Infrared spectroscopic discrimination between α‐ and 310‐helices in globular proteins

We have undertaken a new and more detailed Fourier‐transform infrared (FTIR) spectroscopic study of α‐lactalbumin (in D2O solution) aimed at correlating its secondary structures to observed Amide I’ infrared bands. The spectra reported here were interpreted in light of the recently determined crystal structure of α‐lactalbumin and by comparison with the spectra and structure of the homologous protein lysozyme. Of particular importance is the new evidence supporting the assignment of the band at 1639 cm−1 to 310‐helices. This assignment is in excellent agreement with one based on theoretical and experimental studies of 310‐helical polypeptides. The frequency observed for 310‐helices is distinctly different from that at which α‐helices are typically found (viz., around 1655 cm−1). In the present study, two bands are clearly resolved in the latter region at 1651 and 1659 cm−1. Both are apparently associated with α‐helices. These results suggest that for D2O solutions of globular proteins, FTIR spectroscopy can be a facile method for detecting the presence of these two different types of helical conformation and distinguishing between them. This provides a distinct advantage over ultraviolet circular dichroism spectroscopy (UV‐CD). This work also provides a basis for future studies of α‐lactalbumin which examine the effects of environment (e.g., pH, temperature) and ligands (e.g., Ca2+, Mn2+) on its conformation.