Energetics of ribonuclease T1 structure

Abstract

The energetics of thermal denaturation of two isoforms of ribonuclease T1 (Gln25 and Lys25) in various solvents have been studied by differential scanning calorimetry. It has been shown that the thermal transition of both forms of RNase T1 is strongly affected by slow kinetics, which cause an apparent deviation of the transition from a simple two-state model. By decreasing the heating rate or increasing the transition temperature, the denaturation of RNase approaches an equilibrium two-state transition. This permits determination of the thermodynamic parameters characterizing unfolding of the native structure. These thermodynamic parameters were correlated with the structural features of protein. Analysis of different contributions to the stability of RNase T1 shows that van der Waals interactions and hydrogen bonding are the major contributors to the conformational stability of the protein.

Denaturant m values and heat capacity changes: Relation to changes in accessible surface areas of protein unfolding

“…In addition, Bolen and Santoro (1988) and Santoro and Bolen (1988) have shown that the denaturation of phenylmethanesulfonyl a-chymotrypsin by urea, Gdn HCI, and 1,3-dimethyl urea analyzed by the LEM fit all the requirements of a thermodynamic cycle. The AGHZO values from linear extrapolation of urea denaturation for RNase TI agree remarkably well with free energies obtained from thermal denaturation and DSC under a variety of conditions (Hu et al, 1992b;Yu et al, 1994). Similarly good agreement between urea and Gdn HCI data analyzed by the LEM and thermal denaturation by DSC was found by Santoro and Bolen (1992) for thioredoxin.…”

“…a Swint and Robertson, 1993;' O'Neil et al, 1995;Hurle et al, 1990;Viguera et al, 1994; e Jackson et al, 1993;'Akke and Forsen, 1990;Khorasanizadeh et al, 1993;Wintrode et al, 1994;Scholtz, 1995;J Agashe and Udgaonkar, 1995;Lim et al, 1992; ' Marqusee and Sauer, 1994;McLendon and Smith, 1978;Hagihara et al, 1994; O Privalov and Gill, 1988;P Pace et al, 1990;Shirley et al, 1992;Yu et al, 1994; ' Bowie and Sauer, 1989; ' Egan et al, 1993;Ramdas et al, 1986;"Bryant et al, 1985; " Cohen and Pielak, 1994;x Kelley et al, 1987;Santor0 and Bolen, 1992;'Clarke and Fersht, 1993; aa Pace et al, 1992;" Griko et al, 1994; cc Greene and Pace, 1974;dd Munson et al, 1994;Filimonov et al, 1993; f f Saito and Wada, 1983; gg Ahmad and Bigelow, 1982; hh Taniyama et al, 1992; I' Herning et al, 1992;'' Ropson et al, 1990; kk Shortle and Meeker, 1986;"Carra et al, 1994; mm Craig et al, 1987; "" Makhatadze et al, 1994;O0 De Young et al, 1993...…”

How to measure and predict the molar absorption coefficient of a protein

“…In trying to resolve this question, the Privalov laboratory measured E = 20,630 M" cm-I using amino acid analysis and a Kjeldahl nitrogen method (Yu et al, 1994). Their preferred method is the Kjeldahl procedure of Jaenicke (1974); we tried this method in our laboratory and found 19,630 M" cm".…”

“…The E values in entries 48-57 are from Kalnin et al (1990). Entries 58-66 are from the Privalov lab (Privalov et al, 1989;Griko et al, 1994;Yu et al, 1994). For barnase and mutants, entry 67 is from Lees and Hartley (1966), and 68-72 are from Loewenthal et al (1991).…”

Anion binding to the ubiquitin molecule

“…This heating rate is low enough to provide equilibrium conditions upon thermal unfolding. One of the indications of the equilibrium character of unfolding of any small globular human ubiquitin is equality of the calorimetric and fitted enthalpies as shown in Table 1 (see also Yu et al, 1994). Reversibility of the unfolding transition under several conditions was examined by rescanning the sample.…”