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Proteome‐wide analysis of phospho‐regulated PDZ domain interactions
- Source :
- Molecular Systems Biology
- Publication Year :
- 2018
-
Abstract
- A key function of reversible protein phosphorylation is to regulate protein–protein interactions, many of which involve short linear motifs (3–12 amino acids). Motif‐based interactions are difficult to capture because of their often low‐to‐moderate affinities. Here, we describe phosphomimetic proteomic peptide‐phage display, a powerful method for simultaneously finding motif‐based interaction and pinpointing phosphorylation switches. We computationally designed an oligonucleotide library encoding human C‐terminal peptides containing known or predicted Ser/Thr phosphosites and phosphomimetic variants thereof. We incorporated these oligonucleotides into a phage library and screened the PDZ (PSD‐95/Dlg/ZO‐1) domains of Scribble and DLG1 for interactions potentially enabled or disabled by ligand phosphorylation. We identified known and novel binders and characterized selected interactions through microscale thermophoresis, isothermal titration calorimetry, and NMR. We uncover site‐specific phospho‐regulation of PDZ domain interactions, provide a structural framework for how PDZ domains accomplish phosphopeptide binding, and discuss ligand phosphorylation as a switching mechanism of PDZ domain interactions. The approach is readily scalable and can be used to explore the potential phospho‐regulation of motif‐based interactions on a large scale.
- Subjects :
- Proteome
PDZ domain
Oligonucleotides
Method
PDZ Domains
Methods & Resources
Ligands
Scribble
Peptide Library
Protein Interaction Mapping
Methods
Humans
Amino Acid Sequence
Protein Interaction Maps
Binding Sites
phosphorylation
Biochemistry and Molecular Biology
Post-translational Modifications, Proteolysis & Proteomics
PDZ domainphage displayphosphorylationprotein–protein interactionScribble
protein–protein interaction
ddc:540
Zonula Occludens-1 Protein
phage display
Peptides
Disks Large Homolog 4 Protein
Biokemi och molekylärbiologi
Protein Binding
Subjects
Details
- Language :
- English
- Database :
- OpenAIRE
- Journal :
- Molecular Systems Biology
- Accession number :
- edsair.pmid.dedup....4b78726af2237ad38a869ab900f28427