Determining (beta)-sheet stability by Fourier transform infrared difference spectra

We describe here a new method for determining the conformational stability of antiparallel (beta)-sheets. Due to coupling between the transition dipoles, (beta)-sheet conformations typically exhibit a characteristic high-frequency amide I component centered at ~1680 cm^-1. Using one (beta)-sheet protein and two small (beta)-hairpins, we demonstrate that this high-frequency component, which is fairly narrow (~8-10 cm^-1), can be quantitatively resolved and used in thermal stability determination. Compared with the commonly used CD and fluorescence techniques, this ir method offers advantages. Since the area of this high-frequency component is only proportional to the folded population, it eliminates the need for a priori information of the folded and unfolded baselines encountered in other methods. Thus, it is applicable to a variety of (beta)-sheet systems.