Intramolecular disulfide bonds in protein molecules, whether present in the wild-type protein or engineered via site-directed mutagenesis, are capable of significantly increasing the stability. Establishing thermodynamic parameters associated with the redox formation of this linkage is often encumbered by other global structural changes within the protein molecule. The active site of Escherichia coli thioredoxin possesses a disulfide/dithiol in a short loop, oxidation/reduction of which is accompanied by little structural alteration of the protein. Data for the thermal denaturation of the reduced protein are presented, which on comparison to the data obtained for the oxidized form [Ladbury, J.E., Wynn, R., Hellinga, H.W., & Sturtevant, J.M. (1993) Biochemistry 32, 7526-7530] are used to establish thermodynamic parameters for the redox reaction in this molecule. Data for an isosteric double mutation in the active site of thioredoxin (Cys32Ser/Cys35Ser) are also presented. Although the wild-type and mutated proteins show a similar reduction in free energy compared to the oxidized form (-3.0 +/- 0.4 and -3.1 +/- 0.3 kcal mol-1, respectively), the enthalpic and entropic contributions to this destabilization are different for the two proteins. Study holds ProTherm entries: 4493, 4494, 4495, 4496, 4497 Extra Details: disulfide bonds; redox formation; global structural changes;,active site; enthalpic and entropic contributions
Submitter: Connie Wang
Submission Date: April 24, 2018, 8:25 p.m.
|Number of data points||17|
|Proteins||Thioredoxin 1 ; Thioredoxin 1|
|Assays/Quantities/Protocols||Experimental Assay: dCp pH:8.0 ; Experimental Assay: dHcal pH:8.0 ; Experimental Assay: Tm pH:8.0 ; Experimental Assay: dCp pH:7.0 ; Experimental Assay: dHcal pH:7.0 ; Experimental Assay: Tm pH:7.0 ; Experimental Assay: dCp pH:6.0 ; Experimental Assay: dHcal pH:6.0 ; Experimental Assay: Tm pH:6.0 ; Derived Quantity: dTm pH:7.0 ; Derived Quantity: dTm pH:6.0|
|Libraries||Mutations for sequence SDKIIHLTDDSFDTDVLKADGAILVDFWAEWCGPCKMIAPILDEIADEYQGKLTVAKLNIDQNPGTAPKYGIRGIPTLLLFKNGEVAATKVGALSKGQLKEFLDANLA|