Ribonuclease pressure denatured

A good example of application is given by the protein structural changes of bovine ribonuclease A in the course of its denaturation by pressure. The UV spectrum of RNase is dominated by the absorbance of tyrosine - this RNase does not contain tryptophan. As shown in Figure 6, an increase of pressure from 1 to 500 MPa results in a blue-shift of the 4th derivative maximum from 285.7 0.05 to 283.5 0.05 nm. This shift of 2.2 nm corresponds to an increase of the mean dielectric constant from 25 to 59. It is characteristic of the exposure to the aqueous solvent of part of the 6 tyrosines, as it is expected for a partly denaturation. The transition is fully reversible with clear isosbestic points. The pressure effect can therefore be described by a simple two-state model between the native (e,. = 25) and the partially denatured (e,. = 59) state. A simulation on the basis of this model permitted us to determine the thermodynamic parameters of this transition AG° = 10.3 kJ/mol and AV = - 52 ml/mol. A comparison with results obtained by other methods indicates that the (e,. = 59) state corresponds to an intermediate in the defolding process which has molten globule like characteristics [12]. It thus appears that fourth derivative... 

Investigation of the equilibrium between native and unfolded conformations formed on solvent denaturation, temperature, and pressure have now been performed on a wide range of proteins. For example, James and Sawan have studied the effect of increasing guanidinium chloride concentration on the mobility of individual histidine residues in ribonuclease (pancreatic) C n.m.r. spectroscopy using spin-lattice relaxation in an off-resonance rotating frame. They found that L-histidines-12, -105, and -119 increase in mobility up to a denaturant concentra-... 

Figure 25.28 Pressure can denature proteins, ribonuclease A in this case. Source J Zhang, X Peng, A Jones and J Jones, Biochemistry 34, 8631-8641 (1995).

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