BDB -BiopanningDataSet Details

Biopanning Data

BiopanningDataSet ID: 984

Peptides:

Sequences	Affinity Values
CRIGPITWVC	[10]

Sequences Features:

Motif:

Unique Sequence Number:

Experimental Method:

Phage display (common panning)

Sequencing Platform:

Sequencing Method:

Sanger sequencing

Rounds of Panning:

Reference:

PMID: 8662529

Target Name:

Erythropoietin receptor

Template Name:

Erythropoietin

Structure of Target-Template Complex:

1CN4

1EER

Structure of Target-Peptide Complex:

1EBA

1EBP

Library Name:

ON1203 phage display library (CX8C)

Affinity Measurement Method:

Binding assay

Affinity Measurement Description:

Concentrations required for 50 percent inhibition of tracer (125)I-labeled EPO binding (IC50 values, μM) were determined in a competition binding assay. Under the assay conditions, the IC50 was approximately equal to the affinity (Kd).

Brief Description:

The extracellular domain of the human EPO receptor (EPOR ECD) was expressed in Chinese hamster ovary cells as a fusion protein. Besides EPOR ECD, this fusion protein had a COOH-terminal signal sequence of the human placental alkaline phosphatase (HPAP) to aid in purification and immobilization. Between the ECD and HPAP domains, a protease (thrombin) cleavage site was inserted. The fussion protein was used as the target; target-phage complexes were cleaved from mAbl79 with bovine thrombin at room temperature for 10 minutes.

BiopanningDataSet Comments:

EPO was found to compete with the clone displaying CRIGPITWVC for binding to the immobilized EPOR. The peptide GGCRIGPITWVCGG was capable of competing with EPO binding to immobilized EPOR with an apparent affinity of approximately 10 uM. Higher affinity peptides were found from a secondary mutagenesis library containing peptides of increased length and were verified as full EPOR agonists in vitro and in vivo. The stuctures of EPO mimetic peptides complexed with EPOR have been solved (e.g. 1EBA, 1EBP).

Curator:

Curation Date:

0000-00-00

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