N-terminus of the third PDZ Domain of PSD-95 Orchestrates Allosteric Communication for Selective Ligand Binding

Abstract

PDZ domains constitute common models to study single domain allostery without significant structural changes. The third PDZ domain of PSD-95 (PDZ3) is known to have selective structural features that confer unique modulatory roles to this unit. In this model system two residues, H372 directly connected to the binding site and G330 holding an off-binding-site position, were designated to assess the effect of mutations on binding selectivity. It has been observed that the H372A and G330T/H372A mutations change ligand preferences from class I (T/S amino acid at position −2 of the ligand) to class II (hydrophobic amino acid at the same position). Alternatively, the G330T single mutation leads to the recognition of both ligand classes. We have performed a series of molecular dynamics (MD) simulations for wild-type, H372A, G330T single mutants and a double mutant of PDZ3 in the absence and presence of both types of ligands. With the combination of free energy difference calculations and a detailed analysis of MD trajectories, ‘class switching’ and ‘class bridging’ behavior of PDZ3 mutants, as well as their effects on ligand selection and binding affinities are explained. We show that the dynamics of the charged N-terminus plays a fundamental role in determining the binding preferences in PDZ3 by altering the electrostatic energy. These findings are corroborated by simulations on N-termini truncated versions of these systems. The dynamical allostery orchestrated by the N-terminus offers a fresh perspective to the study of communication pathways in proteins.

Dynamic Community Composition Unravels Allosteric Communication in PDZ3

“…As a result, the flexible N-terminus interacts with the ligand and affects binding specificity, significantly for WT and single mutations. We have shown previously 20 The analyses of the remaining segments do not provide information on binding specificity of PDZ3. For example, the C-terminus does not communicate with structural segments other than the N-terminus.…”

“…35,36 To understand the modularity of PDZ3, the communication between the N/C termini, α2, α3 and the ligand is assessed. 7,9,20,22 Although, node centrality measures are informative to the extent of pinpointing residues whose centrality are shifted depending on the variant studied ( Figure 2), our community composition analysis is more sensitive (Tables 2 and 3). We find that PDZ3 complex variants have diverse community configurations, and the fraction of these changes modulates binding preferences.…”

“…The MD simulations and their detailed energetic analyses have recently been reported elsewhere. 20 Briefly, protein structures are downloaded from the protein data bank (PDB) 38 (Table 1), and missing residues of ligand are inserted by using SWISS-MODEL 39 server. Equal lengths of the structures are used whereby residue index ranges are 299-415 for the protein and 2-9 for the ligand (see Fig.…”

Dynamic Community Composition Unravels Allosteric Communication in PDZ3

“…To understand the molecular basis of binding, we investigate the previously studied structural segments, which are the N and C termini, α2 helix and the aforementioned α3 helix (Figure 1a). 5,[9][10][11][20][21][22] The dynamics of the highly charged N-terminus region (Figure 1a) has recently been shown to be important in the binding mechanism, especially due to its electrostatic contributions to the total free energy. 20 The effect of the N-terminus might be partnered with that of the C-terminus.…”

“…5,[9][10][11][20][21][22] The dynamics of the highly charged N-terminus region (Figure 1a) has recently been shown to be important in the binding mechanism, especially due to its electrostatic contributions to the total free energy. 20 The effect of the N-terminus might be partnered with that of the C-terminus. 21 α2 helix (Figure 1a) lines up the ligand, and its effect has been investigated by deep mutational scan.…”