To eludicate the role in conformational stability of Cys residues buried in the interior of a protein, the thermodynamic properties of denaturation of mutant alpha subunit of Escherichia coli tryptophan synthase, in which Ser, Ala, Val or Gly was substituted for each of the three Cys residues, were analyzed using calorimetry. The mutants were less stable than the wild type, indicating that Cys residues contribute greatly to the stability of the alpha subunit. In most cases, a large decrease in denaturation enthalpy was observed, compensated for by the denaturation entropy to a major extent. The extent of changes in the denaturation Gibbs energy and denaturation enthalpy varied greatly depending on both substituting residues and positions. Of all the mutant proteins, the Cys154Ser mutant showed the greatest decrease in denaturation enthalpy; its denaturation enthalpy was half that of the wild type, and was considerably repaired by adding a ligand of the alpha subunit. Because the enthalpy of ligand binding to Cys154Ser in the native state did not change, it seems that the decrease in the denaturation enthalpy of Cys154Ser and its recovery by ligand binding are caused by conformational changes in the denatured state due to the mutation. Study holds ProTherm entries: 948, 949, 950, 951, 952, 953, 954, 955, 956, 957, 958, 11771, 11772, 11773, 11774, 11775, 11776, 11777, 11778, 11779, 11780, 11781 Extra Details: additive : EDTA(1 mM). tryptophan synthase alpha subunit; conformational stability ;,thermodynamic properties;calorimetry
Submitter: Connie Wang
Submission Date: April 24, 2018, 8:16 p.m.
|Number of data points||77|
|Proteins||Tryptophan synthase alpha chain ; Tryptophan synthase alpha chain|
|Assays/Quantities/Protocols||Experimental Assay: dCp temp:50.0 C ; Experimental Assay: dG ; Experimental Assay: dHcal ; Experimental Assay: Tm ; Experimental Assay: dCp ; Derived Quantity: ddG ; Derived Quantity: dTm|
|Libraries||Mutations for sequence MERYESLFAQLKERKEGAFVPFVTLGDPGIEQSLKIIDTLIEAGADALELGIPFSDPLADGPTIQNATLRAFAAGVTPAQCFEMLALIRQKHPTIPIGLLMYANLVFNKGIDEFYAQCEKVGVDSVLVADVPVEESAPFRQAALRHNVAPIFICPPNADDDLLRQIASYGRGYTYLLSRAGVTGAENRAALPLNHLVAKLKEYNAAPPLQGFGISAPDQVKAAIDAGAAGAISGSAIVKIIEQHINEPEKMLAALKVFVQPMKAATRS|