Transient non-native secondary structures during the refolding of alpha-lactalbumin detected by infrared spectroscopy.

Stopped-flow Fourier-transform infrared spectroscopy (SF-FTIR) was used to identify native as well as non-native secondary structures during the refolding of the calcium-binding protein alpha-lactalbumin. Infrared absorbance spectra were recorded in real time after a pH jump induced refolding of the protein. In the presence of calcium, the refolding is fast with concerted appearance of secondary structures; in its absence, folding is much slower and intricate, with transient formation and disappearance of non-native beta-sheet. The possibility of detecting native as well as non-native structures at the same time is especially valuable in providing insight into the complexity of the refolding process of a protein.