pH-induced conformational transitions in α-lactalbumin investigated with two-dimensional Raman correlation variance plots and moving windows

Raman spectroscopy is being increasingly applied to the investigation of conformational transitions in polypeptides and proteins, and in particular protein unfolding, due to its ability to monitor changes in secondary structure. Here we focus on the application of two different two-dimensional correlation methods, autocorrelation and moving windows techniques, to investigate the pH-induced conformational transitions in α-lactalbumin monitored with Raman scattering. From our results we have identified three distinct stages in the conformational transition occurring at ranges ∼pH 6.5–4.6, ∼pH 4.6–3.6 and ∼pH 3.6–1.8, suggesting the existence of three possible conformational species. The first two transition phases the conformational changes are predominantly occurring in the backbone secondary structure, while in the final transition phase changes are occurring in both secondary structure and side chain residue solvent exposure.