Thermal, chemical and pH induced unfolding of turmeric root lectin: modes of denaturation

PLoS One. 2014 Aug 20;9(8):e103579. doi: 10.1371/journal.pone.0103579. eCollection 2014.

Abstract

Curcuma longa rhizome lectin, of non-seed origin having antifungal, antibacterial and α-glucosidase inhibitory activities, forms a homodimer with high thermal stability as well as acid tolerance. Size exclusion chromatography and dynamic light scattering show it to be a dimer at pH 7, but it converts to a monomer near pH 2. Circular dichroism spectra and fluorescence emission maxima are virtually indistinguishable from pH 7 to 2, indicating secondary and tertiary structures remain the same in dimer and monomer within experimental error. The tryptophan environment as probed by acrylamide quenching data yielded very similar data at pH 2 and pH 7, implying very similar folding for monomer and dimer. Differential scanning calorimetry shows a transition at 350.3 K for dimer and at 327.0 K for monomer. Thermal unfolding and chemical unfolding induced by guanidinium chloride for dimer are both reversible and can be described by two-state models. The temperatures and the denaturant concentrations at which one-half of the protein molecules are unfolded, are protein concentration-dependent for dimer but protein concentration-independent for monomer. The free energy of unfolding at 298 K was found to be 5.23 Kcal mol-1 and 14.90 Kcal mol-1 for the monomer and dimer respectively. The value of change in excess heat capacity upon protein denaturation (ΔCp) is 3.42 Kcal mol-1 K-1 for dimer. The small ΔCp for unfolding of CLA reflects a buried hydrophobic core in the folded dimeric protein. These unfolding experiments, temperature dependent circular dichroism and dynamic light scattering for the dimer at pH 7 indicate its higher stability than for the monomer at pH 2. This difference in stability of dimeric and monomeric forms highlights the contribution of inter-subunit interactions in the former.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Calorimetry, Differential Scanning
  • Curcuma / chemistry*
  • Hydrogen-Ion Concentration
  • Lectins / chemistry*
  • Models, Molecular*
  • Plant Roots / chemistry*
  • Protein Denaturation*
  • Protein Folding*
  • Temperature
  • Thermodynamics

Substances

  • Lectins

Grants and funding

All funding for this study received from Bose Institute, which in turn receives funding from the Department of Science and Technology, Government of India. The funder provided support in the form of salaries for authors [HB,RC], but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section.