Spectroscopic and thermodynamic evidence for a complex denaturation mechanism of bovine β-lactoglobulin A

Abstract

We present a spectroscopic and calorimetric study on the thermal denaturation of bovine β-lactoglobulin (β-lg) variant A. Spectroscopic data allowed detection of a stable intermediate emerging from structural modifications restricted to local regions of the native molecule. It is suggested that this kind of intermediate could be a common property of lipocalins. Using the same set of parameters that has successfully related thermodynamics and structural properties of other proteins, it is shown that the thermally denatured state of β-lg retains a significant amount of buried hydrophobic surface area. Thus, despite being a small protein composed of a single structural domain, β-lg exhibits a complex unfolding mechanism, comprising at least two other species different from the native and completely unfolded states.