Dual Specificity PDZ- and 14-3-3-Binding Motifs: A Structural and Interactomics Study

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• Post-translational modifications, particularly phosphorylation, significantly impact protein-protein interactions.
• A substantial portion of human PDZ domain-binding motifs (PBMs) are potentially phosphorylatable, suggesting widespread functional regulation.
• Phosphorylation can alter the binding affinities of PBMs to PDZ domains and 14-3-3 proteins, thereby rewiring cellular interactomes.

Purpose of the Study:

• To investigate the impact of phosphorylation and phosphomimetic mutations on the binding affinities of specific PBMs.
• To characterize the structural consequences of phosphorylation and phosphomimetic modifications on PBM-PDZ domain interactions.
• To elucidate the structural mechanisms underlying PBM recognition by 14-3-3 proteins.

Main Methods:

• Measurement of binding affinities for native, phosphorylated, and phosphomimetic PBM variants against 266 human PDZ domains.
• Co-crystallization of PBM variants with a selected PDZ domain to determine structural changes.
• Structural elucidation of PBM interactions with 14-3-3 proteins.

Main Results:

• Differential binding affinities were observed between native, phosphorylated, and phosphomimetic PBM variants and various PDZ domains.
• Structural analysis revealed distinct conformational changes induced by phosphorylation and phosphomimetic mutations.
• The study identified specific structural features governing PBM recognition by 14-3-3 proteins.

Conclusions:

• Phosphorylation and phosphomimetic approaches can differentially affect PBM-mediated protein-protein interactions.
• Atomic and interactomic insights were gained into the regulation of dual specificity motifs by phosphorylation.
• Understanding these modifications is crucial for comprehending interactome dynamics and potential therapeutic strategies.