Intrinsic viscosity ribonuclease

The intrinsic viscosity of native ribonuclease is very low. Harrington and Schellman (247) reported 3.3 ml/g at neutral pH in 0.1 M KC1. Buzzell and Tanford (265) found values of 3.3-3.5 ml/g over the entire pH range from 1 to 11 and ionic strengths from 0.05 to 0.25 M. This value increases dramatically on denaturation even without oxidation or reduction of the disulfide bonds to 8.5 ml/g (266). In the presence of reducing agents and 6 M guanidine hydrochloride the value is 16.0 ml/g (267). 

Analogous shifts in protein spectra have been observed as a result of conformational changes associated with denaturation. Initial reports on this phenomenon (see Ref 24) attributed the spectral difference to the transfer of the aromatie amino acids from the hydrophobic interior of the protein to the more aqueous surface environment as a result of the conformational change. Spectral changes for several proteins correlated well with independent measurements of denaturation such as intrinsic viscosity, circular dichroic spectra, and heat capacity measurements. For example, Edelhoch [26] compared the ribonuclease UV spectrum in buffer with that obtained in 6M guanidine hydrochloride (GuHCI). 

PDA analysis was also applied to the study of ribonuclease A in reversed-phase chromatography [33]. Ribonuclease A conformation has been the focus of numerous studies, and its dcnaturation is an excellent example of a reversible unfolding process. Cohen et al. [31] first reported the influence of a hydrophobic stationary phase on ribonuclease dcnaturation. In this work, the change in the 254/280 nm absorbance ratio was followed using multiple injections of the same sample. The increase in the ratio correlated extremely well with a number of other physical measurements of structural changes such as circular dichroism and intrinsic viscosity. 

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