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Article Dans Une Revue Biopolymers Année : 2009

T7 phage display as a method of peptide ligand discovery for PDZ domain proteins

Sudhir Sharma
  • Fonction : Auteur
Adnan Memic
  • Fonction : Auteur
Mark R. Spaller
  • Fonction : Auteur

Résumé

The use of bacteriophage T7 is presented as a peptide display platform to identify short binding sequences for PDZ domain proteins. Two different domains are examined, the 10th PDZ domain (PDZ10) of the multi-PDZ domain protein 1 (MUPP1) and the third PDZ domain (PDZ3) of postsynaptic density-95 (PSD-95) protein. Using the T7Select 415-1b construct, which displays 415 peptides per phage particle, a random heptapeptide and focused octapeptide libraries were constructed and subjected to iterative selection-enrichment cycles against surface-immobilized PDZ3 and PDZ10 proteins. The derived consensus sequences, together with those of high-frequency clones, were used as the basis for individual chemically synthesized peptides. Each peptide was subjected to isothermal titration calorimetry binding determinations against the corresponding PDZ domain under standard solution conditions. For MUPP1 PDZ10, binding analysis demonstrated that one of the heptapeptides, Ac-IGRISRV, displayed a two-fold improved affinity over the octapeptide derived from the carboxy terminus of the hc-Kit protein, which we had recently demonstrated as among the highest affinity ligands reported to date for that domain. In the case of PSD-95 PDZ3, peptides were found that possessed low-micromolar dissociation constants, as well as those that rediscovered the C-terminal sequence (KQTSV) of the protein CRIPT, a known natural binding protein of PDZ3. These successful examples of ligand discovery against two distinctly different PDZ domains demonstrate that the T7 phage platform could prove broadly applicable to the numerous other PDZ domains for which binding peptides are absent or of insufficient affinity.

Dates et versions

hal-01668582 , version 1 (20-12-2017)

Identifiants

Citer

Sudhir Sharma, Adnan Memic, Chamila N. Rupasinghe, Anne-Cécile E. Duc, Mark R. Spaller. T7 phage display as a method of peptide ligand discovery for PDZ domain proteins. Biopolymers, 2009, 92 (3), pp.183 - 193. ⟨10.1002/bip.21172⟩. ⟨hal-01668582⟩
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