- Conference date: 29 Aug - 1 Sep 1999
- Location: Kobe-Sanda (Japan)
The Cro-V55C (cysteine cross-linked) dimer of the λ Cro repressor protein undergoes thermal unfolding in two discrete steps. The secondary structure of the stable equilibrium intermediate exhibits partial unfolding and reorganization at the N-terminal ends while other parts of the structure (some of the β-sheets) remain intact. To test whether the transition from the native to the intermediate state involves sequential events, we used a 2D-IR approach capable of detecting small differences of individual spectral features in response to external factors. The 2D-IR analysis shows that the intermediate state is formed in closely related sequential steps. To interpret the experimental 2D-IR data, 2D correlation plots for single and multiple sequential events were simulated. These plots were compared with the experimental data and translated into structural changes occurring within Cro-V55C. They reveal that the formation of the stable intermediate starts with the unfolding of the short N-terminal β-strand, followed by that of the three α-helices, and ends with the rearrangement of the remaining major β-sheet.
- Fourier transform infrared spectroscopy
- Secondary structure
- Spectrum analysis
- Thermal analysis
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