Thermal and pH-induced conformational changes of a beta-sheet protein monitored by infrared spectroscopy.

The stability of a lentil lectin, an all-beta protein, has been perturbed by changes in pH and temperature. In the pH interval 5.0 --> 10.0, the overall secondary structure does not undergo significant changes. However, if the individual components of the infrared amide I band are considered, changes in band components attributed to variations in beta-sheet and beta-turns cross-interactions are detected. The combined effects of pH and temperature reveal that the protein is more compact at pH 7.5 with lower denaturation temperatures at pH 5.0 or 10.0, indicating a less stable protein under those conditions. According to our results, the structural stability of the beta-sheet would depend not only on the intermolecular interactions among the strands but also on the conformation of the segments connecting these strands. The protein infrared band assignment has also been examined since the three-dimensional structure of the lentil lectin protein is known from X-ray diffraction studies. Two of the bands observed are attributed to beta-sheet. The one at 1620 cm-1, not affected if the medium is deuterated, is assigned to hairpins composed by two strands connected by a rigid turn whereas that located at 1633 cm-1 corresponds to strands associated by more flexible segments. The band appearing at 1645 cm-1 in H2O corresponds to the open, flexible loops that are connecting the beta-strands. The simplest assumption of the various secondary structure components having identical IR extinction coefficients is enough to provide IR-derived data that are in good agreement with the structure solved by X-ray diffraction.