SPECTROSCOPIC AND VOLUMETRIC INVESTIGATION OF CYTOCHROME-C UNFOLDING AT ALKALINE PH - CHARACTERIZATION OF THE BASE-INDUCED UNFOLDED STATE AT 25-DEGREES-C

We have measured at 25 degrees C the relative specific sound velocity increment, [u], and the partial specific volume, v degrees, of cytochr ome c as a function of pH, Our data reveal that the base-induced nativ e to unfolded transition of the protein is accompanied by a volume dec rease of 0.014 cm(3) g(-1) and a compressibility decrease of 3.8 x 10( -6) cm(3) g(-1) bar(-1). These results allow us to conclude that, rela tive to a fully unfolded conformation, the base-denatured state of cyt ochrome c has only 70 to 80% of its surface area exposed to the solven t. Recently, we reported a similar result for the acid-denatured state of cytochrome c. Thus, insofar as solvent exposure is concerned, both the base- and the acid-induced unfolded states of cytochrome c retain some order, with 20 to 30% of their surface areas remaining solvent-i naccessible. We discuss the implications of this result in terms of de fining potential intermediate states in protein folding pathways.