Transition of ribonuclease

Weame, S. and Creighton, T. (1988) Further experimental studies of the disulfide folding transition of Ribonuclease A. Prot. Struct. Funct. Genet. 4, 251-261. 

Table II. Effect of Cosolvents on the Midpoint (Tm) of the Reversible Native Denatured Transition of Ribonuclease A at pH4 2.8°...

Thermal transitions of ribonuclease, chymotrypsinogen (with disulfide bonds intact) were reinvestigated by temperature jump in a time range of observation which could extend down to few microseconds allowing detection of rapid phases, under the same conditions (pH, concentration) used by Pohl (1968a). 

Navon A, Ittah V, Laity JH, et al. Local and long-range interactions in the thermal unfolding transition of bovine pancreatic ribonuclease A. Biochemistry 2001 40 93-104. 

No large conformational changes occur in the enzyme during catalysis, but many small movements take place. The structural basis for the catalytic power of ribonuclease thus resides in several different features tight, specihc binding of a strained conformation of the substrate, general acid-base catalysis by His-12 and His-119, and preferential stabilization of the transition state by ionic interactions with Lys-41. 

Recently the related cyclization of the phenyl ester of c/j-tetrahydrofuran-3,4-diol monophosphate to the corresponding five-membered phosphate with loss of phenol has been shown to be subject to general catalysis by imidazole132. This reaction serves as a model for the first step in the action of ribonuclease which leads to the formation of the nucleoside 2 ,3 -cyclic phosphate. The actual details of the transition state leading to the cyclic phosphate as catalyzed by the enzyme are presently the subject of some debate. One possibility is the in-line mechanism (53)... 

The fluorescence of ribonuclease solutions has been studied extensively by Cowgill. The absence of tryptophan permits the tyrosine fluorescence to be observed. The tyrosine fluorescence of RNase is very low in comparison with the maximum expected from its tyrosine content. All methods of denaturing RNase lead to an increase in fluorescence. Transitions, as indicated by the pattern of fluorescence change vs. denaturant concentration, are about the same as those indicated by other physical techniques [see, e.g., Gaily and Edelman (305) ]. 

Vanadate itself is a much poorer inhibitor of ribonuclease A than is the VUr complex. This makes sense because vanadate alone cannot mimic the transition state. Covalently bound components, the parts of the nucleoside covalently bound to the phosphate moiety, are needed to complete the transition-state-like structure. They... 

Leon-Lai, C.H., M.J. Gresser, and A.S. Tracey. 1996. Influence of vanadium(V) complexes on the catalytic activity of ribonuclease A. The role of vanadate complexes as transition state analogues to reactions at phosphate. Can. J. Chem. 74 38 -8. 

Borah, B., C.W. Chen, W. Egan, M. Miller, A. Wlodawer, and J.S. Cohen. 1985. Nuclear magnetic resonance and neutron diffraction studies of the complex of ribonuclease A with uridine vanadate, a transition-state analogue. J. Biochem. 24 2058-2067. 

The initial decrease in optical rotation found in aqueous solutions of /3-lactoglobulin and ovalbumin is not, however, sufficient to differentiate globular proteins from simpler synthetic polypeptides in their transition behavior, for neither ribonuclease nor human serum albumin appear to exhibit it. The specific rotation of ribonuclease in water-2-chloroethanol mixtures becomes steadily less levorotatory as the proportion of nonpolar solvent increases (Weber and Tanford, 1959). In the case of human serum albumin Bresler (1958) and Bresler el al. (1959) find that only progre.ssive increases in specific rotation occur as the concentration of 2-chloroethanol is increased and that this change is accompanied by a steady rise in viscosity and the corresponding axial ratios characteristic of the formation of rodlike particles. If these proteins do have some initial helical content in water, as can be argued from their optical rotatory dispersion, then it appears that hydrophobic forces are not required for the stability of these regions. 

That c in faet reflects helical content is shown by its rise in proteins for which there is independent evidence that helical content is being increased. The nonaqueous transitions of insulin, native and oxidized ribonuclease, silk fibroin, and oxidized bovine serum albumin (Yang and Doty, 1957) as well as that for carboxymethylated human serum albumin (Bresler... 

FIGURE 7.4 Transition of proteins from the native to the unfolded state or vice versa, (a) Ribonuclease at pH 3.15, as a function of temperature, (b) Lysozyme as a function of guanidinium chloride concentration, (c) Nuclease A as a function of pH. 

Figure 4. Ionic dependence of transition T, Tu, of ribonuclease (29) (—,) Structure T, determined spectroscopically at the same conditions as related Tm values. Indicated is the cause of the change in Tj( by the ion influence of the water...

Fig. 13.26. Active site of ribonuclease A (5RSA, stereo-diagram) with the co-crystallized transition-state analog uridine vanadate. The vanadium atom occupies the center of a distorted TBP with 0(2 ) and 0(7) in the apical positions, 0(3 ), 0(6) and 0(8) in the equatorial plane. The two catalytic His-residues are hydrogen bonded to 0(3 ) and 0(8)...

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