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Canonical and noncanonical sites determine NPT2A binding selectivity to NHERF1 PDZ1

Mamonova, T and Zhang, Q and Khajeh, JA and Bu, Z and Bisello, A and Friedman, PA (2015) Canonical and noncanonical sites determine NPT2A binding selectivity to NHERF1 PDZ1. PLoS ONE, 10 (6).

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Na<sup>+</sup>/H<sup>+</sup> Exchanger Regulatory Factor-1 (NHERF1) is a scaffolding protein containing 2 PDZ domains that coordinates the assembly and trafficking of transmembrane receptors and ion channels. Most target proteins harboring a C-terminus recognition motif bind more-or-less equivalently to the either PDZ domain, which contain identical core-binding motifs. However some substrates such as the type II sodium-dependent phosphate co-transporter (NPT2A), uniquely bind only one PDZ domain. We sought to define the structural determinants responsible for the specificity of interaction between NHERF1 PDZ domains and NPT2A. By performing all-atom/explicit-solvent molecular dynamics (MD) simulations in combination with biological mutagenesis, fluorescent polarization (FP) binding assays, and isothermal titration calorimetry (ITC), we found that in addition to canonical interactions of residues at 0 and -2 positions, Arg at the -1 position of NPT2A plays a critical role in association with Glu43 and His27 of PDZ1 that are absent in PDZ2. Experimentally introduced mutation in PDZ1 (Glu43Asp and His27Asn) decreased binding to NPT2A. Conversely, introduction of Asp183Glu and Asn167His mutations in PDZ2 promoted the formation of favorable interactions yielding micromolar K<inf>D</inf>s. The results describe novel determinants within both the PDZ domain and outside the canonical PDZ-recognition motif that are responsible for discrimination of NPT2A between two PDZ domains. The results challenge general paradigms for PDZ recognition and suggest new targets for drug development.


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Item Type: Article
Status: Published
CreatorsEmailPitt UsernameORCID
Mamonova, Ttbm7@pitt.eduTBM7
Zhang, Q
Khajeh, JA
Bu, Z
Bisello, Aalb138@pitt.eduALB138
Friedman, PApaf10@pitt.eduPAF10
ContributionContributors NameEmailPitt UsernameORCID
Date: 12 June 2015
Date Type: Publication
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Journal or Publication Title: PLoS ONE
Volume: 10
Number: 6
DOI or Unique Handle: 10.1371/journal.pone.0129554
Institution: University of Pittsburgh
Schools and Programs: School of Medicine > Pharmacology and Chemical Biology
School of Medicine > Structural Biology
Refereed: Yes
Date Deposited: 23 Aug 2016 14:19
Last Modified: 03 Apr 2021 09:55


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