Changes in Protein Conformation and Surface Hydrophobicity upon Peroxidase-Catalyzed Cross-Linking of Apo-α-lactalbumin

In this study, we explore the effect of peroxidase-catalyzed cross-linking on the molecular conformation of apo-alpha-lactalbumin (apo-alpha-LA) and the resulting changes in protein surface hydrophobicity. In studying conformational changes, we distinguish between early stages of the reaction (partial cross-linking), in which only protein oligomers (10(6) Da > M-w, >= 10(4) Da) are formed, and a later stage (full cross-linking), in which larger protein particles (M-w >= 10(6) Da) are formed. Partial cross-linking induces a moderate loss of alpha-helical content. Surprisingly, further cross-linking leads to a partial return of a-helices that are lost upon early cross-linking. At the same time, for partially and fully cross-linked apo-alpha-LA, almost all tertiary structure is lost. The protein surface hydrophobicity first increases for partial cross-linking, but then decreases again at full cross-linking. Our results highlight the subtle changes in protein conformation and surface hydrophobicity of apo-alpha-LA upon peroxidase-catalyzed cross-linking.