Crystallographic and computational methods have been used to study the binding of two monosaccharides (glucoside and mannoside) to concanavalin-A. The 2 structure of glucoside bound concanavalin-A is reported and compared with the 2 Ó Ó structure of the mannoside complex. The interaction energies of the substrate in each crystallographic subunit were calculated by molecular mechanics and found to be essentially the same for both sugars. Further energy minimisation of the active site region of the subunits did not alter this conclusion. Information from crystallographic B-factors was interpreted in terms of mobility of the sugars in the combining site. Molecular dynamics (MD) was employed to investigate mobility of the ligands at the binding sites. Switching between di †erent binding states was observed for mannoside over the ensemble in line with the crystallographic B-factors. A calculated average interaction energy was found to be more favourable for mannoside than glucoside, by 4.9 ^3.6 kcal mol~1 (comparable with the experimentally determined binding energy di †erence of 1.6 ^0.3 kcal mol~1). However, on consideration of all terms contributing to the binding enthalpy a di †erence is not found. This work demonstrates the difficulty in relating structure to thermodynamic properties, but suggests that dynamic models are needed to provide a more complete picture of ligandÈreceptor interactions.

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Molecular dynamics (MD) simulations of Erythrina corallodendron lectin binding to a monosaccharide, a-galactose, and a disaccharide, N-acetyl lactosamine, have been performed in order to investigate the relationship between structure and thermodynamics.A simulated annealing protocol has been used to generate ensembles of structures for the two complexes, from which both qualitative and quantitative information on binding dynamics have been extracted. The ensembled averaged lectin±saccharide interaction enthalpy is equivalent for both sugars, whereas the calculation based on the X-ray structures does show a difference. Within large statistical errors, the calculated`binding enthalpy' is also the same for the two systems. These errors arise largely from terms involving solvent and are a typical limitation of current MD simulations.Significant qualitative differences in binding between the two complexes are, however, observed over the ensembles. These could be important for unraveling the structure/thermodynamic relationship. Stated simply, there are a greater number of binding options available to the disaccharide compared to the monosaccharide. The implications of alternative binding states on thermodynamic parameters and the`breaking of enthalpy±entropy compensation' are discussed. The role of solvent in lectin±saccharide complex formation is suggested to be significant.Keywords: lectin±saccharide; structure; thermodynamics; Erythrina corallodendron lectin; molecular dynamics.Many important biological processes, both benign and malign, are based upon protein±carbohydrate recognition [1]. Legume lectins [2] are excellent model systems for the study of protein± saccharide interactions because they are abundant, easy to purify and specifically recognize a variety of complex carbohydrates [3]. The numbers of crystallographically determined legume lectin±saccharide complexes continues to grow each year [4] and in addition, calorimetric studies have provided information on the thermodynamics of complex formation for many of these complexes [5±9]. Despite these increases in data, the relationship between the structure of protein±saccharide complexes and the corresponding thermodynamic data remains to be elucidated. This problem is general to all receptor±ligand interactions and is a key question in molecular biophysics [10±12]. Undoubtedly, the solution of this problem would have an enormous impact in the field of structure-based drug design. A major goal is the ability to predict the free energy of binding for any given ligand/receptor pair [13,14]. Less ambitious is to explain why one ligand should bind more tightly than another to a certain receptor. It is also generally found for lectin± saccharide complex formation that a more favoured free energy of binding (DG bind ) for a given sugar compared to others, is a result of a more favoured enthalpy value (DH bind ) despite some compensation by a less favoured entropy [11]. Thus, one could further simplify an analysis by considering enthalpies rather than free energi...

Peanut lectin binds T-antigen [Galbeta(1-3)GalNAc] with an order of magnitude higher affinity than it binds the disaccharide lactose. The crystal structures of the two complexes indicate that the higher affinity for T-antigen is generated by two water bridges involving the acetamido group. Fresh calorimetric measurements on the two complexes have been carried out in the temperature range 280-313 K. Four sets of nanosecond molecular-dynamics (MD) simulations, two at 293 K and the other two at 313 K, were performed on each of the two complexes. At each temperature, two somewhat different protocols were used to hydrate the complex in the two runs. Two MD runs under slightly different conditions for each complex served to assess the reliability of the approach for exploring protein-ligand interactions. Enthalpies based on static calculations and on MD simulations favour complexation involving T-antigen. The simulations also brought to light ensembles of direct and water-mediated protein-sugar interactions in both the cases. These ensembles provide a qualitative explanation for the temperature dependence of the thermodynamic parameters of peanut lectin-T-antigen interaction and for the results of one of the two mutational studies on the lectin. They also support the earlier conclusion that the increased affinity of peanut lectin for T-antigen compared with that for lactose is primarily caused by additional water bridges involving the acetamido group. The calculations provide a rationale for the observed sugar-binding affinity of one of the two available mutants. Detailed examination of the calculations point to the need for exercising caution in interpreting results of MD simulations: while long simulations are not possible owing to computational reasons, it is desirable to carry out several short simulations with somewhat different initial conditions.