Eva Dejmková 1, Martina Pokorná 1, Michaela Wimmerová1,2


1National Centre for Biomolecular Research and 2Department of Biochemistry, Masaryk University, Kotlářská 2, 611-37 Brno, Czech Republic, E-mail:


Lectins are proteins or glycoproteins of non-immune origin capable agglutinate cells. They are able to specifically recognize saccharides with high affinity. However, the major function of lectins appears to be in the cell recognition process, they generally recognize diverse sugar structures and mediate a variety of biological processes. Bacterial lectins play a crucial role in recognition of sugar moieties on the host cell surface and consequently can cause bacterial adhesion. An attractive approach is the use of agents that interfere with the ability of the bacteria to adhere to the host cell surface, since such adhesion is one of the initial stages of the infectious process. However, the process of ligand binding is very complex and complicated, deeper understanding of the thermodynamics of lectin-saccharide interaction can be useful for the molecular design of potent anti-adhesion therapeutics.

Chromobacterium violaceum is an opportunistic pathogen that commonly occurs in water and soil in tropical and subtropical regions. Infection caused by this bacterium can be fatal for immunocompromised people and children. Bacterial unusual resistance to antibiotics is the reason of high mortality.

The lectin from C. violaceum named CV-IIL is a tetramer (the structural functional unit is a dimer), each monomer is composed of 113 amino acids. This lectin shows ability to bind L-fucose (6-deoxy-L-galactose) and D-mannose with high affinity.  Each monomer contains two calcium ions in the carbohydrate binding site, which mediate binding of the saccharides. The crystal structure of CV-IIL demonstrates that there is also one water molecule, which plays a special role by bridging sugar with backbone nitrogen atom and also with side chain of the amino acid threonine in position 97. (Fig.1) [1] Therefore, several representatives from different groups of amino acids were chosen for the mutagenesis in this position 97.

The complete thermodynamical analysis was performed using isothermal titration microcalorimetry method (ITC 200, Microcal). From the single measurement we can obtain and consequently calculate important thermodynamical values (enthalpy, entropy, Gibbs free energy) as well as binding affinity and stoichiometry of the lectin-saccharide interaction. The interaction with high affinity ligand α-Me-fucoside was measured in case of each prepared mutant lectin at different temperatures in the range from 10 to 40°C at constant pressure. These detail measurements allow the determination of the heat capacity ∆Cp - the slope of the enthalpy dependence on temperature. Accurate values of ∆Cp for individual mutant allow partial discrimination of the entropy change associated with the solvent release and the loss of configurational entropy. The total entropy of binding can be devided into terms for changes in solvation and losses in configurational, rotational and translational entropy.

∆S° = ∆S solv + ∆S config + ∆S rot + ∆S transl [2]

The two major contributions to the binding entropy are the change in conformational and the solvation entropy, which will be discussed. However, the values of the Gibbs free energy are very similar, the difference in enthalpy and entropy contributions were marked in some cases.


The work is supported by Ministry of Education (MSM0021622413, ME08008) and Czech Science Foundation (GA/303/09/1168) of the Czech Republic


1.     Pokorná M., Cioci G., Perret S, Rebuffet E., Kostlánová N., Adam J., Gilboa-Garber N., Mitchell E.P., Imberty A., Wimmerová M., Unusual Entropy-Driven Affinity of Chromobacterium violaceum Lectin CV-IIL toward Fucose and Mannose, Biochemistry         45 (24) 7501 - 7510, 2006

2.     Chervenak M.C., Toone E.J., Calorimetric Analysis of the Binding of Lectin with      Overlapping Carbohydrate-Binding Ligand Specifities, Biochemistry 34 5685 - 5695,             1995