Characterization of PDZ domain–peptide interactions using an integrated protocol of QM/MM, PB/SA, and CFEA analyses

Protein–protein interactions, particularly weak and transient ones, are often mediated by peptide recognition domains. Characterizing the interaction interface of domain–peptide complexes and analyzing binding specificity for modular domains are critical for deciphering protein–protein interaction networks. In this article, we report the successful use of an integrated computational protocol to dissect the energetic profile and structural basis of peptide binding to third PDZ domain (PDZ3) from the PSD-95 protein. This protocol employs rigorous quantum mechanics/molecular mechanics (QM/MM), semi-empirical Poisson–Boltzmann/surface area (PB/SA), and empirical conformational free energy analysis (CFEA) to quantitatively describe and decompose systematic energy changes arising from, respectively, noncovalent interaction, desolvation effect, and conformational entropy loss associated with the formation of 30 affinity-known PDZ3–peptide complexes. We show that the QM/MM-, PB/SA-, and CFEA-derived energy components can work together fairly well in reproducing experimentally measured affinity after a linearly weighting treatment, albeit they are not compatible with each other directly. We also demonstrate that: (1) noncovalent interaction and desolvation effect donate, respectively, stability and specificity to complex architecture, while entropy loss contributes modestly to binding; (2) P0 and P−2 of peptide ligand are the most important positions for determining both the stability and specificity of the PDZ3–peptide complex, P−1 and P−3 can confer substantial stability (but not specificity) for the complex, and N-terminal P−4 and P−5 have only a very limited effect on binding.