BDB -BiopanningDataSet Details

Biopanning Data

BiopanningDataSet ID: 3271

Peptides:

Sequences
CKKNSPTLC(14/18) CIGNSNTLC(4/18)

Sequences Features:

Motif:

Unique Sequence Number:

Experimental Method:

Phage display (subtractive panning)

Sequencing Platform:

Sequencing Method:

Sanger sequencing

Rounds of Panning:

Reference:

PMID: 28029647

Target Name:

G-protein coupled receptor 55

Template Name:

Lysophosphatidylinositol, LPI

Structure of Target-Template Complex:

Not determined.

Structure of Target-Peptide Complex:

Not determined.

Library Name:

Ph.D.-C7C phage display library (CX7C)

Affinity Measurement Method:

Affinity Measurement Description:

Brief Description:

Briefly, phages of approximately 1.0e10 plaque-forming units (PFU) were pre-incubated for 1 h with 5.0e5 wild-type HEK293 cells. The supernatant containing unbound phage particles was subsequently transferred to 35-mm Petri dishes to be incubated with 5 ×105 ev-HEK293 cells for an additional 2 h, to further eliminate phage particles with a specific binding. The ultimate pre-cleaned supernatant was incubated with GPR55-HEK293 cells for 2 h, and unbound phage particles were removed by washing with 50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 0.1% Tween-20. The extracellular bound phages were eluted with 100 mM glycine-HCl (pH 2.2) for 10 min, and neutralized with 1 M Tris-HCl (pH 9.0). An additional step of cryogenic lysis of GPR55-HEK293 cells allowed the recovery of the intracellular bound phages. The internalized phages were collected.

BiopanningDataSet Comments:

Sequencing of selected phagotopes dictated the primary structure for the synthetic FITC-labeled peptide P1 (CKKNSPTLC), which was analyzed for binding specificity to immunoprecipitated HA-GPR55, and to endogenously expressed the lysophosphatidylinositol receptor GPR55, using cells interfered or not for GPR55, as well as for co-localization imaging with HA-GPR55 at the membrane level. The peptide P1 stimulated GPR55 endocytosis and inhibited GPR55-dependent proliferation of EHEB and DeFew cells, two human B-lymphoblastoid cell lines. Our data support the potential therapeutic application of peptide ligands of GPR55 for targeting and inhibiting growth of neoplastic cells, which overexpress GPR55 and are dependent on GPR55 signaling for their proliferation.

Curator:

Bifang He

Curation Date:

2018-01-19

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