Stereochemical Determinants of C-terminal Specificity in PDZ Peptide-binding Domains

Amacher, J.F.; Cushing, Patrick R.; Bahl, Christopher D.; Beck, Tobias; Madden, Dean R.

doi:10.1074/jbc.m112.401588

Cited by 34 publications

(110 citation statements)

References 69 publications

(87 reference statements)

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“…One possible explanation is that the C-terminal carboxylate of the ligand positions differently towards the carboxylate-binding site of the PDZ domains and thereby attains subtle differences in H-bond patterns, which also has been suggested by X-ray crystal structures of PDZ domains in complex with C-terminal ligands 50 . These findings together with our present data suggest that the C-terminal interaction at the carboxylatebinding site could be part of a selectivity mechanism that allows the PDZ domains to discriminate between C-terminal peptide ligands 51 .…”

Section: Discussionsupporting

confidence: 55%

Probing backbone hydrogen bonding in PDZ/ligand interactions by protein amide-to-ester mutations

Pedersen

Anker

et al. 2014

Nat Commun

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PDZ domains are scaffolding modules in protein-protein interactions that mediate numerous physiological functions by interacting canonically with the C-terminus or non-canonically with an internal motif of protein ligands. A conserved carboxylate-binding site in the PDZ domain facilitates binding via backbone hydrogen bonds; however, little is known about the role of these hydrogen bonds due to experimental challenges with backbone mutations. Here we address this interaction by generating semisynthetic PDZ domains containing backbone amide-to-ester mutations and evaluating the importance of individual hydrogen bonds for ligand binding. We observe substantial and differential effects upon amide-to-ester mutation in PDZ2 of postsynaptic density protein 95 and other PDZ domains, suggesting that hydrogen bonding at the carboxylate-binding site contributes to both affinity and selectivity. In particular, the hydrogen-bonding pattern is surprisingly different between the non-canonical and canonical interaction. Our data provide a detailed understanding of the role of hydrogen bonds in protein-protein interactions.

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Section: Discussionsupporting

confidence: 55%

Probing backbone hydrogen bonding in PDZ/ligand interactions by protein amide-to-ester mutations

Pedersen

Anker

et al. 2014

Nat Commun

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“…CALP displayed strong preferences for I/L/V and S/T at position 0 (P 0 ) and position −2 (P −2 ), respectively, where P 0 is defined as the most C-terminal residue. These preferences are in excellent agreement with the assignment of CALP as a bona fide class I PDZ domain 31 , and with our previous studies 27,30,32 . At positions P −6 through P −9 , the wt amino acids can be replaced by any other, which is typical of the relatively non-specific interactions of N-terminal residues with the PDZ domain 14 .…”

supporting

confidence: 91%

Optimization of the process of inverted peptides (PIPEPLUS) to screen PDZ domain ligands

Seisel¹,

Rädisch²,

Gill³

et al. 2017

Bioorganic & Medicinal Chemistry Letters

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PDZ domains play crucial roles in cell signaling processes and are therefore attractive targets for the development of therapeutic inhibitors. In many cases, C-terminal peptides are the physiological binding partners of PDZ domains. To identify both native ligands and potential inhibitors we thus have screened arrays synthesized by the process of inverted peptides (PIPE), a variant of SPOT synthesis that generates peptides with free C-termini. Here, we present the development of a new functionalized cellulose membrane as solid support along with the optimized PIPEPLUS technology. Improved resolution and accuracy of the synthesis were shown with peptide arrays containing both natural and non-natural amino acids. These new screening possibilities will advance the development of active, selective and metabolically stable PDZ interactors.

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“…Given the lack of upstream binding motifs, we decided to explore the free-energy contributions of non-motif residues, using nested sets of chimeric peptides based on reference sequences with both high (iCAL36; K i = 23 μM) and low (CFTR; K i = 420 μM) affinity for the CALP domain (Amacher et al, 2013). Using fluorescence polarization (FP) competition experiments, we determined K i values for each of the chimeric peptides binding to CALP (Table 1).…”

Section: Resultsmentioning

confidence: 99%

Stereochemical Preferences Modulate Affinity and Selectivity among Five PDZ Domains that Bind CFTR: Comparative Structural and Sequence Analyses

et al. 2014

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Summary PDZ domain interactions are involved in signaling and trafficking pathways that coordinate crucial cellular processes. Alignment-based PDZ binding motifs identify the few most favorable residues at certain positions along the peptide backbone. However, sequences that bind the CAL (CFTR-Associated Ligand) PDZ domain reveal only a degenerate motif that overpredicts the true number of high affinity interactors. Here, we combine extended peptide-array motif analysis with biochemical techniques to show that non-motif ‘modulator’ residues influence CAL binding. The crystallographic structures of 13 new CAL:peptide complexes reveal defined, but accommodating stereochemical environments at non-motif positions, which are reflected in modulator preferences uncovered by multi-sequence substitutional arrays. These preferences facilitate the identification of new high-affinity CAL binding sequences and differentially affect CAL and NHERF PDZ binding. As a result, they also help determine the specificity of a PDZ domain network that regulates the trafficking of CFTR at the apical membrane.

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Stereochemical Determinants of C-terminal Specificity in PDZ Peptide-binding Domains

Cited by 34 publications

References 69 publications

Probing backbone hydrogen bonding in PDZ/ligand interactions by protein amide-to-ester mutations

Probing backbone hydrogen bonding in PDZ/ligand interactions by protein amide-to-ester mutations

Optimization of the process of inverted peptides (PIPEPLUS) to screen PDZ domain ligands

Stereochemical Preferences Modulate Affinity and Selectivity among Five PDZ Domains that Bind CFTR: Comparative Structural and Sequence Analyses

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