Thermal Stability and Mode of Oligomerization of the Tetrameric Peanut Agglutinin: A Differential Scanning Calorimetry Study

Reddy, Bhanuprakash G and Bharadwaj, Satish and Surolia, Avadhesha (1999) Thermal Stability and Mode of Oligomerization of the Tetrameric Peanut Agglutinin: A Differential Scanning Calorimetry Study. In: Biochemistry, 38 (14). pp. 4464-4470.

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Abstract

Peanut agglutinin is a homotetrameric legume lectin. The crystal structure of peanut agglutinin shows that the four subunits associate in an unusual manner, giving rise to open quaternary structure [Banarjee, R., et al. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 227-231]. The thermal unfolding of peanut agglutinin has been characterized by differential scanning calorimetry and gel filtration to elucidate its thermal stability and its mode of oligomerization. The unfolding process is reversible and could be described by a three-state model with two transitions occurring at around 331 and 336 K. For the tetramer, the ratio of $\Delta H_c/ \Delta H_v$ for the first transition is close to 4 and for the second transition is close to 0.25, suggesting that 4 and 0.25 cooperative unit(s) of the tetramer are involved in the first and second transitions, respectively. The agreement between the model-independent$ \Delta H_v(S)$ determined from the values of the temperatures of the peak maximum$(T_{p1})$ with the protein concentration with the values of $\Delta H_v$ obtained from the fit of the data to the transition confirms that the first peak is associated with the dissociation of peanut agglutinin tetramers $(A_4)$ to "folded" monomers (4A), whereas the second peak describes the unfolding (4U) of these monomers. The overall process for the thermal unfolding of peanut agglutinin could therefore be summarized as $A_4\leftrightarrow 4A\leftrightarrow 4 U $. Gel filtration studies confirm this process, as peanut agglutinin elutes as a tetramer up to $50^oC$, and at and above $56^oC$ ($T_m$ of first transition), it elutes at a position commensurate with that of the folded monomer of peanut agglutinin. The unfolding behavior of peanut agglutinin in the presence of saturating amounts of carbohydrate ligands is similar to that observed for the unligated form. The temperature of maximal stability of the peanut agglutinin tetramer at pH 7.4 is calculated to be around $33^oC$ with a maximal free energy of stabilization of 8.70 kcal/mol. The results demonstrate that unfolding of peanut agglutinin goes through two distinct phases with folded monomer being the intermediate.

Item Type:	Journal Article
Publication:	Biochemistry
Publisher:	American Chemical Society(ACS)
Additional Information:	Copyright of this article belongs to American Chemical Society(ACS).
Department/Centre:	Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited:	04 Oct 2007
Last Modified:	19 Sep 2010 04:40
URI:	http://eprints.iisc.ac.in/id/eprint/12068

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