Guanidinium hydrochloride unfolded proteins

The intrinsic fluorescence of the eight tryptophanyl residues in MBP can be used to measure its reversible unfolding/refolding reactions. When unfolded by incubation in a denaturant such as guanidinium hydrochloride, the side chains of the tryptophanyl residues are exposed to solvent and their fluorescence is quenched. On dilution of the denaturant, the subsequent refolding reaction can be monitored by the increase in fluorescence that results from burial of the side chains in the interior of the protein and their shielding from solvent (Liu et al., 1988, 1989). 

For example, increasing the temperature or addition of chemical denaturants such as urea or guanidinium hydrochloride (GuHCl) produce changes in fluorescence or circular dichroism spectra of proteins in solution consistent with unfolding of the protein structure (Figures 5.10 and 5.11). 

Conformational changes may also be detected by c.d. The denaturation of a-lactalbumin with guanidinium hydrochloride was studied by Kuwajima. The unfolding process, which involves three stages with an a-helical intermediate state, was followed by c.d. and difference spectra. Similarly, Takahashi et al. noted that disulphide-reduced ribonuclease A had a c.d. spectrum different from either the native or random coil protein. The reoxidation of the reduced enzyme (which results in recovery of activity) was followed by c.d. and it was shown that the change in a-helix was more rapid than that in j -structure. 

The unfolding of a protein may be brought about by treatment with demturants, substances such as guanidinium hydrochloride... 

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