Further examination of the intermediate state in the denaturation of the tryptophan synthase alpha subunit. Evidence that the equilibrium denaturation intermediate is a molten globule.

To further characterize the intermediate state in the denaturation of tryptophan synthase alpha subunit from Escherichia coli, we have carried out differential scanning calorimetry in various concentrations of urea near pH 8.5. The heat capacity curve of the intermediate has no excess heat capacity nor any transition. This indicates that the intermediate is a thermodynamically denatured form. Although the intermediate retains significant CD signal in the far-uv region, the tertiary structure of the intermediate is disrupted as judged by the near-uv CD spectra and 1H NMR spectra in the aromatic region. This intermediate might be similar to a molten globule state. These results do not support our earlier proposal that the intermediate of the alpha subunit in the denaturation process retains an intact N-terminal domain, but that the C-terminal domain unfolds.