1995
DOI: 10.1021/bi00015a036
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A Calorimetric Study of the Thermal Stability of Barstar and Its Interaction with Barnase

Abstract: The temperature-induced unfolding of single, double, and triple mutants of barstar, the specific intracellular protein inhibitor of barnase from Bacillus amyloliquefaciens, has been studied by high-sensitivity differential scanning calorimetry. The thermal unfolding of barstar mutants, where at least one of the two cysteine residues in the molecule had been replaced by alanine, follows a two-state mechanism at neutral and alkaline pH. The unfolding enthalpy and heat capacity changes are slightly lower than tho… Show more

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Cited by 43 publications
(36 citation statements)
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“…DSC transitions of proteins have proved to be very sensitive to ligand binding and so DSC has often been used in the past to analyse the thermodynamics of proteinligand interaction in preference to ITC under some circumstances. [37][38][39][40] We have shown here and elsewhere 17 that intramolecular binding of lowaffinity sequences has a considerable impact upon the stability and dynamic properties of an SH3 domain. Thus, since the thermodynamics of the SH3-peptide interaction are affected considerably by simple mutations, these effects (not only the changes in affinity but also in the enthalpy-entropy balance) can be separated from the unfolding thermodynamics of the SH3 domain by a threestate analysis of the DSC data of the chimeric proteins.…”
Section: Discussionmentioning
confidence: 57%
“…DSC transitions of proteins have proved to be very sensitive to ligand binding and so DSC has often been used in the past to analyse the thermodynamics of proteinligand interaction in preference to ITC under some circumstances. [37][38][39][40] We have shown here and elsewhere 17 that intramolecular binding of lowaffinity sequences has a considerable impact upon the stability and dynamic properties of an SH3 domain. Thus, since the thermodynamics of the SH3-peptide interaction are affected considerably by simple mutations, these effects (not only the changes in affinity but also in the enthalpy-entropy balance) can be separated from the unfolding thermodynamics of the SH3 domain by a threestate analysis of the DSC data of the chimeric proteins.…”
Section: Discussionmentioning
confidence: 57%
“…The enthalpy of unfolding of the apoprotein is about 50 kcal mol", a little less than half that of the holoprotein, and the heat capacity change for unfolding is substantially decreased, to about 0.3 kcal K" mol". These values are generally larger than have been observed in other systems for partially folded species or "molten globules" both in magnitude and as a fraction of the native protein values (Xie et al, 1991;Yutani et al, 1992;Ogasahara et al, 1993;Martinez et al, 1995;Genzor et al, 1996;Kreimer et al, 1996). Dramatic changes occur in the structure of cyt bs62 when the heme is removed (Fig.…”
Section: Apocytochrome B562 Denaturationmentioning
confidence: 73%
“…Barstar C40/82A has been used in several studies related to protein folding (Schreiber & Fersht, 1993;Martínez et al, 1995). One complication of studies justifies the use of the C40/82A mutant for folding studies.…”
Section: Discussionmentioning
confidence: 99%