C are as follows: delta G = -27.3 kJ/mol; delta H = -67~kJ/mol; delta S = -133 J/mol/K. As these values indicate, the interaction between the N-terminus of dystrophin and F-actin is an enthalpy-driven process rather than the entropy-driven process that occurs with myosin binding. Many of the actin residues which have been identified as being involved in myosin binding have also been identified in the interaction of the N-terminus of dystrophin with F-actin. Our microcalorimetric data indicates that even though these two different proteins interact with many of the same amino acids on F-actin, their binding mechanisms are fundamentally different.

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