Determining β‐sheet stability by Fourier transform infrared difference spectra

Abstract

We describe here a new method for determining the conformational stability of antiparallel β-sheets. Due to coupling between the transition dipoles, β-sheet conformations typically exhibit a characteristic high-frequency amide I component centered at ∼ 1680 cm⁻¹. Using one β-sheet protein and two small β-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 β-sheet systems. © 2004 Wiley Periodicals, Inc. Biopolymers, 2004