BDB -BiopanningDataSet Details

Biopanning Data

BiopanningDataSet ID: 3111

Peptides:

Sequences
EWWWW

Sequences Features:

Motif:

Unique Sequence Number:

Experimental Method:

Phage display (common panning)

Sequencing Platform:

Sequencing Method:

Sanger Sequencing

Rounds of Panning:

Reference:

PMID: 28315327

Target Name:

Kelch-like ECH-associated protein 1

Template Name:

Nuclear factor erythroid 2-related factor 2

Structure of Target-Template Complex:

Not determined.

Structure of Target-Peptide Complex:

Not determined.

Library Name:

X4 T7 phage display library

Affinity Measurement Method:

Affinity Measurement Description:

Brief Description:

Biotinylated Avi-tagged Keap1 Kelch domain was immobilized to Dynabeads M280-streptavidin (Invitrogen) in PBS (045e29795, Wako) containing 0.5% BSA. After washing the beads using PBS containing 0.1% Tween 20 (PBST), the beads were incubated with the phage libraries for 1 h and were washed with PBST. The bound phages were eluted with 0.5% SDS and were incubated with E. coli BLT5615 cells (Merck Millipore) in log-phase growth for phage amplification. After bacteriolysis, the phages were recovered from the culture supernatant by centrifugation and PEG-precipitation according to manufacturer's T7 protocol. The recovered phages were suspended in PBS and were used for the next round of panning.

BiopanningDataSet Comments:

We identified a tetrapeptide, EWWW, showing moderate binding affinity, which inhibits the interaction between Nrf2 and Keap1. The tetrapeptide does not include an ETGE motif, which is a commonly found consensus sequence in known peptidic inhibitors. In addition to affinity parameters, IC50, KD, and thermodynamic parameters, the crystal structure of the complex was determined to elucidate the binding conformation. The binding interactions resemble those of known small-molecule inhibitors as opposed to those of substrates and peptidic inhibitors.

Curator:

Yumin

Curation Date:

2017-04-11

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