Partially folded conformations in the folding pathway of bovine carbonic anhydrase II: a fluorescence spectroscopic analysis.

GdmCl-, urea-, and pH-induced unfolding pathways of bovine carbonic anhydrase II have been analyzed by using changes induced by different denaturing agents in intensity, anisotropy, life time, and parameter A value of intrinsic fluorescence as well as intensity and life time of ANS (ammonium salt of 8-anilinonaphthalene-1-sulfonic acid) fluorescence. The formation of several stable unfolding intermediates, some of which were not observed previously, has been established. This was further confirmed by representation of fluorescence data in terms of a "phase diagram", that is, I(lambda1) versus I(lambda2) dependence, where I(lambda1) and I(lambda2) are the fluorescence intensity values measured at wavelengths lambda(1) and lambda(2), respectively.