Effect of temperature and pH on the structure and stability of tumor-specific lectin jacalin and insights into the location of its tryptophan residues: CD, DSC and fluorescence studies

Jacalin, the jackfruit seed lectin, exhibits high specificity for the tumor-specific T-antigen and is used in various biomedical and biotechnological applications. Here, we report biophysical studies on the thermal unfolding of jacalin and the effect of pH and temperature on its secondary structure. Differential scanning calorimetric (DSC) studies revealed that native jacalin unfolds at ∼60 °C and that carbohydrate binding stabilizes the protein structure. Circular dichroism spectroscopic studies indicated that the secondary structure of jacalin remains mostly unaffected over pH 2.0–9.0, whereas considerable changes were observed in the tertiary structure. DSC experiments demonstrated that jacalin exhibits two overlapping transitions between pH 2 and 5, which could be attributed to dissociation of the tetrameric protein into subunits and their unfolding. Interestingly, only one transition between pH 6 and 9 was observed, suggesting that the subunit dissociation and unfolding occur simultaneously. While quenching of the protein intrinsic fluorescence by acrylamide increased significantly upon carbohydrate binding, quenching by succinimide is essentially unaffected. We attribute this difference to increased exposure of Trp-123 in the α-chain as it is involved in carbohydrate binding. Both acrylamide and succinimide gave biphasic Stern-Volmer plots, consistent with differential accessibility of the two tryptophan residues of jacalin to them. [Display omitted] •Secondary structure of jacalin is stable over a wide range of pH (2–9) and temperature.•The unfolding temperature of jacalin increases from 60 °C to 63 °C upon sugar binding.•Two unfolding transitions of jacalin at pH 2–5 are assigned to subunit dissociation and unfolding.•A single transition is observed at pH 6–9 indicating simultaneous subunit dissociation and unfolding.•The two Trp residues of jacalin show different accessibilities to acrylamide and succinimide.