CALCIUM-INDUCED DIMERIZATION OF TROPONIN-C - MODE OF INTERACTION AND USE OF TRIFLUOROETHANOL AS A DENATURANT OF QUATERNARY STRUCTURE

Protein aggregation can be a problem, especially as a large number of proteins become available for structural studies at fairly high concen trations using solution techniques such as NMR spectroscopy. The muscl e regulatory protein troponin C (TnC) undergoes a calcium-induced dime rization at neutral pH with a dissociation constant for the dimerizati on of 0.4 mM at 20 degrees C. The present study indicates that the mod e of dimerization involves the N-domain of one monomer interacting wit h the N-domain of another monomer. Addition of the solvent trifluoroet hanol (TFE) to a concentration of 15%, v/v, results in a 10-fold incre ase in the dimer dissociation constant of calcium-saturated TnC (4 mM at 20 degrees C), making TnC predominantly a monomer for spectroscopic studies. Further, TFE, at the concentrations used herein, acts to per turb the quaternary structure of TnC without adversely affecting the s econdary or tertiary structure as evidenced by minimal changes to its CD spectra and H-1, C-13, and N-15 NMR chemical shifts.