Table 3.

Na⁺/K⁺ selectivity in altered binding sites (kcal/mol)

	Site NA1	Site NA2
Fully Flexible Proteina	6.11	3.22
Frozen protein in a K+ adapted conformationb	−0.11	0.81
Partially frozen protein in a K+ adapted conformationc	4.27	2.11

^aNo constrains were applied on the system during molecular dynamics simulation (T=315 K).

^bNa⁺ ions were replaced by potassium in binding sites followed by energy minimization to insure optimal K+ coordination in binding sites NA1 and NA2, minimized system was fixed during free energy simulations.

^cSame as b, but ligands in the first coordination shell of the ion were free to move.