BDB -BiopanningDataSet Details

Biopanning Data

BiopanningDataSet ID: 3546

Peptides:

Sequences

Affinity Values

NVFTVSP(6)
NSLSISY(3)
SQQGKLN(2)
ELTPLTL(2)
QLAVAPS(2)
DRLSHTR(2)
QTLNVKP(2)
VVTPKTA(2)
TTQVLEA(2)
NLLDSLH(1)
WSLSELH(1)

[0.816657±0.001570, 12.09]
[0.371417±0.001570, NA]
[0.525270±0.002944, NA]
[0.182410±0.003243, NA]
[0.255427±0.001570, NA]
[0.252820±0.002944, NA]
[0.215853±0.001570, NA]
[0.220397±0.001570, NA]
[0.208373±0.000981, NA]
[0.208033±0.001570, NA]
[0.319490±0.002720, NA]

Sequences Features:

Motif:

Unique Sequence Number:

Experimental Method:

Phage display (common panning)

Sequencing Platform:

Sequencing Method:

Sanger sequencing

Rounds of Panning:

Reference:

PMID: 31805343

Target Name:

Fibroblast growth factor 9, FGF-9

Template Name:

FGFR-1, FGFR-2, FGFR-3 and FGFR-4

Structure of Target-Template Complex:

Not determined.

Structure of Target-Peptide Complex:

Not determined.

Library Name:

Ph.D.-7 phage display library (X7)

Affinity Measurement Method:

ELISA

Affinity Measurement Description:

The binding activity of the 11 positive phage clones to FGF9 was verified by ELISA assay. The absorbance was measured at 450 nm. The OD450 value (in the first column) was reproduced from Figure 1A and shown. In addition, to confirm the interaction between P4 peptide and FGF9, the surface plasmon resonance (SPR) on the immobilized FGF9 was applied to determine the affinity constants. The binding kinetics of the peptides to FGF9 was detected by OpenSPR™ (Nicoya Lifesciences, Waterloo, Canada). The Kd (μM) value was shown in the second column of the affinity values.

Brief Description:

A sterile polystyrene petri dish was coated with 10 μg/ml recombinant human FGF9 or PBS at 4 °C in 0.1 M NaHCO3 overnight, and blocked with bovine serum albumin (BSA) at 5 mg/ml in 0.1 M NaHCO3 for 2 h at room temperature. After the dish was washed five times (1 min each) with 0.05 % PBST (PBS containing 0.05 % Tween-20), the diluted original Ph.D.-7 library (2.0e11) was added and incubated for 2 h at room temperature. The unbound phages were removed through washing ten times (1 min each) with 0.05 % PBST prior to elution of the bound phages with continuous shaking in 1 ml of 0.1 M glycine-HCl for 10 min at room temperature and subsequent neutralization with 100 μl of 1 M Tris-HCl. The eluate was amplified and purified for the next round of screening. Two additional rounds of selection were conducted under more rigorous conditions. In short, dishes were coated with less amount of FGF9 (5 μg/ml and 2.5 μg/ml for the second and third round, respectively), incubated with the previous eluate for shorter time (1.5 h and 1 h for the second and third rounds, respectively), and washed with higher concentration of PBST for a longer time (0.1 % PBST for 10 × 2 min and 0.3 % PBST for 10 × 3 min for the second and third round, respectively). After three successive rounds of strict selection, the phage clones obtained by infection of E. coli ER2738, individual selection and amplification were exposed to Enzyme-linked immunosorbent assay (ELISA).

BiopanningDataSet Comments:

Sequence comparison showed that P4 (NVFTVSP) shared high homology with the conserved motif in the immunoglobulin-like (Ig-like) domain II∼III (D2-D3) linker of the FGF9 high-affinity receptor (FGFR3c). The interaction between P4 and FGF9 was confirmed by the surface plasmon resonance (SPR) assay. Functional analysis indicated that P4 counteracted FGF9-induced aggressive phenotype, including cell proliferation, migration, and invasion in vitro, as well as suppressed tumor growth in vivovia down-regulation of the MAPKs and Akt cascades. More importantly, we found that FGF9 served as an underlying mechanism of the chemoresistance in GC and BC cells, and P4 could increase the sensitivity to the chemical agent via antagonizing the suppression effects of FGF9 on cell apoptosis. Taken together, our study identified a novel binding peptide for FGF9, which may serve as a potential therapeutic agent for malignant tumors featured by abnormally up-regulation of FGF9.

Curator:

Chunchao Pu

Curation Date:

2020-12-29

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