The carbohydrate binding protein, Cyanovirin-N, obtained from cyanobacteria, consists of high-affinity and low-affinity binding domains. To avoid the formation of a domain swapped structure in solution and also to better focus on the binding of carbohydrates at the high-affinity site, the Ghirlanda group (Biochemistry, 46, 2007, 9199-9207) engineered the P51G-m4-CVN mutant which does not dimerize nor binds at the low-affinity site. This mutant provides an excellent starting point for the experimental and computational study of further transformations to enhance binding at the high-affinity site as well as to retool this site for the possible binding of different sugars. However, before such endeavors are pursued, detailed understanding of apparently key interactions both present in wild-type and P51G-m4-CVN at the high-affinity site must be derived and controversies about the importance of certain residues must be resolved. One such interaction is that of Glu41, a charged residue in intimate contact with 2'OH of dimannose at the nonreducing end. We do so computationally by performing two mutations using the thermodynamic integration formalism in explicit solvent. Mutations of P51G-m4-CVN Glu41 to Ala41 and Gly41 reveal that whereas the loss of Coulomb interactions result in a free energy penalty of about 2.1 kcal/mol, this is significantly compensated by favorable contributions to the Lennard-Jones portion of the transformation, resulting in almost no change in the free energy of binding. At least in terms of free energetics, and in the case of this particular CVN mutant, Glu41 does not appear to be as important as previously thought. This is not because of lack of extensive hydrogen bonding with the ligand but instead because of other compensating factors.