Absolute and relative binding free energy calculations of the interaction of biotin and its analogs with streptavidin using molecular dynamics/free energy perturbation approaches

Abstract We present calculations of the absolute and relative binding free energies of complexation of streptavidin with biotin and its analogsby means of a thermodynamic free energy perturbation method implemented with molecular dynamics. Using the recently solved crystal structure of the streptavidin–biotin complex, biotin was mutated into a dummy molecule as well as thiobiotin and iminobiotin both in the protein and insolution. The calculated absolute binding free energy was dependent on the simulation model used. Encouragingly, the “best models” provided a reasonable semiquantitative reproduction (−20 to −22 kcal/mol) of the experimental free energy (−18.3 kcal/ mol). Furthermore, the calculated results give clear insights into the binding nature of the protein–ligand complex, showing that the van der Waals energy dominates the electrostatic and hydrogen bonding energies in thebinding of biotin by streptavidin. Specifically, the mutation of biotin into a dummy molecule in solution has a Δ G (van der Waals) ∼ −4 kcal/mol, due to the cancellation of dispersion and repulsion “cavity” effects. On the other hand, in the protein, a very small free energy price must be paid to create a cavity since one already exists and the mutation of biotin into a dummy molecule has a Δ G (van der Waals) ∼ 15 kcal/mol. These results are also consistent with the interpretation that the entropy increase to be expected from hydrophobic interactions from desolvation of biotin is counterbalanced by a decrease in entropy accompanying the formationof buried hydrogen bonds, which have been derived from the apparentlyconflicting experimental data. They provide an alternative interpretationofthe reason for the extremely high affinity of the biotin‐streptavidin interaction than that recently proposed by Weber et al. (J. Am. Chem. Soc. 114:3197, 1992). In the case of the relative binding freeenergies, the calculated values of 3.8 ± 0.6 and 7.2 ± 0.6 kcal/mol compare well with the experimental values of 3.6 and 6.2 kcal/mol for the perturbation of biotin to thiobiotin and iminobiotin, respectively in the related protein avidin. The calculations indicate that desolvation of the ligand is important in understanding the relative affinity of the ligands with the protein. The above successful simulations suggestthat the molecular dynamics/free energy perturbation method is useful for understanding the energetic features affecting the binding between proteins and ligands, since it is generally difficult to determine these factors unambiguously by experiment. This set of studies provide a textbook example of the key elements of protein–ligand recognition: the electrostatic free energy dominates the relative affinities, the van der Waals free energy dominates the absolute free energy; the free energy of desolvation is a key to why iminobiotin is so much more weakly bound than biotin and the free energy of binding explains why thiobiotin is so weakly bound relative to biotin. © 1993 Wiley‐Liss, Inc.