Molecular Basis of Class III Ligand Recognition by PDZ3 in Murine Protein Tyrosine Phosphatase PTPN13

Abstract

Protein tyrosine phosphatase PTPN13, also known as PTP-BL in mice, represents a large multi-domain non-transmembrane scaffolding protein that contains five consecutive PDZ domains. Here, we report the solution structures of the extended murine PTPN13 PDZ3 domain in its apo form and in complex with its physiological ligand, the carboxy-terminus of protein kinase C-related kinase-2 (PRK2), determined by multidimensional NMR spectroscopy. Both in its ligand-free state and when complexed to PRK2, PDZ3 of PTPN13 adopts the classical compact, globular D/E fold. PDZ3 of PTPN13 binds five carboxy-terminal amino acids of PRK2 via a groove located between the EB-strand and the DB-helix. The PRK2 peptide resides in the canonical PDZ3 binding cleft in an elongated manner and the amino acid side chains in position P0 and P-2, cysteine and aspartate, of the ligand face the groove between EB-strand and DB-helix, whereas the PRK2 side chains of tryptophan and alanine located in position P-1 and P-3 point away from the binding cleft. These structures are rare examples of selective class III ligand recognition by a PDZ domain and now provide a basis for the detailed structural investigation of the promiscuous interaction between the PDZ domains of PTPN13 and their ligands. They will also lead to a better understanding of the proposed scaffolding function of these domains in multi-protein complexes assembled by PTPN13 and could ultimately contribute to low molecular weight antagonists that might even act on the PRK2 signaling pathway to modulate rearrangements of the actin cytoskeleton.

The binding affinity of PTPN13’s tandem PDZ2/3 domain is allosterically modulated

“…The NMR chemical shift perturbations observed in 2D 1 H- 15 N-HSQC spectra for the PDZ2-PRK2 complex demonstrate that, in comparison to PDZ3 (K D = 318 ± 47 μM), the PRK2 peptide only weakly binds to the PDZ2 domain (K D = 661 ± 71 μM) (Fig. 1a, b, c) [12]. In particular, NMR chemical shift differences were observed for the canonical ligand binding groove of PDZ2 including G23, S24, which are also involved in peptide recognition, and S28, and G31.…”

“…Averaged dissociation constants for the different complexes were calculated from least-squares fitting of the NMR chemical shift perturbations observed for several amino acids as a function of ligand concentration. NMR chemical shift perturbation data were analysed with the ORIGIN software package (www.originlab.com) as previously published [12, 52]. The error for K D values was calculated from the individual fit of several significant chemical shift perturbations found for the amide resonances located in and/or next to the canonical PDZ binding cleft.…”

“…Here, the carboxylate moiety of the last ligand residue, i.e. P 0 , is bound to GLGF motif of PDZ domains, which causes ligand amino acid side chains of P 0 and P − 2 to be orientated towards the PDZ binding pocket and side chains in position P − 1 and P − 3 to point away from the PDZ binding cleft into the solvent [12]. Recently, more general concepts have been introduced, which do not (entirely) rely on specific definitions of PDZ ligand classes [13, 14].…”

Conformational flexibility of GRASPs and their constituent PDZ subdomains reveals structural basis of their promiscuous interactome

“…For instance, the modified binding preference of PTPN13 PDZ2 domain for APC could be explained by an allosteric modulation induced by the interaction of PDZ3 with PDZ2 [41]. Moreover, the potential of PTP-BL PDZ2 domain to interact with class III ligands was found to be modulated by the presence of PDZ1, suggesting that the PDZ microenvironment is determinant for their ligand binding specificity [42,43]. This allosteric control seems to take place even with recent evidence suggesting that PDZ-tandem protein have structurally independent PDZs within the full-length scaffold protein [44].…”

The order of PDZ3 and TrpCage in fusion chimeras determines their properties—a biophysical characterization

“…and act as a scaffold domain mediating the formation of intracellular multiprotein complexes. [11][12][13][14] PDZ domains possess a very broad specificity for various substrates, 15 and the specificity of the PDZ-ligand binding pocket has been shown to be modulated via intramolecular allosteric networks. [16][17][18] Therefore, it could be expected that the dynamics of PDZ is highly dependent on the domains in its vicinity.…”