title: Structural comparison between oxidized and reduced Escherichia coli thioredoxin. Proton NMR and CD studies.
abstract: Escherichia coli thioredoxin (Mr 11,700) usually functions as a hydrogen carrier protein that undergoes reversible oxidation/reduction reactions of its active-site disulfide linkage. By use of a number of assigned and identified resonances in one- and two-dimensional 1H NMR spectra, the two forms of the protein have been compared. Only groups that are relatively close to the active-site Cys-32, Cys-35 linkage such as Trp-28, Trp-31, Phe-27, Ala-29, and Val-25 undergo substantial changes in their 1H NMR chemical shift upon reduction. Various residues that are further removed from the active site, like Tyr-49, Tyr-70, His-6, Phe-12, Phe-81, and Phe-102, appear to be little affected (less than 0.02 ppm) by the reduction, suggesting that the rest of the protein structure is not much affected. Thus, the structural changes that occur upon reduction appear to be localized to the disulfide-containing turn and the central strand of the twisted beta-sheet that directly leads to this turn. Notwithstanding the apparent similarity in the secondary and tertiary structures of the oxidized and reduced forms of the protein, the thermal stability of the protein decreases by 10 degrees C upon the reduction of the single disulfide. This was found by both 1H NMR and near- and far-ultraviolet circular dichroism studies. Oxidized thioredoxin was also more resistant to alkaline denaturation. Furthermore, the exchange rate of the relatively stable slow-exchanging backbone amide protons that are part of the core of the twisted five-stranded beta-sheet of thioredoxin increases substantially after reduction.(ABSTRACT TRUNCATED AT 250 WORDS)
mesh_terms: D001426:Bacterial Proteins; D002942:Circular Dichroism; D004926:Escherichia coli / Q000032:analysis*; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D011489:Protein Denaturation; D013696:Temperature; D013879:Thioredoxins
chemical_list: D001426:Bacterial Proteins; D013879:Thioredoxins