A direct conflict between the stabilization free
energy parameters of cytochrome c determined by
optical methods and by hydrogen exchange (HX) is quantitatively
explained when the partially folded intermediates seen
by HX are taken into account. The results support the previous
HX measurements of intermediate populations, show how intermediates
can elude the standard melting analysis, and illustrate
how they confuse the analysis when they are significantly
populated within the melting transition region.