title: Domain orientation in the N-Terminal PDZ tandem from PSD-95 is maintained in the full-length protein.
abstract: Tandem PDZ domains have been suggested to form structurally independent supramodules. However, dissimilarity between crystallography and NMR models emphasize their malleable conformation. Studies in full-length scaffold proteins are needed to examine the effect of tertiary interactions within their native context. Using single-molecule fluorescence to characterize the N-terminal PDZ tandem in PSD-95, we provide the first direct evidence that PDZ tandems can be structurally independent within a full-length scaffold protein. Molecular refinement using our data converged on a single structure with an antiparallel alignment of the ligand-binding sites. Devoid of interaction partners, single-molecule conditions captured PSD-95 in its unbound, ground state. Interactions between PDZ domains could not be detected while fluctuation correlation spectroscopy showed that other conformations are dynamically sampled. We conclude that ultra-weak interactions stabilize the conformation providing a "low-relief" energy landscape that allows the domain orientation to be flipped by environmental interactions.
authors.affiliation: Department of Pharmacological Sciences, Stony Brook University, Stony Brook, NY 11794, USA.
authors.forename: James J
authors.initials: JJ
authors.lastname: McCann
authors.forename: Liqiang
authors.initials: L
authors.lastname: Zheng
authors.forename: Salvatore
authors.initials: S
authors.lastname: Chiantia
authors.forename: Mark E
authors.initials: ME
authors.lastname: Bowen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003545:Cysteine / Q000737:chemistry; D000076122:Disks Large Homolog 4 Protein; D031541:Fluorescence Resonance Energy Transfer; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry* / Q000235:genetics; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics; D056004:Molecular Dynamics Simulation; D010446:Peptide Fragments / Q000737:chemistry; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics
chemical_list: D000076122:Disks Large Homolog 4 Protein; C501449:Dlg4 protein, rat; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D003545:Cysteine